仙客来体细胞胚胎发生、发育及SERK基因在体细胞胚性转化过程的表达特性
|
摘要
本论文以仙客来(Cyclamen persicum Mill)已开花植株的新生叶片和叶柄为外植体,系统研究了仙客来体细胞胚的诱导条件并建立了完整的仙客来体细胞胚胎发生和发育体系;在此基础上,选择各发育期的体细胞胚为外植体,进一步建立了以球形胚为外植体的体细胞胚高频、高效诱导体系;同时以球形胚直接投放液体培养成功建立了胚性悬浮体系并筛选获得另一胚性丧失体系。以上述不同胚性培养体系为材料,对仙客来SERK基因(CpSERK)在体细胞胚性转化过程中的表达特性进行了初步的研究,获得的研究结果如下:
以仙客来“葡萄酒红火焰纹”(Cyclamen cv“Wine Red Flame”)的新生叶片为外植体,成功诱导、筛选出胚性愈伤组织。所选择的愈伤组织诱导培养基为1/2MS附加2,4-D(2-4mg.L~(-1))和2iP( 0.2-0.8mg.L~(-1)),胚性愈伤组织继代增殖培养基1/2MS+ 2 , 4-D2mg.L~(-1)+6-BA0.2mg.L~(-1)和1/2MS+2 , 4-D2mg.L~(-1) +2iP0.2mg.L~(-1)。初培养所获得的松散愈伤组织经过3-4次的继代培养后,根据颜色、松散程度、水渍化程度、细胞的形态特征等指标进一步分类筛选。通过胚性能力的检测,证明并获得了两种不同形态的胚性愈伤组织,分别为棕红色松软愈伤组织和松脆愈伤组织,胚性愈伤组织固体继代一年以上仍能保持其胚性能力。
将两种胚性愈伤组织进一步继代培养于不含激素的1/2MS培养基上,14d左右,体细胞胚开始在愈伤组织表面发生。松软愈伤组织的体细胞胚均以多发生的状态出现,一丛可达几十个甚至上百个。松脆愈伤组织的体细胞胚以单发生和多发生两种状态出现,即使多发生,体细胞胚发育的数目最多只有十几个。体细胞胚发生和发育具明显的不同步化,同一丛体胚中会出现球形胚、心形胚、鱼雷胚甚至成熟胚并存的现象,大多数体胚60-90d可发育成完整的小植株。连续7-8次继代培养的胚性愈伤体胚正常率为34.7%,随着继代次数的增加,胚性愈伤组织的胚性能力和正常体胚率明显下降,继代培养一年以上的胚性愈伤组织,其体胚正常率下降到7%以下。
对两种胚性愈伤组织和松散的非胚性愈伤组织进行了活体细胞的比较观察,结果表明,胚性和非胚性愈伤组织中均含有不同大小的细胞和细胞团。两种胚性愈伤组织细胞中均含有胚性细胞和非胚性细胞,但数量上存在明显的差异。松软胚性愈伤组织多为含有数十个胚性细胞的胚性细胞团,而松脆胚性愈伤组织中的胚性细胞团胚性细胞数目只有几个或十几个,且在随机选取的相同视野中胚性细胞团的数量前者明显多于后者。通过连续的活体细胞追踪观察发现,胚性细胞团起源于胚性母细胞。一个胚性母细胞通过持续分裂形成包含有几个或十几个胚性细胞的胚性细胞团,而包含有几十个胚性细胞的胚性细胞团来源于成团存在的几个胚性母细胞。
对体细胞胚胎发生和发育整个过程进行了细胞和组织学追踪,结合活体细胞和石蜡包埋切片技术显微观察结果表明,该培养体系的体细胞胚胎属于单细胞发生体系。体细胞胚的发生起源于胚性细胞团表面的单个胚性细胞,经历了二分体、四分体、八分体、多细胞原胚、球形胚、心形胚和鱼雷胚等时期,发育成完整植株。仙客来体细胞胚发育植株再生方式有两种:第一种为胚性细胞经多细胞原胚、球形胚、心形胚和鱼雷胚再生为小植株;第二种再生方式为球形胚不经过鱼雷胚而发育先形成球茎,然后是根或芽的形成,最后形成一完整植株。
利用组织化学切片技术对体细胞胚发生和发育过程中淀粉的代谢进行了动态跟踪观察。结果显示在体细胞胚胎整个发生发育期内出现了4个淀粉粒积累的高峰期,分别为胚性细胞期、球形胚期、早期鱼雷胚期和体细胞胚再生为小植株的球茎和不定芽的发育期,淀粉的积累和消减与体细胞胚的发生、发育密切相关。
为进一步研究高频率体胚体系,以再生胚胎为外植体,采用上述的胚性愈伤组织继代培养基1/2MS+2,4-D 2mg.L~(-1) +6-BA 0.2 mg.L~(-1)和1/2MS+2,4-D 2 mg.L~(-1) +2iP 0.2 mg.L~(-1)使其重新脱分化,初培养25-28d即成功诱导获得了胚性愈伤组织。在无激素的基本培养基上促使体细胞胚的发育,实验结果表明,除小球形胚外,以不同再生胚胎为外植体初培养愈伤组织诱导率均达95%以上,而且初代培养获得的愈伤组织就具备胚性能力。比较不同再生胚胎诱导获得的愈伤组织的胚性能力,表现为以大球形胚为外植体获得的胚性愈伤组织胚性能力最佳,以小球形胚和小无极球为外植体次之,无激素培养基培养30d每克愈伤组织获得体细胞胚数分别为734、423和342个,来源于单个胚胎外植体的体胚发生总数为118、80、69;同时,相同类型体细胞胚为外植体获得的愈伤组织的胚性能力个体间存在显著差异。
以大球形胚为外植体,研究继代培养次数对胚性愈伤组织胚性能力的影响。1/2MS基本培养基培养50d,根据对体细胞胚的发生率、每克愈伤组织体胚发生总数、来源单个再生胚胎外植体的体胚发生总数以及正常体胚率四个指标综合考察,继代次数对体胚发生率无显著影响,但对愈伤组织的胚性能力和体胚正常率影响显著。胚性能力最强的愈伤组织为继代一次的愈伤,体细胞胚的发生率、每克愈伤组织体胚发生总数、来源单个再生胚胎外植体的体胚发生总数以及正常体胚率分别为100%,3304/g.FW,1296/ g.FW,8.54%。
以大球形胚为外植体诱导获得的胚性愈伤组织为实验材料,以蔗糖浓度(0, 10, 30, 60, 90, 120g.L~(-1))和不同的ABA浓度(0, 0.2, 0.5, 1.0, 2.0mg.L~(-1))为指标,对体细胞胚的发育进行了进一步的条件优化。结果表明,1/2MS基本培养基中附加一定浓度的ABA可显著提高体细胞胚的正常率,但随着ABA浓度的增加,2.0mg.L~(-1)ABA明显抑制体细胞胚的发生,体胚发生数仅为292/ g.FW;培养基中附加0.5 mg.L~(-1)ABA,每克愈伤组织体细胞胚的发生数量为648,正常体胚率为12.65%;而未附加ABA的对照组,每克愈伤组织体细胞胚发生数为957,体胚的正常率仅有4.09%。体细胞胚的发生和发育均需要在基本培养中附加适量浓度的蔗糖,以体细胞胚的发生数量和正常体胚率两个指标综合考察,适宜范围为培养基附加30-60g.L~(-1)的蔗糖;其中60g.L~(-1)蔗糖胚性愈伤组织体胚的发生数量和正常体胚率均为最高,分别为1384/ g.FW,8.24%。蔗糖浓度低于10g.L~(-1)或高于120 g.L~(-1)均显著抑制体细胞胚的正常发育。
以再生球形胚为外植体采用直接液体投放的方式进行悬浮培养建立胚性悬浮体系,结果表明,球形胚初代液体培养即可脱分化形成大量的胚性母细胞,2-3次继代胚性母细胞分裂形成胚性细胞团。悬浮培养体系的最佳接种量为5%PCV浓度,最适继代时间为14d;此胚性悬浮体系连续继代培养一年以上仍具有胚性能力,且获得的胚性细胞在无激素的液体和固体培养基中均能发育;液体培养基中体胚的发育、植株再生方式只有第二种方式,而固体发育时两种发育、再生方式兼备。悬浮培养一年半(36代)筛选获得的米黄色细胞团块经两种不同方式验证,已完全丧失胚性和体胚发生的能力,但可以稳定增殖。
以上述获得的不同胚性体系为实验材料,研究仙客来体胚发生、发育相关基因的特异表达,利用Genbank搜索到的仙客来SERK基因cDNA序列,根据基因的保守序列利用Primer Premier 5.0软件设计了SERK基因的两对特异引物(SKF1/SKR1或SKF2/SKR2),克隆获得两个SERK基因cDNA片段,分别命名为CpSERKa和CpSERKb;测序结果显示cDNA长度分别为464bp,编码154aa和768bp,编码255aa。CpSERKa和CpSERKb在核苷酸水平上有89.2%的相似性,而在蛋白质水平上相似性可达90.9%,是两个不同的SERK基因。获得的基因序列在GenBank比对搜索,结果显示CpSERKa与该物种的CpSERK3同源性最高,核苷酸水平相似性为98.5%,而蛋白质水平相似性为98.1%,推测可能为该基因的部分序列;而CpSERKb为与CpSERK3同源性很高的另一新的家族成员。CpSERKa和CpSERKb与拟南芥、胡萝卜、马铃薯和蜜柑等物种具有较高的同源性,在分子系统进化树中与AtSERK3邻近,位于同一分支,可能属于同一SERK基因家族。
在不同的胚性体系中,CpSERKa和CpSERKb在具有胚性发生能力的组织和细胞中均有表达,而未获得或丧失胚性能力的组织中均不表达。对体细胞胚胎不同发育时期的表达结果表明,在多细胞原胚和球形胚期也有明显的表达; CpSERKa在鱼雷胚期仅表现出微弱的表达,而CpSERKb则没有表达产物产生。
A protocol for somatic embryogenesis and development has been carried out,using young leaves and petioles from flowering plants as explants in Cyclamen persicum Mill. Another rapid and high-frequency somatic embryogenesis system has also been successfully achieved from globular embryo-explants. And an embryogenic suspension culture is further established by inoculating globular embryos into liquid medium. Meanwhile expression pattern of Cyclamen somatic embryogenesis receptor-like kinase (CpSERK) is investigated during acquisition of embryogenic competence of somatic cell from liquid and solid culture systems.
The results show that embryogenic calli have been successfully induced and identified. The auxin 2,4-D (2-4mg.L~(-1)) and 2iP (0.2-0.8mg.L~(-1)) in half strength MS medium are important factors for the induction of somatic embryogenesis. The optimal sub-cultural media for embryogenic callus are supplemented with 1/2MS +2,4-D2mg.L~(-1) + 6-BA0.2mg.L~(-1) or 1/2MS+2,4-D2mg.L~(-1) + 2iP0.2mg.L~(-1). According to the color, friability, watery grades and morphological character et al., calli are classified and respectively proliferated after 3-4 sub-cultural periods on induction medium. Two types of embryogenic callus are verified and obtained, which one is white-brown and soft, and the other is similar to the former except for friability. Embryogenic competence of both calli can be retained for more than one year.
Somatic embryos start to appear on the surface of both embryogenic calli on the same medium without growth regulators for about 14 days. Somatic embryos occur in clusters from soft embryogenic callus, and several scores of embryoids are observed in one cluster. However, somatic embryos from friable embryogenic callus occur in clusters in more cases, or occasionally in individual way, with a few or more than ten embryoids in one cluster. Somatic embryos developed asynchronously in the same cluster, containing different developmental stage of embryoids, such as globular embryos, heart-shaped embryos, torpedo-shaped embryos and even mature embryos. Majority of somatic embryos can be regenerated into whole plantlets for 60-90 days. Normal embryo frequency is 34.7% from embryogenic calli after 7-8 periods of subculture. Embryogenic competence and normal embryo frequency have decreased remarkably as subculture prolongs, being only below 7% of normal embryo frequency for more than one year.
The observations of viable cells show that both embryogenic and non-embryogenic calli comprise different sizes and shapes of cells and cell aggregates. The number of embryogenic and non-embryogenic cells is obviously different in the two types of embryogenic calli. The soft embryogenic callus mostly comprises aggregates with more than several scores of embryogenic cells, but the friable embryogenic callus contains aggregates with several or more than ten embryogenic cells. And the number of embryogenic aggregates in the former callus is more than that of the latter callus in the same random scope. Embryogenic aggregates are derived from mother embryogenic cells by time-lapse track of viable cell. Embryogenic aggregates with several or more than ten cells are originated from an embryogenic mother cell by continuous division, but embryogenic aggregates with more than several scores of embryogenic cells could be from a few of mother embryogenic cells in groups.
Histological and cytological analysis during somatic embryogenesis of cyclamen show that somatic embryo is originated from the single embryogenic cell, being developed into a whole plantlet through different stages, such as two-cell proembryo, four-cell proembryo, eight-cell proembryo, multicellular proembryo, globular, heart-shaped, torpedo-shaped embryo. The two pathways have been observed during cyclamen somatic embryo development and plant regeneration. The first pathway is that a whole plantlet is regenerated through different developmental stages, such as embryogenic cell, multicellular proembryos, globular embryo, heart-shaped embryo and torpedo-shaped embryo. The second pathway is that a tuber is firstly formed from globular embryo without torpedo-shaped embryo, then roots or buds appear, finally a whole plantlet is regenerated.
Dynamic changes of starch metabolism are investigated during the development and regeneration of somatic embryos. Four accumulation peaks of starch grains respectively appear at the stage of embryogenic cells, globular embryos, the early torpedo-shaped embryos and the developing tuber of a plantlet, which may be closely related to somatic embryogenesis, development and plant regeneration for energy and
substance. Embryogenic calli are successfully induced after 25-28 days from regenerated somatic embryos-explants on the above sub-cultural media(1/2MS+2,4-D 2mg.L~(-1) +6-BA 0.2 mg.L~(-1) and 1/2MS+2,4-D 2 mg.L~(-1) +2ip 0.2 mg.L~(-1)). Primary callus percentages come to above 95%, except for small globular embryo-explants, using different types of somatic embryos regenerated from leaf-explants as explants. The initial calli have embryogenic capacity, which is optimal in embryogenic callus from large globular embryo-explants, secondly from small non-polar tuber-explants and small globular embryo-explants. The number of somatic embryos from the above three embryo-explants is respectively 734/g.FW, 423/g.FW and 342/g.FW and that of embryogenic calli is 118, 80 and 69 from single embryo-explants on free-growth-regulators medium for 30 days. Embryogenic competence shows significant difference among individuals from the same type of embryo-explants.
Effects of sub-cultural periods on embryogenic competence are studied as large globular embryo explants. The results show that sub-cultural periods have no effects on embryogenesis percentage, but significant effects on embryogenic competence and normal somatic embryo frequency on the-free-growth-regulator medium for 50 days. Calli for the first sub-cultural period show the best embryogenic competence, and the somatic embryogenesis percentage, the total number of somatic embryos of per gram callus, the total number of somatic embryos from single embryo-explants and the frequency of normal somatic embryos are respectively 100%, 3304/g.FW, 1296/ g.FW and 8.54%.
Effects of different concentration of sucrose (0, 10, 30, 60, 90, 120g.L~(-1)) and ABA(0, 0.2, 0.5, 1.0, 2.0mg.L~(-1)))have been evaluated on late development of somatic embryos. The frequency of normal somatic embryo has remarkably increased on the 1/2MS media supplemented with appropriate concentration of ABA. Higher concentration of ABA can clearly inhibit somatic embryogenesis, and the average number is only 292/g.FW on the medium with ABA 2.0 mg.L~(-1). ABA (0.5 mg.L~(-1)) is the most effective for development of somatic embryos, which the average number is 648/g.FW and normal embryo frequency is 12.65%. The normal embryo frequency (4.09%)on the control medium was low as compared to the frequency of 9.92–12.65% on experimental medium supplemented with ABA. Appropriate concentration supplement of sucrose (30-60 g.L~(-1)) is also helpful to somatic embryogenesis and development. The total number of somatic embryos and the normal embryo frequency are optimal on the medium with 60 g.L~(-1) sucrose, respectively 1384/ g.FW and 8.24%. Development of somatic embryos can be inhibited on the medium with less than 10g.L~(-1) or more than 120 g.L~(-1) of sucrose.
Embryogenic suspension culture has been successfully established, using regenerated globular embryos as starting materials inoculated directly in liquid medium. Globular embryo-explants are dedifferentiated into embryogenic mother cells after initial suspension culture, from which embryogenic cell aggregates are derived for 2-3 periods of subculture. The optimal inoculated density is 5% PCV (packed cell volume) and the optimal period of subculture is 14 days in the suspension system. Somatic embryos have developed on solid or liquid media without growth regulators, and embryogenic competence in suspension culture can be retained for more than a year. The developmental pathway of somatic embryos belongs to the second pathway in liquid culture, but both developmental pathways occur on solid medium. It is further verified that yellow cells aggregates have completely lost embryogenic competence with better proliferation capacity.
Specific expression of the related genes with somatic embryogenesis and development is investigated using the above different embryogenic systems as experimental materials. Two pairs of specific primers of SERK genes (SKF1/SKR1 或SKF2/SKR2)are designed using Primer Premier 5.0 software, according to the cDNA conserved sequences of cyclamen SERK genes which are BLAST searched against the GenBank database at the National Centre for Biotechnology Information (NCBI). Two putative SERK gene fragments are cloned and named CpSERKa and CpSERKb, respectively. The 464bp fragment is CpSERKa, coding 154 amino acids and the 768bp fragment is CpSERKb, coding 255 amino acids. The two CpSERK fragments are distinct but similar at both nucleotide level (89.2% similarity) and protein level (90.9% similarity) within the overlapped segment. The results from GenBank BLAST search and multiple sequence comparison reveal that CpSERKa is most similar to the SERK homologue CpSERK3 in the same species, with the similarities of 98.5% at nucleotide levels and 98.1% at protein levels. And CpSERKa is the partial sequence of CpSERK3, but CpSERKb is the other member of the same family. The results are confirmed by the phylogenic analysis in which CpSERKa, CpSERKb and CpSERK3 are tightly grouped together adjacent to AtSERK3. CpSERKa and CpSERKb are also highly homologous to SERK genes in Daucus carota, Solanum tuberosum and some other species.
Expression of both CpSERKa and CpSERKb has successfully detected in tissues and cells of embryogenic competence from different embryogenic systems, but not in non-embryogenic callus and cell aggregates of losing embryogenic competence. The results show that SERK genes are also expressed in multicellular proembryos and globular embryos. The slight expression of CpSERKa is detected, but CpSERKb is not expressed at the stage of torpedo-shaped embryos.
以仙客来“葡萄酒红火焰纹”(Cyclamen cv“Wine Red Flame”)的新生叶片为外植体,成功诱导、筛选出胚性愈伤组织。所选择的愈伤组织诱导培养基为1/2MS附加2,4-D(2-4mg.L~(-1))和2iP( 0.2-0.8mg.L~(-1)),胚性愈伤组织继代增殖培养基1/2MS+ 2 , 4-D2mg.L~(-1)+6-BA0.2mg.L~(-1)和1/2MS+2 , 4-D2mg.L~(-1) +2iP0.2mg.L~(-1)。初培养所获得的松散愈伤组织经过3-4次的继代培养后,根据颜色、松散程度、水渍化程度、细胞的形态特征等指标进一步分类筛选。通过胚性能力的检测,证明并获得了两种不同形态的胚性愈伤组织,分别为棕红色松软愈伤组织和松脆愈伤组织,胚性愈伤组织固体继代一年以上仍能保持其胚性能力。
将两种胚性愈伤组织进一步继代培养于不含激素的1/2MS培养基上,14d左右,体细胞胚开始在愈伤组织表面发生。松软愈伤组织的体细胞胚均以多发生的状态出现,一丛可达几十个甚至上百个。松脆愈伤组织的体细胞胚以单发生和多发生两种状态出现,即使多发生,体细胞胚发育的数目最多只有十几个。体细胞胚发生和发育具明显的不同步化,同一丛体胚中会出现球形胚、心形胚、鱼雷胚甚至成熟胚并存的现象,大多数体胚60-90d可发育成完整的小植株。连续7-8次继代培养的胚性愈伤体胚正常率为34.7%,随着继代次数的增加,胚性愈伤组织的胚性能力和正常体胚率明显下降,继代培养一年以上的胚性愈伤组织,其体胚正常率下降到7%以下。
对两种胚性愈伤组织和松散的非胚性愈伤组织进行了活体细胞的比较观察,结果表明,胚性和非胚性愈伤组织中均含有不同大小的细胞和细胞团。两种胚性愈伤组织细胞中均含有胚性细胞和非胚性细胞,但数量上存在明显的差异。松软胚性愈伤组织多为含有数十个胚性细胞的胚性细胞团,而松脆胚性愈伤组织中的胚性细胞团胚性细胞数目只有几个或十几个,且在随机选取的相同视野中胚性细胞团的数量前者明显多于后者。通过连续的活体细胞追踪观察发现,胚性细胞团起源于胚性母细胞。一个胚性母细胞通过持续分裂形成包含有几个或十几个胚性细胞的胚性细胞团,而包含有几十个胚性细胞的胚性细胞团来源于成团存在的几个胚性母细胞。
对体细胞胚胎发生和发育整个过程进行了细胞和组织学追踪,结合活体细胞和石蜡包埋切片技术显微观察结果表明,该培养体系的体细胞胚胎属于单细胞发生体系。体细胞胚的发生起源于胚性细胞团表面的单个胚性细胞,经历了二分体、四分体、八分体、多细胞原胚、球形胚、心形胚和鱼雷胚等时期,发育成完整植株。仙客来体细胞胚发育植株再生方式有两种:第一种为胚性细胞经多细胞原胚、球形胚、心形胚和鱼雷胚再生为小植株;第二种再生方式为球形胚不经过鱼雷胚而发育先形成球茎,然后是根或芽的形成,最后形成一完整植株。
利用组织化学切片技术对体细胞胚发生和发育过程中淀粉的代谢进行了动态跟踪观察。结果显示在体细胞胚胎整个发生发育期内出现了4个淀粉粒积累的高峰期,分别为胚性细胞期、球形胚期、早期鱼雷胚期和体细胞胚再生为小植株的球茎和不定芽的发育期,淀粉的积累和消减与体细胞胚的发生、发育密切相关。
为进一步研究高频率体胚体系,以再生胚胎为外植体,采用上述的胚性愈伤组织继代培养基1/2MS+2,4-D 2mg.L~(-1) +6-BA 0.2 mg.L~(-1)和1/2MS+2,4-D 2 mg.L~(-1) +2iP 0.2 mg.L~(-1)使其重新脱分化,初培养25-28d即成功诱导获得了胚性愈伤组织。在无激素的基本培养基上促使体细胞胚的发育,实验结果表明,除小球形胚外,以不同再生胚胎为外植体初培养愈伤组织诱导率均达95%以上,而且初代培养获得的愈伤组织就具备胚性能力。比较不同再生胚胎诱导获得的愈伤组织的胚性能力,表现为以大球形胚为外植体获得的胚性愈伤组织胚性能力最佳,以小球形胚和小无极球为外植体次之,无激素培养基培养30d每克愈伤组织获得体细胞胚数分别为734、423和342个,来源于单个胚胎外植体的体胚发生总数为118、80、69;同时,相同类型体细胞胚为外植体获得的愈伤组织的胚性能力个体间存在显著差异。
以大球形胚为外植体,研究继代培养次数对胚性愈伤组织胚性能力的影响。1/2MS基本培养基培养50d,根据对体细胞胚的发生率、每克愈伤组织体胚发生总数、来源单个再生胚胎外植体的体胚发生总数以及正常体胚率四个指标综合考察,继代次数对体胚发生率无显著影响,但对愈伤组织的胚性能力和体胚正常率影响显著。胚性能力最强的愈伤组织为继代一次的愈伤,体细胞胚的发生率、每克愈伤组织体胚发生总数、来源单个再生胚胎外植体的体胚发生总数以及正常体胚率分别为100%,3304/g.FW,1296/ g.FW,8.54%。
以大球形胚为外植体诱导获得的胚性愈伤组织为实验材料,以蔗糖浓度(0, 10, 30, 60, 90, 120g.L~(-1))和不同的ABA浓度(0, 0.2, 0.5, 1.0, 2.0mg.L~(-1))为指标,对体细胞胚的发育进行了进一步的条件优化。结果表明,1/2MS基本培养基中附加一定浓度的ABA可显著提高体细胞胚的正常率,但随着ABA浓度的增加,2.0mg.L~(-1)ABA明显抑制体细胞胚的发生,体胚发生数仅为292/ g.FW;培养基中附加0.5 mg.L~(-1)ABA,每克愈伤组织体细胞胚的发生数量为648,正常体胚率为12.65%;而未附加ABA的对照组,每克愈伤组织体细胞胚发生数为957,体胚的正常率仅有4.09%。体细胞胚的发生和发育均需要在基本培养中附加适量浓度的蔗糖,以体细胞胚的发生数量和正常体胚率两个指标综合考察,适宜范围为培养基附加30-60g.L~(-1)的蔗糖;其中60g.L~(-1)蔗糖胚性愈伤组织体胚的发生数量和正常体胚率均为最高,分别为1384/ g.FW,8.24%。蔗糖浓度低于10g.L~(-1)或高于120 g.L~(-1)均显著抑制体细胞胚的正常发育。
以再生球形胚为外植体采用直接液体投放的方式进行悬浮培养建立胚性悬浮体系,结果表明,球形胚初代液体培养即可脱分化形成大量的胚性母细胞,2-3次继代胚性母细胞分裂形成胚性细胞团。悬浮培养体系的最佳接种量为5%PCV浓度,最适继代时间为14d;此胚性悬浮体系连续继代培养一年以上仍具有胚性能力,且获得的胚性细胞在无激素的液体和固体培养基中均能发育;液体培养基中体胚的发育、植株再生方式只有第二种方式,而固体发育时两种发育、再生方式兼备。悬浮培养一年半(36代)筛选获得的米黄色细胞团块经两种不同方式验证,已完全丧失胚性和体胚发生的能力,但可以稳定增殖。
以上述获得的不同胚性体系为实验材料,研究仙客来体胚发生、发育相关基因的特异表达,利用Genbank搜索到的仙客来SERK基因cDNA序列,根据基因的保守序列利用Primer Premier 5.0软件设计了SERK基因的两对特异引物(SKF1/SKR1或SKF2/SKR2),克隆获得两个SERK基因cDNA片段,分别命名为CpSERKa和CpSERKb;测序结果显示cDNA长度分别为464bp,编码154aa和768bp,编码255aa。CpSERKa和CpSERKb在核苷酸水平上有89.2%的相似性,而在蛋白质水平上相似性可达90.9%,是两个不同的SERK基因。获得的基因序列在GenBank比对搜索,结果显示CpSERKa与该物种的CpSERK3同源性最高,核苷酸水平相似性为98.5%,而蛋白质水平相似性为98.1%,推测可能为该基因的部分序列;而CpSERKb为与CpSERK3同源性很高的另一新的家族成员。CpSERKa和CpSERKb与拟南芥、胡萝卜、马铃薯和蜜柑等物种具有较高的同源性,在分子系统进化树中与AtSERK3邻近,位于同一分支,可能属于同一SERK基因家族。
在不同的胚性体系中,CpSERKa和CpSERKb在具有胚性发生能力的组织和细胞中均有表达,而未获得或丧失胚性能力的组织中均不表达。对体细胞胚胎不同发育时期的表达结果表明,在多细胞原胚和球形胚期也有明显的表达; CpSERKa在鱼雷胚期仅表现出微弱的表达,而CpSERKb则没有表达产物产生。
A protocol for somatic embryogenesis and development has been carried out,using young leaves and petioles from flowering plants as explants in Cyclamen persicum Mill. Another rapid and high-frequency somatic embryogenesis system has also been successfully achieved from globular embryo-explants. And an embryogenic suspension culture is further established by inoculating globular embryos into liquid medium. Meanwhile expression pattern of Cyclamen somatic embryogenesis receptor-like kinase (CpSERK) is investigated during acquisition of embryogenic competence of somatic cell from liquid and solid culture systems.
The results show that embryogenic calli have been successfully induced and identified. The auxin 2,4-D (2-4mg.L~(-1)) and 2iP (0.2-0.8mg.L~(-1)) in half strength MS medium are important factors for the induction of somatic embryogenesis. The optimal sub-cultural media for embryogenic callus are supplemented with 1/2MS +2,4-D2mg.L~(-1) + 6-BA0.2mg.L~(-1) or 1/2MS+2,4-D2mg.L~(-1) + 2iP0.2mg.L~(-1). According to the color, friability, watery grades and morphological character et al., calli are classified and respectively proliferated after 3-4 sub-cultural periods on induction medium. Two types of embryogenic callus are verified and obtained, which one is white-brown and soft, and the other is similar to the former except for friability. Embryogenic competence of both calli can be retained for more than one year.
Somatic embryos start to appear on the surface of both embryogenic calli on the same medium without growth regulators for about 14 days. Somatic embryos occur in clusters from soft embryogenic callus, and several scores of embryoids are observed in one cluster. However, somatic embryos from friable embryogenic callus occur in clusters in more cases, or occasionally in individual way, with a few or more than ten embryoids in one cluster. Somatic embryos developed asynchronously in the same cluster, containing different developmental stage of embryoids, such as globular embryos, heart-shaped embryos, torpedo-shaped embryos and even mature embryos. Majority of somatic embryos can be regenerated into whole plantlets for 60-90 days. Normal embryo frequency is 34.7% from embryogenic calli after 7-8 periods of subculture. Embryogenic competence and normal embryo frequency have decreased remarkably as subculture prolongs, being only below 7% of normal embryo frequency for more than one year.
The observations of viable cells show that both embryogenic and non-embryogenic calli comprise different sizes and shapes of cells and cell aggregates. The number of embryogenic and non-embryogenic cells is obviously different in the two types of embryogenic calli. The soft embryogenic callus mostly comprises aggregates with more than several scores of embryogenic cells, but the friable embryogenic callus contains aggregates with several or more than ten embryogenic cells. And the number of embryogenic aggregates in the former callus is more than that of the latter callus in the same random scope. Embryogenic aggregates are derived from mother embryogenic cells by time-lapse track of viable cell. Embryogenic aggregates with several or more than ten cells are originated from an embryogenic mother cell by continuous division, but embryogenic aggregates with more than several scores of embryogenic cells could be from a few of mother embryogenic cells in groups.
Histological and cytological analysis during somatic embryogenesis of cyclamen show that somatic embryo is originated from the single embryogenic cell, being developed into a whole plantlet through different stages, such as two-cell proembryo, four-cell proembryo, eight-cell proembryo, multicellular proembryo, globular, heart-shaped, torpedo-shaped embryo. The two pathways have been observed during cyclamen somatic embryo development and plant regeneration. The first pathway is that a whole plantlet is regenerated through different developmental stages, such as embryogenic cell, multicellular proembryos, globular embryo, heart-shaped embryo and torpedo-shaped embryo. The second pathway is that a tuber is firstly formed from globular embryo without torpedo-shaped embryo, then roots or buds appear, finally a whole plantlet is regenerated.
Dynamic changes of starch metabolism are investigated during the development and regeneration of somatic embryos. Four accumulation peaks of starch grains respectively appear at the stage of embryogenic cells, globular embryos, the early torpedo-shaped embryos and the developing tuber of a plantlet, which may be closely related to somatic embryogenesis, development and plant regeneration for energy and
substance. Embryogenic calli are successfully induced after 25-28 days from regenerated somatic embryos-explants on the above sub-cultural media(1/2MS+2,4-D 2mg.L~(-1) +6-BA 0.2 mg.L~(-1) and 1/2MS+2,4-D 2 mg.L~(-1) +2ip 0.2 mg.L~(-1)). Primary callus percentages come to above 95%, except for small globular embryo-explants, using different types of somatic embryos regenerated from leaf-explants as explants. The initial calli have embryogenic capacity, which is optimal in embryogenic callus from large globular embryo-explants, secondly from small non-polar tuber-explants and small globular embryo-explants. The number of somatic embryos from the above three embryo-explants is respectively 734/g.FW, 423/g.FW and 342/g.FW and that of embryogenic calli is 118, 80 and 69 from single embryo-explants on free-growth-regulators medium for 30 days. Embryogenic competence shows significant difference among individuals from the same type of embryo-explants.
Effects of sub-cultural periods on embryogenic competence are studied as large globular embryo explants. The results show that sub-cultural periods have no effects on embryogenesis percentage, but significant effects on embryogenic competence and normal somatic embryo frequency on the-free-growth-regulator medium for 50 days. Calli for the first sub-cultural period show the best embryogenic competence, and the somatic embryogenesis percentage, the total number of somatic embryos of per gram callus, the total number of somatic embryos from single embryo-explants and the frequency of normal somatic embryos are respectively 100%, 3304/g.FW, 1296/ g.FW and 8.54%.
Effects of different concentration of sucrose (0, 10, 30, 60, 90, 120g.L~(-1)) and ABA(0, 0.2, 0.5, 1.0, 2.0mg.L~(-1)))have been evaluated on late development of somatic embryos. The frequency of normal somatic embryo has remarkably increased on the 1/2MS media supplemented with appropriate concentration of ABA. Higher concentration of ABA can clearly inhibit somatic embryogenesis, and the average number is only 292/g.FW on the medium with ABA 2.0 mg.L~(-1). ABA (0.5 mg.L~(-1)) is the most effective for development of somatic embryos, which the average number is 648/g.FW and normal embryo frequency is 12.65%. The normal embryo frequency (4.09%)on the control medium was low as compared to the frequency of 9.92–12.65% on experimental medium supplemented with ABA. Appropriate concentration supplement of sucrose (30-60 g.L~(-1)) is also helpful to somatic embryogenesis and development. The total number of somatic embryos and the normal embryo frequency are optimal on the medium with 60 g.L~(-1) sucrose, respectively 1384/ g.FW and 8.24%. Development of somatic embryos can be inhibited on the medium with less than 10g.L~(-1) or more than 120 g.L~(-1) of sucrose.
Embryogenic suspension culture has been successfully established, using regenerated globular embryos as starting materials inoculated directly in liquid medium. Globular embryo-explants are dedifferentiated into embryogenic mother cells after initial suspension culture, from which embryogenic cell aggregates are derived for 2-3 periods of subculture. The optimal inoculated density is 5% PCV (packed cell volume) and the optimal period of subculture is 14 days in the suspension system. Somatic embryos have developed on solid or liquid media without growth regulators, and embryogenic competence in suspension culture can be retained for more than a year. The developmental pathway of somatic embryos belongs to the second pathway in liquid culture, but both developmental pathways occur on solid medium. It is further verified that yellow cells aggregates have completely lost embryogenic competence with better proliferation capacity.
Specific expression of the related genes with somatic embryogenesis and development is investigated using the above different embryogenic systems as experimental materials. Two pairs of specific primers of SERK genes (SKF1/SKR1 或SKF2/SKR2)are designed using Primer Premier 5.0 software, according to the cDNA conserved sequences of cyclamen SERK genes which are BLAST searched against the GenBank database at the National Centre for Biotechnology Information (NCBI). Two putative SERK gene fragments are cloned and named CpSERKa and CpSERKb, respectively. The 464bp fragment is CpSERKa, coding 154 amino acids and the 768bp fragment is CpSERKb, coding 255 amino acids. The two CpSERK fragments are distinct but similar at both nucleotide level (89.2% similarity) and protein level (90.9% similarity) within the overlapped segment. The results from GenBank BLAST search and multiple sequence comparison reveal that CpSERKa is most similar to the SERK homologue CpSERK3 in the same species, with the similarities of 98.5% at nucleotide levels and 98.1% at protein levels. And CpSERKa is the partial sequence of CpSERK3, but CpSERKb is the other member of the same family. The results are confirmed by the phylogenic analysis in which CpSERKa, CpSERKb and CpSERK3 are tightly grouped together adjacent to AtSERK3. CpSERKa and CpSERKb are also highly homologous to SERK genes in Daucus carota, Solanum tuberosum and some other species.
Expression of both CpSERKa and CpSERKb has successfully detected in tissues and cells of embryogenic competence from different embryogenic systems, but not in non-embryogenic callus and cell aggregates of losing embryogenic competence. The results show that SERK genes are also expressed in multicellular proembryos and globular embryos. The slight expression of CpSERKa is detected, but CpSERKb is not expressed at the stage of torpedo-shaped embryos.
引文
卞福花.仙客来体细胞胚胎的发生、变异及其蛋白质组学研究:[博士学位论文].北京:北京林业大学,2008
曹有龙,贾勇炯,陈放,罗青,曲琳.枸杞花药愈伤组织悬浮培养条件下胚状体发生与植物株再生.云南植物研究,1999,21(3):346-350
车美建,赖钟雄,赖呈纯,等.荔枝体细胞胚胎发生早期的3种内源激素含量变化.热带作物学报,2005,26(2):55-61
陈以峰,周燮,汤日圣,张金渝,梅传生.水稻体细胞培养中胚性细胞出现与IAA的关系.植物学报,1998,40(5):474-477
陈健辉,方璟.玉米种子萌发过程幼叶细胞中淀粉粒的积累观察.广西植物,2003,23(5) :440-444
程玉兰,黄美娟,刁丰秋等.蔗糖调控培养对胡萝卜体细胞胚内源ABA水平的效应.植物学报,1999 ,41(7):761-765
崔德才和徐培文.植物组织培养与工厂化育苗.北京:化学工业出版社,2004
崔凯荣,戴若兰.植物体细胞发生的分子生物学.北京:科学出版社,2000,50-171
崔凯荣,陈克明,王晓哲,等.植物体细胞胚发生研究的某些现状.植物学通报,1993,10(3):14-20
崔凯荣,刑更生,周功克,等.植物激素对体细胞胚发生的诱导和调节.遗传,2000,22(5):349-354
邓向阳,卫志明.幼胚长度、2 ,4-D浓度、光强度等对花生体细胞胚发生的影响及高效再生系统的建立.植物生理学报,2000,26 (6):525-531
范昌发,郭骁才,贾敬芬,牛天堂.植物细胞中淀粉代谢与离体形态发生途径决定的关系.华北农学报,2000,15:52-57
范现丽,王高,申晓辉.蓝百合体细胞胚胎发生及植株再生的研究.见:张启翔,编.中国观赏园艺研究进展2008——中国园艺学会观赏园艺专业委员会2008年学术年会论文集.中国黑龙江哈尔滨:中国园艺学会观赏园艺专业委员会、国家花卉工程技术研究中心, 2008. 124-127
高述民. ABA和PEG对胡萝卜体细胞胚诱导和调控的影响.西北农林科技大学学报,2001,29(2):13-16
宫相忠,黄健.苜蓿组织培养体细胞胚发生的细胞学观察.青岛海洋大学学报,1997, 24(4):504-508
顾玉红,高述民,郭惠红,李凤兰.文冠果的体细胞胚胎发生.植物生理学通讯,2004 ,40(3):311-313
韩碧文,刘淑兰.植物离体体细胞胚的发生.植物生理学通讯,1988,(1):9-15
韩晓玲,王冰雪,林雪,贾敬芬.小冠花高效体细胞胚胎发生与植株再生的研究.西北大学学报(自然科学版),2006,36(3):420-423
何丽君,于卓.濒危植物四合木(Tetraena mongolica Maxim)的离体培养组织学观察及再生植株的研究.内蒙古农业大学学报,2003,24(2):27-32
何丽君,齐冰杰.四合木体细胞胚胎发生中细胞组织学的淀粉积累动态的研究.内蒙古草业),2002,14(3):39-41
侯喜林.仙客来实生黄化叶柄培养的研究.园艺学报,1991,18(1): 81-86
胡博然,徐文彪,马峰旺.枸杞胚细胞悬浮培养系统建立的研究.西北农林科技大学学报(自然科学版),2003, 31(1):97-100
胡忠,徐艳,高欢欢,关秀琼.影响宁夏枸杞愈伤组织体细胞胚发生的因素.汕头大学学报(自然科学版),2006,21( 4): 57-63
胡适宜.同时显示胚中贮藏的淀粉、蛋白、脂类的制片方法植物学通报. 1994,11(4):49-51
胡适宜.被子植物胚胎学.北京:高等教育出版社,1982. 237-247
[39]黄学林,李筱菊,傅家瑞,劳彩玲. Thidiazuron对苜蓿愈伤组织的乙烯生成及其体细胞胚胎发生的影响.植物生理学报,1994,20:367-372
黄丹枫,刘佩瑛.魔芋再生植株形态发生途径的细胞组织学观察.上海农学院学报,1994,12(1):25-30
黄美娟,黄绍兴,朱微,等.采用调控培养方法提高胡萝卜体细胞胚活力的研究.科学通报,1993,38(6):550-553
赖钟雄,陈春玲.龙眼体细胞胚胎发生过程中的内源激素变化.热带作物学报,2002, 23(2): 41-47
赖钟雄,陈振光.龙眼胚性细胞悬浮培养及其体胚发生再生植株.应用与环境生物学报,2002,8(5): 485-491
赖钟雄,黄浅,林秀莲等.荔枝胚性悬浮细胞系的快速建立及其体胚植株的再生.农业生物技术科学,2007,23(1):28-32
李成浩,刘宝光,王伟达,刘立琨,Yu Chang-Yeon.温度对积棋次生胚发生和植株再生的影响.植物生理学通讯,2007,43(3): 453-456
李会宁,闫锋.仙客来叶片的组织培养体细胞胚诱导研究.汉中师范学院学报,2001,19(2): 81-83
李丽,万勇善,刘风珍. 2,4-D浓度对花生体细胞胚发生的影响.生物技术,2005,15(3):77-79
李杉,邢更妹,崔凯荣,王亚馥.植物体细胞胚发生中ATP酶活性时空分布动态与内源激素的变化.植物学通报,2001,18 (3):308-317
李天宪,赵林,冯锋,等.人巨细胞病毒小鼠模型的建立.微生物学报,1996,36(4):292-294
李学红,李冬玲,张举仁,等.玉米芽尖培养中的高频率体细胞胚胎发生与植株再生(简报). 植物生理学通讯,2000,36(5):430-433
李中奎,刘成运,刘文平.水稻愈伤组织内部胚性细胞的形成及发育.植物研究,2001,21(1):62-64
林荣双,王庆华,梁丽琨,郑秋生,肖显华.花生体细胞胚发生发育过程中淀粉代谢的动态变化.烟台大学学报(自然科学与工程版),2000,13(3):226-229
林荣双,王庆华,梁丽琨,肖显华. TDZ诱导花生幼叶的不定芽和体细胞胚发生的组织学观察.植物研究,2003,23(2):169-172
刘福平,陈移亮.细胞分裂素对蝴蝶兰胚性愈伤组织诱导的影响.热带作物学报,2008,29(1):42-46
刘华英,萧浪涛,鲁旭东,吴顺.柑橘体细胞胚发生的组织细胞学研究.果树学报,2004,21(4):311-314
刘华英,萧浪涛,鲁旭东,吴顺.柑橘体细胞胚发生的组织细胞学研究.果树学报,2004,21(4):311-314
刘艳芝,韦正乙,谭化,高明.花生体细胞胚发生及植株再生.吉林农业科学,2008, 33(4):7-10
刘一鸣,刁丰秋,张雷,等.胡萝卜LEA基因家族新成员DcLEA1的克隆及其结构与表达特征分析.自然科学进展,2004,8(8):882-891
牛德水,邵启全,张敬等.枸杞悬浮培养条件下的胚状体发生.遗传,1990,6:7-9
潘建伟,王卫军,潘伟槐,朱睦元.大麦花药培养及其胚性悬浮细胞系的高频快速建立.浙江大学学报(理学版),2002,29(4):454-458
潘炽.植物组织培养.广州:广东高等教育出版社,2000. 20-59
乔琦,肖娅萍.防风体细胞胚发生发育中的淀粉体和多糖动态.西北植物学报,2007,27(2):0388-0391
秦凡,周吉源.不同植物生长调节剂对蝴蝶兰快速繁殖的影响.武汉植物研究,2003,21(5):452-456
曲桂芹,张贤泽,霍俊伟.影响大豆体细胞胚诱导因素的研究.植物研究,2001,21(2):210-214
曲复宁,由翠荣,康黎芳,等.利用仙客来种苗组培快繁建立单株无性系的研究.西北农林科技大学学报,2003,31(3):81-86
曲复宁,由翠荣,龚雪芹,王云山,康黎芳.仙客来(Cyclamen persicum Mill)无性繁殖体系建立中不定芽分化能力的数量性分析.北方园艺,2001,(5): 62-63
曲复宁,由翠荣,龚雪芹,等.仙客来组织培养中再生器官类型及增殖稳定性比较.植物学通报,2004,21(5):559-564
盛德策,李凤兰,刘忠华.,植物体细胞胚发生的细胞生物学研究进展.,西北植物学报,2008,28(1):0204-0215
施小龙,邢更妹,汪丽虹,王亚馥.植物钙信号系统与体细胞胚发生.生命科学,2002,14(5):302-304
苏浴源,吴永祯,曲占礼,等.仙客来幼芽组织培养的研究.华北农学报,2002,17(增刊):203-205
Thomas T D.糖、GA3及ABA对印度娃儿藤( Tylophora indica ( Burm. f.) Merrill)体细胞胚发生的影响.生物工程学报, 2006, 22(3): 465-471
王波,王鹤,谭巍,等.仙客来组织培养及植株再生.北方园艺,2007,1:166-167
王杰,刘国锋,包满珠,黄莉.麝香百合胚性愈伤组织状态的调整与植株再生.园艺学报,2008,35(12:1795-1802
王康,朱慧森,董宽虎.植物LEA蛋白及其基因家族成员PM16研究进展.草业科学,2008,25(2):97-102
王丽,鲍晓明,黄百渠,郝水.香雪兰外植体形态学极性决定的体细胞胚胎发生.植物学报, 1998,40(2):138-143
王丽,段晓刚,郝水.香雪兰的直接与间接体细胞胚胎发生.实验生物学报,1999,32(2):175-182
汪丽虹,王星,崔凯荣,焦成瑾,王亚馥.石刁柏及党参体细胞胚发生中的淀粉代谢动态. 植物学通报,1996,13:41-45
王清连,王敏,师海荣.植物激素对棉花体细胞胚胎发生的诱导及调节作用.生物技术通讯,2004,15(6):577-579
王仑山,杨汉民,王亚馥,康文隽.伊贝母组织培养中体细胞胚的形成及细胞组织学观察.西北植物学报,1989,9(2):76-81
王亚馥,崔凯荣,陈克明,高清祥,张德华,焦成瑾.小麦组织培养中体细胞胚胎发生的细胞胚胎学及淀粉消长动态的研究.实验生物学报,1993,26(3):259-267
王义,赵文君,孙春玉,杨晶,蒋世翠,张美萍. 2,4-D、BA对人参体细胞胚胎发生过程的影响研究.中国生物工程杂志,2008,28(10):106-112
王义,赵文君,杨忠,孙春玉,张美萍.人参体细胞胚胎发生及植株再生研究.药物生物技术,2008,15(4):278-28
马和平,李毅,邸利,马彦军,魏继辉.枸杞体细胞胚胎发生及植株再生的研究.甘肃农业大学学报,2005,40(1):26-30
马义德,邢更生,戴若兰,等.生物组织切片图像的计算机三维重建技术现状.生物技术通讯,2000,11(4):303-308
魏岳荣.香蕉(Musa spp.)胚性细胞悬浮培养及其超低温保存和植株再生的研究:[博士学位论文].广州:中山大学生命科学学院,2005
魏岳荣,黄学林,李佳等.贡蕉胚性细胞悬浮系的建立和植株再生.生物工程学报,2005,21(1):58-65
吴春霞.植物细胞悬浮培养的影响因素.安徽农业科学,2009,37(1):36-38
夏光敏,陈惠敏.植物生长调节物质对石防风体细胞胚发生与发育的影响.植物学报,1993,35(8):644-648
夏光敏,李忠谊,于家驹,等.峨参原生质的体细胞胚同步化发生及植株再生.实验生物学报,1991,24(4):395-401
肖显华,林荣双,王庆华,王顺珍. 2,4-D诱导的花生体细胞胚发生的组织学研究.植物学通报,1999,16(6):691-695
肖望,黄学林.贡蕉胚性细胞悬浮培养过程中重要生长参数的研究.广东教育学院学报,2005,25(3):77-79
邢更生,崔凯荣,戴若兰,李杉,山仑.小麦胚性细胞发生中淀粉代谢动态的量化研究.兰州大学学报(自然科学版),1998,34:128-132
邢更生,邢更妹,崔凯荣,王亚馥.枸杞胚性细胞分化的超微结构研究.电子显微学报,1999,18(4):395-400
刑更生,崔凯荣,山仑,等.植物体细胞胚发生的分子基础.遗传,1999,21(1):30-34
邢登辉,赵云云,黄承芳.皇冠草体细胞胚胎发生及其体胚发生过程中内源激素的变化.生
物工程学报,1999,15(1): 98-101
邢登辉,吴琴生,刘大钧.禾谷类作物细胞培养.生物学通报,1994,29(7): 1-3
徐林林,芦笛,陆拢等.水稻悬浮细胞系的建立及培养条件对生物产量的影响.植物生理学通讯,2006,42(4):612-616
徐元红,朱四易.伊贝母与平贝母体细胞胚诱导条件的比较.陕西师范大学学报,1998,26(2):119-120
颜昌敬.农作物组织培养.上海:上海科学技术出版社,1991. 283-299
杨汉民,高清祥.枸杞体细胞胚的诱导与形态发.兰州大学学报(自然科学版),1992,28(1):87-89
杨会钗,李彦群.改良的苏木精染色液配制方法探讨.中华中西医杂志,2003,4(16):
叶克难,李文雷,徐增富,黄俊潮,李宝健.苜蓿体细胞胚胎发生过程中DNA、RNA和蛋白质的合成动态.实验生物学报,1992,25(4):403-411
尹文兵,李丽娟,黄勤妮,等.胡萝卜愈伤组织的诱导及细胞悬浮培养研究.山西师范大学学报,2004,18(2):71-76
俞长河,陈振光.荔枝胚性悬浮培养物的建立、保持和优化原生质体分离的研究.热带作物学报,1998,19(3):16-20
袁澍,贾勇炯,林宏辉.诱导植物体细胞胚发生的几个生理因素.植物生理学通讯,2003,39(5):508-512
张宝红,李秀兰. TDZ (thidiazuron)在棉花组织培养中的效应.作物学报,1995,21:253-258
张栋,陈季楚. ABA、NAA诱导水稻胚性愈伤组织的研究.实验生物学报,1995,28(3):329-334
张健,吕柳新,黄春梅等.柑桔体细胞胚的发生及其发育过程中淀粉粒动态研究.洛阳师范学院学报,2005,5:104-106
张树录,郑国錩,聂秀菀.糜子离体体细胞胚胎发生的组织学研究.西北植物学报,1992, 12(1):17-21
张树录.禾本科植物组织培养中的体细胞胚胎发生.植物生理学通讯,1985,(6):15-20
张新英,黄亚斌,尹光初,周思君.离体培养下大豆体细胞胚胎发生的组织学研究.植物学报,1992,34(3):214-218
赵桂兰,刘艳芝,尹爱平等.大豆花药培养中体细胞胚萌发的研究.科学通报,1998,43(14):1512-1516
周春江,李卫东,葛会渡.草莓悬浮细胞系的建立及某些影响因素.植物生理学通讯,2002,38(1):22-24
周俊彦.植物体细胞在组织培养中产生的胚状体.植物生理学报,1981,7(4):389-396
周连霞,马锋旺,陈登文,秦静远,程瑞红. TDZ对仙客来不同外植体再生的影响.西北农林科技大学学报(自然科学版),2006,34(4):39-42
周燕,高述民,李凤兰.胡萝卜体细胞胚胎发生中的细胞组织化学和蛋白质组成变化.植物生理学通讯,2004,40:181-183
庄伟建,张书标,刘思衡.花生体细胞胚的诱导及其植株再生.热带亚热带植物学报,1999 7(2):153-158
郑泳,王君晖,黄纯农.通过花药直接液体培养建立大麦胚性悬浮细胞系.杭州大学学报(自然科学版),1996,23(2):202-203
朱至清,王敬驹,孙敬三.在无激素的培养基上水稻和小麦花粉胚的发育.植物学报,1976,18:239-244
Albertini E, Marconi G, Reale L, Barcaccia G, Porceddu A, Ferranti F, Falcinelli M. SERK and APOSTART, candidate genes for apomixis in poa pratensis. Plant Physiol, 2005, 138(4): 2185-2199
Ammirato PV. Techniques for propagation and breeding. In: Evans DA, Sharp WR, Ammirato PV (eds). Handbook of Plant Cell Culture.Vol I. New York: Macmillan Pub, 1983. 82-84
Anil VS, Rao KS. Calcium mediated signaling during sandal wood somatic embryogenesis. Role for exogenous calcium as second messenger. Plant Physiol, 2000, 123: 1301-1311
Arnold S, Sabara I, Bozhkov P, et al. Developmental pathways of somatic embryogenesis. Plant Cell, Tissue and Organ Culture, 2002, 69: 233-249
Arnholdt-Schmit B. Physiological aspects of genome variability in tissue culture. II. Growth phase-dependent quantitative variability of repetitive BstN I fragments of primary cultures of Daucus carota L. Theor. Appl. Genet, 1995, 91:816-823
Baker CM, Wetzstein H. Influence of auxin type and concertration on peanut somatic embryogenesis. Plant Cell, tissue and Organ Culture, 1994, 36: 361-368
Baker CM and Wetzstein HY. Repetitive somatic embryogenesis in peanut cotyledon cultures by continual exposure to 2, 4-D. Plant Cell, tissue and Organ Culture, 1995, 40: 249-254
Balestrazzi A, Bernacchia G, Pitto L, et al. Spatial expression of DNA topoisomeraseI genes during cell proliferation in Daucus carota. Eur J Histochem, 2001, 45: 31-38
Baudino S, Hansen S, Brettschneider R, et al. Molecular characterization of two novel maize LRR receptor-like kinases, which belong to the SERK gene family. Planta, 2001, 213: 1-10
Beyer EM. Mechanism of action of ethylene:biological activity of deuterated ethylene and evidence against isotopic exchange and cis-transisomerisation. Plant Physiol, 1979, 63: 169-173
Bolandi AR, Branchard M, Alibert G, Gentzbittel L, Berville A and Sarrafi A. Combining ability analysis of somatic embryogenesis from epidermis layers in sunflower. Theor. Appl. Genet, 2000, 100: 621-624
Boxtel JV and Berthouly M. High frequency somatic embryogenesis from coffee leaves. Plant Cell, Tissue and Organ Culture, 1996, 44: 7-17
Cangahuala-Inocente GC, Steiner N, Santos M, Guerra MP. Morphohistological analysis and histochemistry of Feijoa sellowiana somatic embryogenesis. Protoplasma, 2004, 224: 33-40
Charrière F, SottaB, Miginiac E, Hahne G. Induction of adventitious shoots or somatic embryos on in vitro cultured zygotic embryos of Heli anthus annuus: Variation of endogenous hormone levels. Plant Physiol Biochem, 1999, 37: 751-757.
Che P, Gingerich DJ, Lall S, Howel SH. Global and Hormone-Induced Gene Expression Changes during Shoot Development in Arabidopsis. Plant Cell, 2002, 14(11): 2771-2785.
Cheliak WM, Klimaszewska K. Genetic variation in somatic embryogenic response in open-pollinated families of black spruce. Theor Appl Genet, 1991, 82: 185-190
Chengalrayan K, Mhaske VB, Hazra S. High frequency of conversion of abnormal peanut somatic embryo embryogenesis. Plant cell Rep, 1997, 16: 783-786
Choi YE, Jeong JH. Dormancy induction of somatic embryos of Siberian ginseng by high sucrose concentrations enhances the conservations of hydrated artificial seeds and dehydration resistance. Plant Cell Rep, 2002, 20 : 1112-1116
Choi YE, Kim JW, Soh WY. Somatic embryogenesis and plant regeneration from suspensioncultures of Acanthopanax koreanum Nakai. Plant Cell Reports, 1997, 17: 84-88
Conner AJ, Falloon PG. Osmotic versus nutritional effects when rooting in vitro asparagus minicrown on high sucrose media. Plant Sci, 1993, 89: 101-106
Corredoira E, Valladares S, Vieitez AM. Morphohistological analysis of the origin and development of somatic embryos from leaves of mature Quercus robur. In Vitro Cellular & Developmental Biology Plant, 2006, 42 (6): 525-533
Cuming AC, Lane BG. Protein synthesis in imbibing wheat embryos. Eur J Biochem, 1979, 99: 217-224
Daveletova S, Meszaros T, Miskolczi P, et al. Auxin and heat shock activation of a novel member of the calmodulin like domain protein kinase gene family in cultured alfalfa cells. J Exp Bot, 2001, 52: 215-221
Dharmasiri N, Dharmasiri S, Estelle M. The F-box protein TIR1 is an auxin receptor. Nature, 2005a, 435 (7041): 441-445.
Dharmasiri, N, Dharmasiri S, Weijers D, et al. Plant development is regulated by a family of auxin receptor F box proteins. Dev Cell, 2005b, 9: 109-119
Dillen W, Dijkstra I, Oud J. Shoot regeneration in long-term callus cultures derived from mature flowering plants of Cyclamen persicum Mill. Plant Cell Reports, 1996, 15: 545-548
Dohm A, Schwenkel HG, Grunewaldt J. Histological Analysis of the in-Vitro Regeneration in Cyclamen-Persicum Mill. Gartenbauwissenschaft, 1991, 56: 58-65
Dong JZ, Dunstan DI. Characterization of three heat-shock-protein genes and their developmental regulation during somatic embryogenesis in white spruce [picea glauca(moench) voss]. Planta, 1996, 200: 85-91
Dong J, Dunstan DI. Expression of abundant mRNAs during somatic embryogenesis of white spruce. Planta, 2000, 199: 459-466
Dong JZ, Perras MR, Abrams SR, Dunstan DI. Induced gene expression following ABA uptake in embryogenic suspension cultures of picea glauca.Plant Physiol.Plant Physiol Biochem, 1996, 34: 579-587
Dougall DK, Verma DC. Growth and embryo formation in wild carrot suspension culture with ammoniumion as a sole nitrogen source. In Vitro, 1978, 14: 180-182.
Du L, Zhou S, Bao MZ. Effect of plant growth regulators on direct somatic embryogenesis incamphor tree ( Cinnamomum camphora L.) from immature zygotic embryos and embryogenic calli induction. For. Stud. China, 2007, 9(4): 267-271
Dusits D, Bogre L, Gyorgye Y. Molecular and cellular approaches to the plant embryo development from somatic cells in vitro. J Cell Sci, 1991, 99(3): 473-482
Ellen WH, Weining T, Karl WN, et al. Expression and maintenance of embryogenic potential is enhanced through constitutive expression of AGAMOUS-Like 15. Plant Physiol, 2003, 133(2): 653-663.
Feher A, Pasternak TP, Dudits D. Transition of somatic plant cell to an embryogenic state. Plant Cell, Tissue and Organ Culture, 2003, 74: 201-228
Fireozabady E, DeBoer D L. Plant regeneration via somatic embryogenesis in many cultivars of cotton (Gossypium hirsutum L.). In Vitro Cell Dev. Biol, 1993, 29: 166-173
Fowler M R, Ong LM, Russinova E, et al. Early changes in gene expression during direct somatic embryogenesis in alfalf are veiled by RAP-PCR. J Exp Bot, 1998, 49: 249-253
Furukawa K, Kakihara F, Kato M. Somatic embryos produced from aseptic seedlings of wild Cyclamen species. Journal of Society of High Technology in Agriculture, 2002, 14(2): 71-80
Furukawa K, Kakihara F, Kato M. Somatic embryogenesis in seven wild Cyclamen species. Journal of Society of High Technology in Agriculture, 2001, 13(4): 270-278
Fuminori K, Shigeo T. Regulation by Vrn-1/Fr-1chromosomal intervals of CBF-mediated Cor/Leagene expression and freezing tolerance in common wheat. Journal of Experimental Botany, 2005, (3): 887-895
Fischer-Igisics C, Sundgerg B, Neuhaus G, Jones AM. Auxin distribution and transport during embryonic pattern formation in wheat. Plant J, 2001, 26(2): 115-129
Filonova L, Bozhkov P, von Arnold S. Developmental pathway of somatic embryogenesis in Picea abies as revealed by time-lapse tracking. J. Exp. Bot, 2000, 51: 249-264
Gaj MD. Direct somatic embyogenesis as a rapid and efficient system for in vitro regeneration of Arabidopsis thaliana (L.) Heynh. Plant Cell, Tissue and Organ Culture, 2001, 64: 39-46
Giroux RW, Pauls KP. Characterization of somatic embryogenesis-related cDNAs from alfalfa (MedicagosativaL). Plant Mol Biol, 1997, 33: 393-404
Gleddie S, Keller W, Setterfield G. Somatic embryogenesis and plant regeneration from leaf explants and cell suspensions of Solanum melongena (egg plant). Can J Bot, 1983, 61: 656-666
Gonzilez-Benito ME, Alderson PG. In vitro culture of Alstroemeria somatic embryos. Phyton, 1992, 53: 95-101
Gray WM, del Pozo JC, Walker L, et al. Identification of an SCF ubiquitin–ligase complex required for auxin response in Arabidopsis thaliana. Genes & Dev, 1999, 13(13): 1678-1691
Gupta PK, Pullman G. Method for reproducing coniferous plants by somatic embryogenesis using abscisic acid and osmotic potential variation. US Patent, 5036007, 1991.
Haccis B. Question of unicellar origin on non-zygotic embryos in callus cultures. Phytomotphology, 1978, 28: 74-81
Halper NW, Jensen AJ. Ultrastructural changes during growth and embryogenesis in carrot cell culture. Ultra Res., 1967, 18: 428 -443
Harada T, Yakuwa T. Studies on the morphogenesis of Asparagus VI. Effect of sugar on callus and organ formation in the in vitro culture of shoot segments of the seedlings. J Fac Agr Hokkaido Univ, 1983, 61: 307-314
Hatzopoulos P, Fong F, Sung ZR. Absicsic acid regulation of DC8 a carrot embryogenic gene. Plant Physiol, 1990, 94: 690-695
Hecht V, Vielle-Calzada JP, Hartog MV , Schmidt EDL, Boutilier K, Grossniklaus U, and Vries SC. The Arabidopsis somatic embryogenesis receptor kinase-1 gene is expressed in developing ovules and embryos and enhances embryogenic competence in culture. Plant Physiol, 2001, 127: 803-816
He DG, Tanner GT, Scott KJ. Somatic embryogenesis and morphogenesis in callus derived from the epiblast of immature embryo of wheat. Plant Sci, 1986, 45: 119-124
Henry Y, Vain P, Buyser J. Genetic analysis of in vitro plant tissue culture responses and regeneration capacities. Euphytica, 1994, 79: 45-48.
Hess JR, Carman JG. Embryogenic competence of immature wheat embryos: genotype, donor plant, environment and endogenous hormone concentrations. Crop Sci, 1998, 38: 249-253
Higashi K, Shiota H, Kamada H. Patterns of expression of the genes for glutamine synthetase isoforms during somatic and zygotic embryogenesis in carrot. Plant Cell Physiol, 1998, 39: 418-424
Hohe A, Winkelmann T, Schwenkel HG. CO2 accumulation in bioreactor suspension cultures of Cyclamen persicum Mill and its effect on cell growth and regeneration of somatic embryos.Plant Cell Reports, 1999a, 18(10): 863-867
Hohe A, Winkelmann T, Schwenkel HG. The effect of oxygen partial pressure in bioreactors on cell proliferation and subsequent differentiation of somatic embryos of Cyclamen persicum. Plant Cell Tissue Org Cult, 1999b, 59: 39-45
Hohe A, Winkelmann T, Schwenkel HG. Development of somatic embryos of Cyclamen persicum Mill in liquid culture. Gartenbauwissenschaft, 2001, 66: 219-224
Hu H, Xiong L, Yang Y. Rice SERK1 gene positively regulates somatic embryogenesis of cultured cell and host defense response against fungal infection. Planta, 2005, 222(1): 107-117
Hutchinson MJ, Saxena PK. Acetylsalicylic acid enhances and synchronizes thidiazuron-induced somatic embryogenesis in geranium (Pelargonium x hortorum Bailey) tissue cultures. Plant Cell Reports, 1996, 15 (7) : 512-515
Ikeda-Iwai M, Satoh S, Kamada H. Establishment of a reproducible tissue culture system for the induction of Arabidopsis somatic embryos. Exp Bot, 2002, 53: 1575-1580
Imamara J, Hiroshi H. Effects of abscisic acid and water stress on the embryo and plantlet formation in anther culture of Nicotiana tabacum cultivar S amsun Z. Pflanzenphysiol, 1980, 100: 285-289
Ito Y, Takaya K, Kurata N. Expression of SERK family receptor-like protein kinase gene in rice. Biochimica et Biophysica Acta, 2005, 1730(3): 253-258
Jelaska S. Recent development in somatic embryogenesis. Acta Pharmaceutica (Zagreb), 1994, 44(3): 387-406
Jianru Z, Qi-Wen N, Giovanna F, Nam HC. The WUS gene promotes vegetative-to-embryonic transition in Arabid. Plant J, 2002, 30(3): 349-359
Jianru Z, Niu QW, Yoshihisa L, Chua NH. Marker-free transformation: increasing transformation frequency by the use of regeneration-promoting genes. Curr Opi in Biol, 2002, 13(2): 173-180
Jiménez VM, Bangerth F. Endogenous hormone levels in explants and in embryogenic and non-embryogenic cultures of carrot. Physiol Plant, 2001a, 111: 389-395
Jimenez VM, Bangerth F. Endogenous hormone concentrations and embryogenic callus development in wheat. Plant Cell, Tissue and Organ Culture, 2001b, 67: 37-46
Jordy MN. Seasonal variation of organogenetic activity and reserves allocation in the shoot apex of Pinus pinaster Ait. Ann Bot, 2004, 93: 25-37
Kamada H, Harada H. Studies on the organogenesis in carrot tissue culture.Ⅰ. Effects of growth regulators on somatic embryogenesis and root formation. Z. P flonzenphysiol, 1979, 91: 255-266
Kamada H, Ishikawa K, Saga H, Harada H. Induction of somatic embryogenesis in carrot by osmotic stress. Plant Tissue Cult Lett, 1993, 10: 38-44
Kamo K, Jones B, Castillon J, Bolar J, Smith F. Dispersal and size fractionation of embryogenic callus increases the frequency of embryo maturation and conversion in hybrid tea roses. Plant Cell Reports, 2004, 22 (11): 787-792
Kaparakis G and Alderson PG. Role for Cytokinins in Somatic Embryogenesis of Pepper (Capsicum annuum L.)?. J Plant Growth Regul, 2008, 27: 110-114
Karim MZ, Yokota S, Azad M, et al. Relationship between starch accumulation and organ development at the different growth stages of callus in Kihada (Phellodendron amurense Rupr.). Plant Biotechnology, 2006, 23: 239-245
Kikuchi A, Sanuki N, Higashi K, et al. Abscisic acid and stress treatment are essential for the acquisition of embryogenic competence by carrot somatic cells. Planta, 2006, 223: 637-645
Kim B, Remko O, Vijay KS, et al. Ectopic expression of BABY BOOM triggers a conversion from vegetative to embryonic growth. Plant Cell, 2002, 14(8): 1737-1749
King SW. DNA sequence of an ABA-responsive gene (rab 15) from water-stress wheat roots. Plant Mol.Biol, 1992, 18: 119-121
Kitamiya E, Suzuki S, Sano T, et al. Isolation of two genes that were induced upon initiation of somatic embryogenesis on carrot hypocotyls by high concentrations of 2,4-D. Plant Cell Rep, 2000, 19: 551-557
Kiviharju E, Tuominen U, Tormala T. The effect of explant material on somatic embryogenesis of Cyclamen persicum Mill. Plant Cell, Tissue and Organ Culture, 1992, 28(2): 187-194
Kiyosue T and Shinozaki K. Cloning of a carrot cDNA for a member of the family of ADP- ribosylationfactors (ARFs) and characterization of the binding of nucleotides by its product after expression in e- coli. Plant Cell Physiol, 1995, 36: 849-856
Kiyosue T, Kamada H and Harada H. Induction of Somatic embryogenesis from carrot Seeds by Hypochlorite Treatment Plant Tissue Culture Letters, 1989, 6(3): 138-143
Kiyosue T, Kamada H, Harada H. Induction of somatic embryogenesis by salt stress in carrot. Plant Tissue Cult Lett, 1989, 6: 162-164
Kiyosue T, Nakajima M, Yamaguchi I, et al. Endogeneous levels of abscisic acid in embryogenic cells, non-embryogenic cells and somatic embryos of carrot (Daucus carota L.). Biochem Physiol Pflanzen, 1992b, 188: 343-347
Kiyosue T, Takano K, Kamada H, Harada H. Induction of somatic embryogenesis in carrot by heavy metal ions. Can J Bot, 1990, 68: 2021-2033
Kiyosue T, Yamaguchi-Shinozaki K, Shinozaki K, et al. Partial amino-acid sequence of ECP31, a carrot embryogenic-cell protein, and enhancement of its accumulation by abscisic acid in somatic embryos. Planta, 1992a, 186: 337-342
Kiyosue T, Yamaguchi-Shinozaki K, Shinozaki K, et al. Isolation and characterization of a cDNA that encodes ECP31, an embryogenic-cell protein from carrot. Plant Mol Biol, 1992, 19: 239-249
Kong L, Yeung EC. Effects of silver nitrate and polyethylene glycol on white spruce (Picea glauca) somaticembryo development: enhancing cotyledonary embryo formation and endogenous ABA content. Physiol Plant, 1995, 93: 298-304
Kochba J , Spiegel-Roy P, Saad S, et al. Stimulation of embryogenesis in Citrus tissue culture by galactose. Naturwissenschaften, 1978, 65: 261-262
Kopertekh LG and Butenko RG. Naturally occurring phytohormones in wheat explants as related to wheat morphogenesis in vitro. Rus J Plant Physiol., 1995, 42: 488-491
Kreuger M, Postma E, Brouwer Y et al. Somatic embryogenesis of Cyclamen persicum in liquid medium. Physiol Plant, 1995, 94: 605-612
Kuklin AD, Denchev PD, Atanassov AI, Scragg AH. Alfalfa embryo production in airlift vessels via direct somatic embryogenesis. Plant Cell, Tissue and Organ Culture, 1994, 38: 19-23
Landschulz WH, Johnson PF, Maknight SL. The leucine zipper: a hypothetical structure common to a new class of DNA binding proteins. Science, 1988, 240(4860): 1759-1764
Laurent L, Philippe V, Francoise C, et al. Effect of abascisic acid and high concentrations of PEG on Hevea brasiliensis somatic embryos development. Plant Sci, 1997, 124(2): 183-191
Leal I, Misra S. Molecular cloning and characterization of alegumin-like storage protein cDNA of Douglasfir seeds. Plant Mol Biol, 1993,21: 709-715
Lee KP and Lee DW. Somatic embryogenesis and plant regeneration from seeds of wild Dicentra spectabilis (L.) LEM. Plant Cell Rep, 2003, 22: 105-109
Libbenga KR, Mennes AM. Plant hormones. In: Davies PJ (ed). Physiology, Biochemistry andMolecular Biology. Dordrecht, the Netherlands: Kluwer Academic Pub, 1995, 270-275
Li BC and Wolyn DJ. Abscisic acid and ancymidol promote conversion of somatic embryos to plantlets and secondary embryogenesis in Asparagus officinalis L. In Vitro Cell Dev. Biol.-Plant, 1996b, 32: 223-226
Li BC and Wolyn DJ. Temperature and genotype affect asparagus somatic embtyogenesis. In Vitro Cell Dev. Biol.-Plant, 1996a, 32: 136-139
Li MH, Steven EU, Andris K, et al. Improvement of anther culture methods or doubled haploid production in barley breeding. Plant Cell Rep, 1993, 12: 328-334
Lindzen E, Choi JH. A carrot cDNA encoding an atypical protein kinase homologous to plant calcium-dependent protein kinases.Plant Mol Biol, 1995, 28: 785-797
Liu CQ, Xia XL, Yin WL, Huang LC, Zhou JH. Shoot regeneration and somatic embryogenesis from needles of redwood (Sequoia sempervirens (D.Don.) Endl.) Plant Cell Rep, 2006, 25(7): 621 -628
Lo Shiavo F, Filippini F, Cozzani F, et al. Modulation of auxin-binding proteins in cell suspension I. Different responses of carrot embryo cultures.Plant Physiol, 1991, 97: 60-64
Lyngved R, Renaut J, Hausman JF, et al. Embryo-specific Proteins in Cyclamen persicum Analyzed with 2-D DIGE. J Plant Growth Regul, 2008, 27: 353-369
Mahesh-Wari SC, Tyagi AK, Malhotra K. Induction of haploidy from pollen grains in angiosperms the current status. Theor Appl Genet, 1980, 58: 193-206
Mamiya K, Sakamoto Y. Effects of sugar concentration and strength of basal medium on conversion of somatic embryos in Asparagus officinalis L. Sci Hort, 2000, 84: 15-26
Manickam A, vanDamme EJM, Kalaiselvi K, et al. Isolation and cDNA cloning of an Em-like protein from mungbean (Vigna radiata) axes. Physiol Plantarum, 1996, 97(3): 524-530
Manisha Vand BansalYK. Somatic embryogenesis and plantlet regeneration in semul (Bombax ceiba). Plant Cell, Tissue and Organ Culture, 2004, 79: 115-118
Mathew MM and Philip VJ. Somatic embryogenesis versus zygotic embryogenesis in Ensete superbum. Plant Cell, Tissue and Organ Culture, 2003, 72: 267-275
MedicagoT. Root-forming and embryogenic cultures. Plant Physiol, 2003, 133(1): 218-230.
Meskaoui AE, Desjardins Y, Tremblay FM. Kinetics of ethylene biosynthesis and its effects during maturation of white spruce somatic embryos. Physiol Plant, 2000, 109: 333-342
Michalczuk L, Cooke TJ, Conen JD. Auxin levels at different stages of carrot somatic embryogenesis. Phytochemistry, 1992, 31: 1097-1103
Michalczuk L, Ribnicky DM, Cooke TJ, et al. Regulation of indole-acetic acid biosynthetic pathway in carrot cell cultures. Plant Physiol, 1992, 100: 1346-1353
Michler CH, Lineberger RD. Effects of light abscisic levels in carrot suspension cultures. Plant Cell Tiss Org Cult, 1987, 11: 189-207
Miku?a A, Rybczyński JJ, Skierski J,et al. Somatic embryogenesis of Gentiana genus IV: Characterisation of Gentiana cruciata and Gentiana tibetica embryogenic cell suspensions. In: Hvolsef-Eide AK and Preil W, eds. Liquid Culture Systems for In Vitro Plant Propagation, Netherlands: Springer, 2005. 345-358
Mithila J, Hall JC, Victor JMR, Saxena PK. Thidiazuron induces shoot organogenesis at low concentrations and somatic embryogenesis at high concentrations on leaf and petiole explants of African violet (Saintpaulia ionantha Wendl.). Plant Cell Rep, 2003, 21: 408-414
Mohanna AM, Karam NS. In vitro propagation of wild Cyclamen persicum Mill from seedling tissue. Acta Horticilturae, 2000,(530): 243-250
Motoike SY, Skirvin RM, Norton MA, et al. Somatic embryogenesis and long term maintenance of embryogenic lines from fox grapes. Plant Cell Tissue Org Cult, 2001, 66: 121-131
Murasaki K and Tsurushima H. Improvement on clonal propagation of cyclamen in vitro by the use of etiolated petioles. Acta Hort, 1988, 226: 721-724
Murashige T, Skoog F. A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol Plant, 1962, 15: 473-497
Nakagawa H, Saijyo T, Yamauchi N, et al. Effects of sugars and abscisic acid on somatic embryogenesis from melon (Cucumis melo L.) expanded cotyledon. Sci Hort, 2001, 90: 85-92
Nishiwaki M, FujinoK, KodaY, Masuda K, Kikuta Y. Somatic embryogenesis induced by the simple application of abscisic acid to carrot (Daucus carota L.) seedling in culture. Planta, 2000, 211: 756-759
Nitsch C, Norreel B. Effect d′u m choc thermique sue le pouvoir embryogene dupollen Datura innoxia culture dans I′anther ouisolede I′anthere. C R Acad Sci Paris, 1973, 276: 303-306
Nokta M, EatonD, Steinsland OS, Albrecht T. Ca 2+ responses in cytomegalovirus-infected fibroblasts of human origin. Virology, 1987, 157: 259-267
Nolan KE, Irwanto RR, Rose RJ. Auxin up-regulates MtSERK1 expression in both medicago truncatula root-forming and embryogenic cultures. Plant Physiol, 2003, 133(1): 218-230
Osuga K, Masuda H, Komamine A. Synchronization of somatic embryogenesis at high frequency using carrot suspension cultures: model systems and application in plant development. Methods in Cell Science, 1999, 21: 129-140
Overvoorde PJ, Grimes HD. The role of calcium and calmodulin in carrot somatic embryogenesis. Plant Cell Physiol, 1994, 35: 135-144
Owen HR, Miller AR. Haploid plant regeneration from anther cultures of three North American cultivars of strawberry ( Fragaria ananassa Duch). Plant Cell Rep, 1996, 15(12): 905-909
Padmanabhan K, Cantliffe DJ, Koch KE. Auxin-regulated gene expression and embryogenic competence in callus cultures of sweet potato, Ipomoeabatata(L.)Lam. Plant Cell Rep, 2001, 20: 187-192
Quiroz-Figueroa FR, Rojas-Herrera R, Galaz-Avalos RM, Loyola-Vargas VM. Embryo Production through somatic embryogenesis can be used to study cell differentiation in plants. Plant Cell, Tiss and Organ Culture, 2006, 86(3): 285-301
Rensing SA, Lang D, Schumann E, Reski R, Hohe A. EST sequencing from embryogenic Cyclamen persicum cell cultures identifies a high proportion of transcripts homologous to plant genes involved in somatic embryogenesis. J Plant Growth Regul, 2005, 24: 102-115
Ribas LLF, Guerra MP, Zanette F, Kulchetscki L. Somatic embryogenesis in Aspidosperma polyneuron Mull. Arg. In: Jain SM, Gupta PK, Newton RJ, eds. Somatic Embryogenesis in Woody Plants. Dordrecht, Boston, London: Kluwer Academic Publishers, 2000. 509-537
Rougtan JP, Latche A, Fallot J. Inhibition of ethylene production and stimulation of carrot somatic embryogenesis by salicylic acid. Biol Plant, 1990, 32: 273-278
Ruffoni B, Semeria L, Profumo P. Cyclamen persicum Mill. Somatic embryos developed in suspension cultures: histological analysis and conversion to plant. Acta Horticulturae, 2000, 520: 83-90
Ruffoni B. Embryogenic suspension cultures of cyclamen for the production of artificial seeds. Colture Protette, 1998, 27(11): 91-96
Russinova E, Slater A, Atanassov AI, et al. Cloning novel alfalfa cyclin sequences a RACE- PCR approach. Cellular and Molecular Biology, 1995, 41: 703-714
Sandra LS, Linda WK, Kelly MY, et al. LEAFY COTYLEDON2 encodes a B3 domain transcription factor that induces embryo development. Plant Biol, 2001, 20(98): 11806-11811
Santos MDO, Romano E, Yotoko KSC, et al. Characterisation of the cacao somatic embryogenesis receptor-like kinase (SERK) gene expressed during somatic embryogenesis. Plant Sci, 2005, 168(3): 723-729
Sasaki K, Shimomura K, Kamada H, Harada H. IAA metabolism in embryogenic and non-embryogenic carrot cells. Plant Cell Physiol, 1994, 35: 1159-1164
Sato S, Toya T, Kawahara R, et al. Isolation of a carrot gene expressed specifically during early-stage somatic embryogenesis. Plant Mol Biol, 1995, 28: 39-46
Savona M, Ruffoni B, Falasca G, et al (2007). NCBI GenBank database, #EF661828, #EF672247, #DQ350614. http://www.ncbi.nlm.nih.gov
Schmidt ED, Guzzo F, Toonen MAJ, et al. A leucine-rich repeat containing receptor-like kinase marks somatic plant cell competent to form embryos. Development, 1997, 124(10): 2049-2062
Schmidt Th, Ewald A, Seyring M, Hohe A. Comparative analysis of cell cycle events in zygotic and somatic embryos of Cyclamen persicum indicates strong resemblance of somatic embryos to recalcitrant seeds. Plant Cell Rep, 2006, 25: 643-650
Schwenkel HG, Grunewaldt J. Somaclonal Variation in Cyclamen Persicum Mill after in Vitro Mass Propagation. Gartenbauwissenschaft, 1991, 56, 16-20
Schwenkel HG, Winkelmann T. Plant regeneration via somatic embryogenesis from ovules of Cyclamen persicum Mill.Plant Tiss Cult Biotechnol, 1998, 4: 28-34
Sengupta C, Raghavan V. Somatic embryogenesis in carrot cell suspension.Ⅰ. Pattern protein and nucleic acid synthesis. J Exper Bot, 1980, 30: 247-258
Seyring M, Hohe A. Induction of desiccation tolerance in somatic embryos of C. persicum MILL. J Hortic Sci Biotechol, 2005, 80: 65-69
Shah K, Gadella TW, Jrvan EH, et al. Subcellular localization and oligomerization of the Arabidopsis thaliana somatic embryogenesis receptor kinase1 protein. J Mol Biol, 2001, 309: 641-655
Sharp W R, Sondahl M R, Caldas L S, et al. The physiology of in vitro asexual embryogenesis. Hort Rev, 1980, 2: 268 -310
Shibli RA, Ajlouni MM. Somatic embryogenesis in the endemic black iris. Plant Cell Tiss OrgCult, 2000, 61: 15-21
Shimada T, Hirabayashi T, Endo T, et al. Isolation and characterization of the somatic embryogenesis receptor-like kinase gene homologue (CitSERK1)from Citrus unshiu Marc. Sci Hortic, 2005, 103(2): 233-238
Smeeken S. Sugar-induced signal transduction in plants. Annu Rev Plant Physiol Plant Mol Biol, 2000, 51: 49-81
Sofiari E, Raemakers CJJM, Kanju E, et al. Comparison of NAA and 2, 4-D induced somatic embryogenesis in Cassava. Plant Cell, Tissue and Organ Culture, 1997, 50: 45-56
Somleva MN, Schmidt EDL, Vries SC. Embryogenic cells in Dactylis glomerata L.(Poaceae) explants identified by cell tracking and by SERK expression. Plant Cell Rep, 2000, 19(1): 718-726
Songstad DD. Effect of aminocyclopropane carboxylic acid, silver nitrate and norbornadiene on plant regeneratin from maize callus cultures. Plant Cell Rep, 1988, 7: 262-265
Stange L, Osborne D J. Cell specificity in auxin and ethylene induced‘supergrowth’in Riella helicophylla. Planta, 1988, 175: 341 - 347.
Steward FC, Mapes MO, Hears K. Growth and organized development of cultured cells. II. Growth and division of freely suspended cells. Am.J.Bot, 1958, 45: 705-708
Stitn S, Jacobsen HJ. Marker proteins for embryogenic differentiation patterns in pea callus. Plant Cell Reports, 1987, 6: 50 -54
Suprasanna P, Desai NS, Nishanth G, et al. Differential gene expression in embryogenic, non-embryogenic and desiccation induced cultures of sugarcane. Sugar Technol, 2004, 6(4): 305-309
Takamura T, Miyajima I, Matsuo E. Somatic embryogenesis of Cyclamen periscum Mill. 'Anneke' from aseptic seedlings. Plant Cell Reports, 1995, 15(2): 22-25
Takamura T, Nagita Y and Tanaka M. Effects of temperature on in vitro plant regeneration through somatic embryogenesis in Cyclamen persicum Mill. Hot Res, 2003, 2(1): 25-28
Takamura T. and Tanaka M. Somatic embryogenesis from the etiolated petiole of cyclamen (Cyclamen persicum Mill). Plant Tissue Culture Letters, 1996, 13(1): 43-48
Tetteroo FAA, Bino RJ, Bergervoet JHW et al. Effect of ABA and slow drying on DNA replication in carrot (daucus carota) embryoids. Physiologia Plantarum, 1995, 95: 154-158
ThiL T, Pleschka E. Somatic embryogenesis of some Daucus species influenced by ABA. J ApplBot Food Qual, 2005, 79: 1- 4
Thomas C, Meyer D, Himber C, Steinmetz A. Spatial expression of a sunflower SERK gene during induction of somatic embryogenesis and shoot organogenesis. Plant Physiol Biochem, 2004, 1(42): 35-42
Thomas TL. Gene expression during plant embryogenesis and germination: An overview. Plant Cell, 1993, 5(10): 1401-1410
Thorpe TA, Murashige T. Some hischemical changes underlying shoot initiation in tabacco callus cultures. Can J Bot, 1970, 48: 277 -285
Tisserat B, Murashige T. Effects of ethephon, ethylene and 2, 4-dichlorophenoxyacetic acid on asexual embryogenesis in vitro. Plant Physiol, 1977, 60(3): 437-439
Toivonen L, Laakso S, Rosenqvist H. The effects of temperature on growth, indole alkaloid accumulation and lipid composition of Catharanthus roseus cell suspension cultures. Plant Cell Rep, 1992, 11: 390-394
TornéJM, Moysset L, Claparols I, et al. Photo control of somatic embryogenesis and polyamine content in Araujia sericifera petals. Physiol Plant, 1996, 98: 413-418
Tremblay L,Tremblay FM. Effect of gelling agents ,ammonium nitrate and light on the development of Picea mariana (Mill) B. S. P. (black spruce) and Picea rubens Sarg. (red spruce) somatic embryos. Plant Sci, 1991, 77: 233-242
Verhagen SA, Wann SR. Norway spruce somatic embryogenesis High frequency initiation from light cultured mature embryos. Plant Cell Tiss Org Cult, 1989, 16: 103-111
Verma DC, Dougall DK. Influence of carbohydrates on quantitative aspects of growth and embryo formation in wild carrot suspension cultures. Plant Physiol, 1977, 59: 81- 85
Vikrant and Rashid A. Direct as well as indirect somatic embryogenesis from immature (unemerged) inflorescence of a minor millet Paspalum scrobiculatum L. Euphytica, 2001, 120: 167-172
Von Aderkas P, Thompson RG, Zaki M, Benkrima L. Somatic embryogenesis of western larch (Larix occidentalis). In: Bajaj YPS ed. Biotechnology in Agriculture and Forestry Somatic Embryogenesis and Synthetic Seeds, Berlin: Springer-Verlag, 1995. 378-387
Wang YH and Bhalla PL. Somatic embryogenesis from leaf explants of Australian fan flower Scaevola aemula R. Br. Plant Cell Rep, 2004, 22: 408-414
Weatherwax SC, Ong MS, Degenhardt J, et al. The interaction of light and abscisic acid in the regulation of plant gene expression. Plant Physiol, 1996, 111: 363-370
Welander M, Maheswaran G. Shoot regeneration from leaf explants of dwarfing apple rootstocks. J Plant Physiol, 1992, 140: 223-228
Wetherell DF, Dougall DK. Sources of nitrogen supporting growth and embryogenesis in cultured wild carrot issue. Physiol Plant, 1976, 37: 97-103
Wicart G, Mouras A, Lutz A. Histological study of organogenesis and embryogenesis in Cyclamen persicum Mill. Tissue cultures: evidence for a single organogenesis pattern. Protoplasma, 1984, 119(3): 159-167
Wilson SM, Thorpe TA . Somatic embryogenesis in Picea glauca (white spruce), P. engelmannii (Engelman spruce) and P. engelmannii complex (interior spruce). In: Mohan J S, Gupta P K, Newton R J eds. Somatic Embryogenesis in Woody Plants, Vol. 1. Dordrecht, The Netherlands: Kluwer Academic Publishers, 1995. 37-54
Winkelmann T, Heintz D, van Dorsselaer A, Serek M, Braun HP. Proteomic analysis of somatic and zygotic embryos of Cyclamen persicum Mill. reveal new insights into seed and germination physiology. Planta, 2006, 224: 508-519
Winkelmann T, Hohe A, Schwenkel HG. Establishing embryogenic suspension cultures in Cyclamen persicum 'Purple Flamed'. Advances in Horticultural Science, 1998, 12(1): 25-30
Winkelmann T, Meyer L, Serek M. Desiccation of somatic embryos of Cyclamen persicum Mill. J Horticult Sci Biotechnol, 2004, 79 (3): 479-483
Winkelmann T, Schwenkel H G, Hohe A. Development of somatic embryos of Cyclamen persicum Mill in liquid culture. Gartenbauwissenschaft, 2001, 66(5): 219-224
Winklemann T, Serek M. Genotypic differences in callus formation and regeneration of somatic embryos in Cyclamen persicum Mill. Euphytica, 2005, 144: 109-117
Xiang TH, Liang HM, Zhong HX. Physiological and biochemical changes during the establishment of embryogenic cell suspension cultures of rice (Oryza satira L.). II: changes of the contents of amino acid, polyamine and endohormone. ActaAgronomica Sinica, 1997, 23(3): 353-359
Yang H, Saitou T, Komeda Y, et al. Arabidopsis thaliana ECP63 encoding a LEA protein is located in chromosome 4. Gene, 1997, 184: 83-88
Yang H, Saitou T, Komeda Y, et al. Late embryogenesis abundant protein in Arabidopsis thaliana homologous to carrot ECP31. Physiol Plant, 1996, 98: 661-666
Yasuda H, Nakajiima M, Ito T, et al. Partial characterization of genes whose transcripts accumulate preferentially in cell clusters in the early stages of carrot somatic embryogenesis.Plant Mol Biol, 2001, 45: 705-712
Yasuda H, Sato T, Masuda H. Rapid and frequent embryogenesis from single cells of regenerated carrot plantlets. Biosci Biotechnol Biochem, 1998, 62: 1273-1278
Yeung EC, Stasolla C. Somatic embryogenesis-apical meristem formation and conversion. Korean J Plant Tiss Cult, 2000, 27: 253-258
Yeung EC, Stasolla C, Kong L. Apical meristem formation during zygotic embryo development of white spruce. Can J Bot, 1998, 76: 751-761
Zhu C, Kamada H, Harada H, et al. Isolation and characterization of a cDNA encoded an embryogenic cell protein-63 related to embryogenesis from carrot. Acta Bot Sin, 1997, 39: 1091-1098
Zimmerman JL. Somatic embryogenesis: a model for early development in higher plants. Plant Cell, 1993, 5: 1411-1423
曹有龙,贾勇炯,陈放,罗青,曲琳.枸杞花药愈伤组织悬浮培养条件下胚状体发生与植物株再生.云南植物研究,1999,21(3):346-350
车美建,赖钟雄,赖呈纯,等.荔枝体细胞胚胎发生早期的3种内源激素含量变化.热带作物学报,2005,26(2):55-61
陈以峰,周燮,汤日圣,张金渝,梅传生.水稻体细胞培养中胚性细胞出现与IAA的关系.植物学报,1998,40(5):474-477
陈健辉,方璟.玉米种子萌发过程幼叶细胞中淀粉粒的积累观察.广西植物,2003,23(5) :440-444
程玉兰,黄美娟,刁丰秋等.蔗糖调控培养对胡萝卜体细胞胚内源ABA水平的效应.植物学报,1999 ,41(7):761-765
崔德才和徐培文.植物组织培养与工厂化育苗.北京:化学工业出版社,2004
崔凯荣,戴若兰.植物体细胞发生的分子生物学.北京:科学出版社,2000,50-171
崔凯荣,陈克明,王晓哲,等.植物体细胞胚发生研究的某些现状.植物学通报,1993,10(3):14-20
崔凯荣,刑更生,周功克,等.植物激素对体细胞胚发生的诱导和调节.遗传,2000,22(5):349-354
邓向阳,卫志明.幼胚长度、2 ,4-D浓度、光强度等对花生体细胞胚发生的影响及高效再生系统的建立.植物生理学报,2000,26 (6):525-531
范昌发,郭骁才,贾敬芬,牛天堂.植物细胞中淀粉代谢与离体形态发生途径决定的关系.华北农学报,2000,15:52-57
范现丽,王高,申晓辉.蓝百合体细胞胚胎发生及植株再生的研究.见:张启翔,编.中国观赏园艺研究进展2008——中国园艺学会观赏园艺专业委员会2008年学术年会论文集.中国黑龙江哈尔滨:中国园艺学会观赏园艺专业委员会、国家花卉工程技术研究中心, 2008. 124-127
高述民. ABA和PEG对胡萝卜体细胞胚诱导和调控的影响.西北农林科技大学学报,2001,29(2):13-16
宫相忠,黄健.苜蓿组织培养体细胞胚发生的细胞学观察.青岛海洋大学学报,1997, 24(4):504-508
顾玉红,高述民,郭惠红,李凤兰.文冠果的体细胞胚胎发生.植物生理学通讯,2004 ,40(3):311-313
韩碧文,刘淑兰.植物离体体细胞胚的发生.植物生理学通讯,1988,(1):9-15
韩晓玲,王冰雪,林雪,贾敬芬.小冠花高效体细胞胚胎发生与植株再生的研究.西北大学学报(自然科学版),2006,36(3):420-423
何丽君,于卓.濒危植物四合木(Tetraena mongolica Maxim)的离体培养组织学观察及再生植株的研究.内蒙古农业大学学报,2003,24(2):27-32
何丽君,齐冰杰.四合木体细胞胚胎发生中细胞组织学的淀粉积累动态的研究.内蒙古草业),2002,14(3):39-41
侯喜林.仙客来实生黄化叶柄培养的研究.园艺学报,1991,18(1): 81-86
胡博然,徐文彪,马峰旺.枸杞胚细胞悬浮培养系统建立的研究.西北农林科技大学学报(自然科学版),2003, 31(1):97-100
胡忠,徐艳,高欢欢,关秀琼.影响宁夏枸杞愈伤组织体细胞胚发生的因素.汕头大学学报(自然科学版),2006,21( 4): 57-63
胡适宜.同时显示胚中贮藏的淀粉、蛋白、脂类的制片方法植物学通报. 1994,11(4):49-51
胡适宜.被子植物胚胎学.北京:高等教育出版社,1982. 237-247
[39]黄学林,李筱菊,傅家瑞,劳彩玲. Thidiazuron对苜蓿愈伤组织的乙烯生成及其体细胞胚胎发生的影响.植物生理学报,1994,20:367-372
黄丹枫,刘佩瑛.魔芋再生植株形态发生途径的细胞组织学观察.上海农学院学报,1994,12(1):25-30
黄美娟,黄绍兴,朱微,等.采用调控培养方法提高胡萝卜体细胞胚活力的研究.科学通报,1993,38(6):550-553
赖钟雄,陈春玲.龙眼体细胞胚胎发生过程中的内源激素变化.热带作物学报,2002, 23(2): 41-47
赖钟雄,陈振光.龙眼胚性细胞悬浮培养及其体胚发生再生植株.应用与环境生物学报,2002,8(5): 485-491
赖钟雄,黄浅,林秀莲等.荔枝胚性悬浮细胞系的快速建立及其体胚植株的再生.农业生物技术科学,2007,23(1):28-32
李成浩,刘宝光,王伟达,刘立琨,Yu Chang-Yeon.温度对积棋次生胚发生和植株再生的影响.植物生理学通讯,2007,43(3): 453-456
李会宁,闫锋.仙客来叶片的组织培养体细胞胚诱导研究.汉中师范学院学报,2001,19(2): 81-83
李丽,万勇善,刘风珍. 2,4-D浓度对花生体细胞胚发生的影响.生物技术,2005,15(3):77-79
李杉,邢更妹,崔凯荣,王亚馥.植物体细胞胚发生中ATP酶活性时空分布动态与内源激素的变化.植物学通报,2001,18 (3):308-317
李天宪,赵林,冯锋,等.人巨细胞病毒小鼠模型的建立.微生物学报,1996,36(4):292-294
李学红,李冬玲,张举仁,等.玉米芽尖培养中的高频率体细胞胚胎发生与植株再生(简报). 植物生理学通讯,2000,36(5):430-433
李中奎,刘成运,刘文平.水稻愈伤组织内部胚性细胞的形成及发育.植物研究,2001,21(1):62-64
林荣双,王庆华,梁丽琨,郑秋生,肖显华.花生体细胞胚发生发育过程中淀粉代谢的动态变化.烟台大学学报(自然科学与工程版),2000,13(3):226-229
林荣双,王庆华,梁丽琨,肖显华. TDZ诱导花生幼叶的不定芽和体细胞胚发生的组织学观察.植物研究,2003,23(2):169-172
刘福平,陈移亮.细胞分裂素对蝴蝶兰胚性愈伤组织诱导的影响.热带作物学报,2008,29(1):42-46
刘华英,萧浪涛,鲁旭东,吴顺.柑橘体细胞胚发生的组织细胞学研究.果树学报,2004,21(4):311-314
刘华英,萧浪涛,鲁旭东,吴顺.柑橘体细胞胚发生的组织细胞学研究.果树学报,2004,21(4):311-314
刘艳芝,韦正乙,谭化,高明.花生体细胞胚发生及植株再生.吉林农业科学,2008, 33(4):7-10
刘一鸣,刁丰秋,张雷,等.胡萝卜LEA基因家族新成员DcLEA1的克隆及其结构与表达特征分析.自然科学进展,2004,8(8):882-891
牛德水,邵启全,张敬等.枸杞悬浮培养条件下的胚状体发生.遗传,1990,6:7-9
潘建伟,王卫军,潘伟槐,朱睦元.大麦花药培养及其胚性悬浮细胞系的高频快速建立.浙江大学学报(理学版),2002,29(4):454-458
潘炽.植物组织培养.广州:广东高等教育出版社,2000. 20-59
乔琦,肖娅萍.防风体细胞胚发生发育中的淀粉体和多糖动态.西北植物学报,2007,27(2):0388-0391
秦凡,周吉源.不同植物生长调节剂对蝴蝶兰快速繁殖的影响.武汉植物研究,2003,21(5):452-456
曲桂芹,张贤泽,霍俊伟.影响大豆体细胞胚诱导因素的研究.植物研究,2001,21(2):210-214
曲复宁,由翠荣,康黎芳,等.利用仙客来种苗组培快繁建立单株无性系的研究.西北农林科技大学学报,2003,31(3):81-86
曲复宁,由翠荣,龚雪芹,王云山,康黎芳.仙客来(Cyclamen persicum Mill)无性繁殖体系建立中不定芽分化能力的数量性分析.北方园艺,2001,(5): 62-63
曲复宁,由翠荣,龚雪芹,等.仙客来组织培养中再生器官类型及增殖稳定性比较.植物学通报,2004,21(5):559-564
盛德策,李凤兰,刘忠华.,植物体细胞胚发生的细胞生物学研究进展.,西北植物学报,2008,28(1):0204-0215
施小龙,邢更妹,汪丽虹,王亚馥.植物钙信号系统与体细胞胚发生.生命科学,2002,14(5):302-304
苏浴源,吴永祯,曲占礼,等.仙客来幼芽组织培养的研究.华北农学报,2002,17(增刊):203-205
Thomas T D.糖、GA3及ABA对印度娃儿藤( Tylophora indica ( Burm. f.) Merrill)体细胞胚发生的影响.生物工程学报, 2006, 22(3): 465-471
王波,王鹤,谭巍,等.仙客来组织培养及植株再生.北方园艺,2007,1:166-167
王杰,刘国锋,包满珠,黄莉.麝香百合胚性愈伤组织状态的调整与植株再生.园艺学报,2008,35(12:1795-1802
王康,朱慧森,董宽虎.植物LEA蛋白及其基因家族成员PM16研究进展.草业科学,2008,25(2):97-102
王丽,鲍晓明,黄百渠,郝水.香雪兰外植体形态学极性决定的体细胞胚胎发生.植物学报, 1998,40(2):138-143
王丽,段晓刚,郝水.香雪兰的直接与间接体细胞胚胎发生.实验生物学报,1999,32(2):175-182
汪丽虹,王星,崔凯荣,焦成瑾,王亚馥.石刁柏及党参体细胞胚发生中的淀粉代谢动态. 植物学通报,1996,13:41-45
王清连,王敏,师海荣.植物激素对棉花体细胞胚胎发生的诱导及调节作用.生物技术通讯,2004,15(6):577-579
王仑山,杨汉民,王亚馥,康文隽.伊贝母组织培养中体细胞胚的形成及细胞组织学观察.西北植物学报,1989,9(2):76-81
王亚馥,崔凯荣,陈克明,高清祥,张德华,焦成瑾.小麦组织培养中体细胞胚胎发生的细胞胚胎学及淀粉消长动态的研究.实验生物学报,1993,26(3):259-267
王义,赵文君,孙春玉,杨晶,蒋世翠,张美萍. 2,4-D、BA对人参体细胞胚胎发生过程的影响研究.中国生物工程杂志,2008,28(10):106-112
王义,赵文君,杨忠,孙春玉,张美萍.人参体细胞胚胎发生及植株再生研究.药物生物技术,2008,15(4):278-28
马和平,李毅,邸利,马彦军,魏继辉.枸杞体细胞胚胎发生及植株再生的研究.甘肃农业大学学报,2005,40(1):26-30
马义德,邢更生,戴若兰,等.生物组织切片图像的计算机三维重建技术现状.生物技术通讯,2000,11(4):303-308
魏岳荣.香蕉(Musa spp.)胚性细胞悬浮培养及其超低温保存和植株再生的研究:[博士学位论文].广州:中山大学生命科学学院,2005
魏岳荣,黄学林,李佳等.贡蕉胚性细胞悬浮系的建立和植株再生.生物工程学报,2005,21(1):58-65
吴春霞.植物细胞悬浮培养的影响因素.安徽农业科学,2009,37(1):36-38
夏光敏,陈惠敏.植物生长调节物质对石防风体细胞胚发生与发育的影响.植物学报,1993,35(8):644-648
夏光敏,李忠谊,于家驹,等.峨参原生质的体细胞胚同步化发生及植株再生.实验生物学报,1991,24(4):395-401
肖显华,林荣双,王庆华,王顺珍. 2,4-D诱导的花生体细胞胚发生的组织学研究.植物学通报,1999,16(6):691-695
肖望,黄学林.贡蕉胚性细胞悬浮培养过程中重要生长参数的研究.广东教育学院学报,2005,25(3):77-79
邢更生,崔凯荣,戴若兰,李杉,山仑.小麦胚性细胞发生中淀粉代谢动态的量化研究.兰州大学学报(自然科学版),1998,34:128-132
邢更生,邢更妹,崔凯荣,王亚馥.枸杞胚性细胞分化的超微结构研究.电子显微学报,1999,18(4):395-400
刑更生,崔凯荣,山仑,等.植物体细胞胚发生的分子基础.遗传,1999,21(1):30-34
邢登辉,赵云云,黄承芳.皇冠草体细胞胚胎发生及其体胚发生过程中内源激素的变化.生
物工程学报,1999,15(1): 98-101
邢登辉,吴琴生,刘大钧.禾谷类作物细胞培养.生物学通报,1994,29(7): 1-3
徐林林,芦笛,陆拢等.水稻悬浮细胞系的建立及培养条件对生物产量的影响.植物生理学通讯,2006,42(4):612-616
徐元红,朱四易.伊贝母与平贝母体细胞胚诱导条件的比较.陕西师范大学学报,1998,26(2):119-120
颜昌敬.农作物组织培养.上海:上海科学技术出版社,1991. 283-299
杨汉民,高清祥.枸杞体细胞胚的诱导与形态发.兰州大学学报(自然科学版),1992,28(1):87-89
杨会钗,李彦群.改良的苏木精染色液配制方法探讨.中华中西医杂志,2003,4(16):
叶克难,李文雷,徐增富,黄俊潮,李宝健.苜蓿体细胞胚胎发生过程中DNA、RNA和蛋白质的合成动态.实验生物学报,1992,25(4):403-411
尹文兵,李丽娟,黄勤妮,等.胡萝卜愈伤组织的诱导及细胞悬浮培养研究.山西师范大学学报,2004,18(2):71-76
俞长河,陈振光.荔枝胚性悬浮培养物的建立、保持和优化原生质体分离的研究.热带作物学报,1998,19(3):16-20
袁澍,贾勇炯,林宏辉.诱导植物体细胞胚发生的几个生理因素.植物生理学通讯,2003,39(5):508-512
张宝红,李秀兰. TDZ (thidiazuron)在棉花组织培养中的效应.作物学报,1995,21:253-258
张栋,陈季楚. ABA、NAA诱导水稻胚性愈伤组织的研究.实验生物学报,1995,28(3):329-334
张健,吕柳新,黄春梅等.柑桔体细胞胚的发生及其发育过程中淀粉粒动态研究.洛阳师范学院学报,2005,5:104-106
张树录,郑国錩,聂秀菀.糜子离体体细胞胚胎发生的组织学研究.西北植物学报,1992, 12(1):17-21
张树录.禾本科植物组织培养中的体细胞胚胎发生.植物生理学通讯,1985,(6):15-20
张新英,黄亚斌,尹光初,周思君.离体培养下大豆体细胞胚胎发生的组织学研究.植物学报,1992,34(3):214-218
赵桂兰,刘艳芝,尹爱平等.大豆花药培养中体细胞胚萌发的研究.科学通报,1998,43(14):1512-1516
周春江,李卫东,葛会渡.草莓悬浮细胞系的建立及某些影响因素.植物生理学通讯,2002,38(1):22-24
周俊彦.植物体细胞在组织培养中产生的胚状体.植物生理学报,1981,7(4):389-396
周连霞,马锋旺,陈登文,秦静远,程瑞红. TDZ对仙客来不同外植体再生的影响.西北农林科技大学学报(自然科学版),2006,34(4):39-42
周燕,高述民,李凤兰.胡萝卜体细胞胚胎发生中的细胞组织化学和蛋白质组成变化.植物生理学通讯,2004,40:181-183
庄伟建,张书标,刘思衡.花生体细胞胚的诱导及其植株再生.热带亚热带植物学报,1999 7(2):153-158
郑泳,王君晖,黄纯农.通过花药直接液体培养建立大麦胚性悬浮细胞系.杭州大学学报(自然科学版),1996,23(2):202-203
朱至清,王敬驹,孙敬三.在无激素的培养基上水稻和小麦花粉胚的发育.植物学报,1976,18:239-244
Albertini E, Marconi G, Reale L, Barcaccia G, Porceddu A, Ferranti F, Falcinelli M. SERK and APOSTART, candidate genes for apomixis in poa pratensis. Plant Physiol, 2005, 138(4): 2185-2199
Ammirato PV. Techniques for propagation and breeding. In: Evans DA, Sharp WR, Ammirato PV (eds). Handbook of Plant Cell Culture.Vol I. New York: Macmillan Pub, 1983. 82-84
Anil VS, Rao KS. Calcium mediated signaling during sandal wood somatic embryogenesis. Role for exogenous calcium as second messenger. Plant Physiol, 2000, 123: 1301-1311
Arnold S, Sabara I, Bozhkov P, et al. Developmental pathways of somatic embryogenesis. Plant Cell, Tissue and Organ Culture, 2002, 69: 233-249
Arnholdt-Schmit B. Physiological aspects of genome variability in tissue culture. II. Growth phase-dependent quantitative variability of repetitive BstN I fragments of primary cultures of Daucus carota L. Theor. Appl. Genet, 1995, 91:816-823
Baker CM, Wetzstein H. Influence of auxin type and concertration on peanut somatic embryogenesis. Plant Cell, tissue and Organ Culture, 1994, 36: 361-368
Baker CM and Wetzstein HY. Repetitive somatic embryogenesis in peanut cotyledon cultures by continual exposure to 2, 4-D. Plant Cell, tissue and Organ Culture, 1995, 40: 249-254
Balestrazzi A, Bernacchia G, Pitto L, et al. Spatial expression of DNA topoisomeraseI genes during cell proliferation in Daucus carota. Eur J Histochem, 2001, 45: 31-38
Baudino S, Hansen S, Brettschneider R, et al. Molecular characterization of two novel maize LRR receptor-like kinases, which belong to the SERK gene family. Planta, 2001, 213: 1-10
Beyer EM. Mechanism of action of ethylene:biological activity of deuterated ethylene and evidence against isotopic exchange and cis-transisomerisation. Plant Physiol, 1979, 63: 169-173
Bolandi AR, Branchard M, Alibert G, Gentzbittel L, Berville A and Sarrafi A. Combining ability analysis of somatic embryogenesis from epidermis layers in sunflower. Theor. Appl. Genet, 2000, 100: 621-624
Boxtel JV and Berthouly M. High frequency somatic embryogenesis from coffee leaves. Plant Cell, Tissue and Organ Culture, 1996, 44: 7-17
Cangahuala-Inocente GC, Steiner N, Santos M, Guerra MP. Morphohistological analysis and histochemistry of Feijoa sellowiana somatic embryogenesis. Protoplasma, 2004, 224: 33-40
Charrière F, SottaB, Miginiac E, Hahne G. Induction of adventitious shoots or somatic embryos on in vitro cultured zygotic embryos of Heli anthus annuus: Variation of endogenous hormone levels. Plant Physiol Biochem, 1999, 37: 751-757.
Che P, Gingerich DJ, Lall S, Howel SH. Global and Hormone-Induced Gene Expression Changes during Shoot Development in Arabidopsis. Plant Cell, 2002, 14(11): 2771-2785.
Cheliak WM, Klimaszewska K. Genetic variation in somatic embryogenic response in open-pollinated families of black spruce. Theor Appl Genet, 1991, 82: 185-190
Chengalrayan K, Mhaske VB, Hazra S. High frequency of conversion of abnormal peanut somatic embryo embryogenesis. Plant cell Rep, 1997, 16: 783-786
Choi YE, Jeong JH. Dormancy induction of somatic embryos of Siberian ginseng by high sucrose concentrations enhances the conservations of hydrated artificial seeds and dehydration resistance. Plant Cell Rep, 2002, 20 : 1112-1116
Choi YE, Kim JW, Soh WY. Somatic embryogenesis and plant regeneration from suspensioncultures of Acanthopanax koreanum Nakai. Plant Cell Reports, 1997, 17: 84-88
Conner AJ, Falloon PG. Osmotic versus nutritional effects when rooting in vitro asparagus minicrown on high sucrose media. Plant Sci, 1993, 89: 101-106
Corredoira E, Valladares S, Vieitez AM. Morphohistological analysis of the origin and development of somatic embryos from leaves of mature Quercus robur. In Vitro Cellular & Developmental Biology Plant, 2006, 42 (6): 525-533
Cuming AC, Lane BG. Protein synthesis in imbibing wheat embryos. Eur J Biochem, 1979, 99: 217-224
Daveletova S, Meszaros T, Miskolczi P, et al. Auxin and heat shock activation of a novel member of the calmodulin like domain protein kinase gene family in cultured alfalfa cells. J Exp Bot, 2001, 52: 215-221
Dharmasiri N, Dharmasiri S, Estelle M. The F-box protein TIR1 is an auxin receptor. Nature, 2005a, 435 (7041): 441-445.
Dharmasiri, N, Dharmasiri S, Weijers D, et al. Plant development is regulated by a family of auxin receptor F box proteins. Dev Cell, 2005b, 9: 109-119
Dillen W, Dijkstra I, Oud J. Shoot regeneration in long-term callus cultures derived from mature flowering plants of Cyclamen persicum Mill. Plant Cell Reports, 1996, 15: 545-548
Dohm A, Schwenkel HG, Grunewaldt J. Histological Analysis of the in-Vitro Regeneration in Cyclamen-Persicum Mill. Gartenbauwissenschaft, 1991, 56: 58-65
Dong JZ, Dunstan DI. Characterization of three heat-shock-protein genes and their developmental regulation during somatic embryogenesis in white spruce [picea glauca(moench) voss]. Planta, 1996, 200: 85-91
Dong J, Dunstan DI. Expression of abundant mRNAs during somatic embryogenesis of white spruce. Planta, 2000, 199: 459-466
Dong JZ, Perras MR, Abrams SR, Dunstan DI. Induced gene expression following ABA uptake in embryogenic suspension cultures of picea glauca.Plant Physiol.Plant Physiol Biochem, 1996, 34: 579-587
Dougall DK, Verma DC. Growth and embryo formation in wild carrot suspension culture with ammoniumion as a sole nitrogen source. In Vitro, 1978, 14: 180-182.
Du L, Zhou S, Bao MZ. Effect of plant growth regulators on direct somatic embryogenesis incamphor tree ( Cinnamomum camphora L.) from immature zygotic embryos and embryogenic calli induction. For. Stud. China, 2007, 9(4): 267-271
Dusits D, Bogre L, Gyorgye Y. Molecular and cellular approaches to the plant embryo development from somatic cells in vitro. J Cell Sci, 1991, 99(3): 473-482
Ellen WH, Weining T, Karl WN, et al. Expression and maintenance of embryogenic potential is enhanced through constitutive expression of AGAMOUS-Like 15. Plant Physiol, 2003, 133(2): 653-663.
Feher A, Pasternak TP, Dudits D. Transition of somatic plant cell to an embryogenic state. Plant Cell, Tissue and Organ Culture, 2003, 74: 201-228
Fireozabady E, DeBoer D L. Plant regeneration via somatic embryogenesis in many cultivars of cotton (Gossypium hirsutum L.). In Vitro Cell Dev. Biol, 1993, 29: 166-173
Fowler M R, Ong LM, Russinova E, et al. Early changes in gene expression during direct somatic embryogenesis in alfalf are veiled by RAP-PCR. J Exp Bot, 1998, 49: 249-253
Furukawa K, Kakihara F, Kato M. Somatic embryos produced from aseptic seedlings of wild Cyclamen species. Journal of Society of High Technology in Agriculture, 2002, 14(2): 71-80
Furukawa K, Kakihara F, Kato M. Somatic embryogenesis in seven wild Cyclamen species. Journal of Society of High Technology in Agriculture, 2001, 13(4): 270-278
Fuminori K, Shigeo T. Regulation by Vrn-1/Fr-1chromosomal intervals of CBF-mediated Cor/Leagene expression and freezing tolerance in common wheat. Journal of Experimental Botany, 2005, (3): 887-895
Fischer-Igisics C, Sundgerg B, Neuhaus G, Jones AM. Auxin distribution and transport during embryonic pattern formation in wheat. Plant J, 2001, 26(2): 115-129
Filonova L, Bozhkov P, von Arnold S. Developmental pathway of somatic embryogenesis in Picea abies as revealed by time-lapse tracking. J. Exp. Bot, 2000, 51: 249-264
Gaj MD. Direct somatic embyogenesis as a rapid and efficient system for in vitro regeneration of Arabidopsis thaliana (L.) Heynh. Plant Cell, Tissue and Organ Culture, 2001, 64: 39-46
Giroux RW, Pauls KP. Characterization of somatic embryogenesis-related cDNAs from alfalfa (MedicagosativaL). Plant Mol Biol, 1997, 33: 393-404
Gleddie S, Keller W, Setterfield G. Somatic embryogenesis and plant regeneration from leaf explants and cell suspensions of Solanum melongena (egg plant). Can J Bot, 1983, 61: 656-666
Gonzilez-Benito ME, Alderson PG. In vitro culture of Alstroemeria somatic embryos. Phyton, 1992, 53: 95-101
Gray WM, del Pozo JC, Walker L, et al. Identification of an SCF ubiquitin–ligase complex required for auxin response in Arabidopsis thaliana. Genes & Dev, 1999, 13(13): 1678-1691
Gupta PK, Pullman G. Method for reproducing coniferous plants by somatic embryogenesis using abscisic acid and osmotic potential variation. US Patent, 5036007, 1991.
Haccis B. Question of unicellar origin on non-zygotic embryos in callus cultures. Phytomotphology, 1978, 28: 74-81
Halper NW, Jensen AJ. Ultrastructural changes during growth and embryogenesis in carrot cell culture. Ultra Res., 1967, 18: 428 -443
Harada T, Yakuwa T. Studies on the morphogenesis of Asparagus VI. Effect of sugar on callus and organ formation in the in vitro culture of shoot segments of the seedlings. J Fac Agr Hokkaido Univ, 1983, 61: 307-314
Hatzopoulos P, Fong F, Sung ZR. Absicsic acid regulation of DC8 a carrot embryogenic gene. Plant Physiol, 1990, 94: 690-695
Hecht V, Vielle-Calzada JP, Hartog MV , Schmidt EDL, Boutilier K, Grossniklaus U, and Vries SC. The Arabidopsis somatic embryogenesis receptor kinase-1 gene is expressed in developing ovules and embryos and enhances embryogenic competence in culture. Plant Physiol, 2001, 127: 803-816
He DG, Tanner GT, Scott KJ. Somatic embryogenesis and morphogenesis in callus derived from the epiblast of immature embryo of wheat. Plant Sci, 1986, 45: 119-124
Henry Y, Vain P, Buyser J. Genetic analysis of in vitro plant tissue culture responses and regeneration capacities. Euphytica, 1994, 79: 45-48.
Hess JR, Carman JG. Embryogenic competence of immature wheat embryos: genotype, donor plant, environment and endogenous hormone concentrations. Crop Sci, 1998, 38: 249-253
Higashi K, Shiota H, Kamada H. Patterns of expression of the genes for glutamine synthetase isoforms during somatic and zygotic embryogenesis in carrot. Plant Cell Physiol, 1998, 39: 418-424
Hohe A, Winkelmann T, Schwenkel HG. CO2 accumulation in bioreactor suspension cultures of Cyclamen persicum Mill and its effect on cell growth and regeneration of somatic embryos.Plant Cell Reports, 1999a, 18(10): 863-867
Hohe A, Winkelmann T, Schwenkel HG. The effect of oxygen partial pressure in bioreactors on cell proliferation and subsequent differentiation of somatic embryos of Cyclamen persicum. Plant Cell Tissue Org Cult, 1999b, 59: 39-45
Hohe A, Winkelmann T, Schwenkel HG. Development of somatic embryos of Cyclamen persicum Mill in liquid culture. Gartenbauwissenschaft, 2001, 66: 219-224
Hu H, Xiong L, Yang Y. Rice SERK1 gene positively regulates somatic embryogenesis of cultured cell and host defense response against fungal infection. Planta, 2005, 222(1): 107-117
Hutchinson MJ, Saxena PK. Acetylsalicylic acid enhances and synchronizes thidiazuron-induced somatic embryogenesis in geranium (Pelargonium x hortorum Bailey) tissue cultures. Plant Cell Reports, 1996, 15 (7) : 512-515
Ikeda-Iwai M, Satoh S, Kamada H. Establishment of a reproducible tissue culture system for the induction of Arabidopsis somatic embryos. Exp Bot, 2002, 53: 1575-1580
Imamara J, Hiroshi H. Effects of abscisic acid and water stress on the embryo and plantlet formation in anther culture of Nicotiana tabacum cultivar S amsun Z. Pflanzenphysiol, 1980, 100: 285-289
Ito Y, Takaya K, Kurata N. Expression of SERK family receptor-like protein kinase gene in rice. Biochimica et Biophysica Acta, 2005, 1730(3): 253-258
Jelaska S. Recent development in somatic embryogenesis. Acta Pharmaceutica (Zagreb), 1994, 44(3): 387-406
Jianru Z, Qi-Wen N, Giovanna F, Nam HC. The WUS gene promotes vegetative-to-embryonic transition in Arabid. Plant J, 2002, 30(3): 349-359
Jianru Z, Niu QW, Yoshihisa L, Chua NH. Marker-free transformation: increasing transformation frequency by the use of regeneration-promoting genes. Curr Opi in Biol, 2002, 13(2): 173-180
Jiménez VM, Bangerth F. Endogenous hormone levels in explants and in embryogenic and non-embryogenic cultures of carrot. Physiol Plant, 2001a, 111: 389-395
Jimenez VM, Bangerth F. Endogenous hormone concentrations and embryogenic callus development in wheat. Plant Cell, Tissue and Organ Culture, 2001b, 67: 37-46
Jordy MN. Seasonal variation of organogenetic activity and reserves allocation in the shoot apex of Pinus pinaster Ait. Ann Bot, 2004, 93: 25-37
Kamada H, Harada H. Studies on the organogenesis in carrot tissue culture.Ⅰ. Effects of growth regulators on somatic embryogenesis and root formation. Z. P flonzenphysiol, 1979, 91: 255-266
Kamada H, Ishikawa K, Saga H, Harada H. Induction of somatic embryogenesis in carrot by osmotic stress. Plant Tissue Cult Lett, 1993, 10: 38-44
Kamo K, Jones B, Castillon J, Bolar J, Smith F. Dispersal and size fractionation of embryogenic callus increases the frequency of embryo maturation and conversion in hybrid tea roses. Plant Cell Reports, 2004, 22 (11): 787-792
Kaparakis G and Alderson PG. Role for Cytokinins in Somatic Embryogenesis of Pepper (Capsicum annuum L.)?. J Plant Growth Regul, 2008, 27: 110-114
Karim MZ, Yokota S, Azad M, et al. Relationship between starch accumulation and organ development at the different growth stages of callus in Kihada (Phellodendron amurense Rupr.). Plant Biotechnology, 2006, 23: 239-245
Kikuchi A, Sanuki N, Higashi K, et al. Abscisic acid and stress treatment are essential for the acquisition of embryogenic competence by carrot somatic cells. Planta, 2006, 223: 637-645
Kim B, Remko O, Vijay KS, et al. Ectopic expression of BABY BOOM triggers a conversion from vegetative to embryonic growth. Plant Cell, 2002, 14(8): 1737-1749
King SW. DNA sequence of an ABA-responsive gene (rab 15) from water-stress wheat roots. Plant Mol.Biol, 1992, 18: 119-121
Kitamiya E, Suzuki S, Sano T, et al. Isolation of two genes that were induced upon initiation of somatic embryogenesis on carrot hypocotyls by high concentrations of 2,4-D. Plant Cell Rep, 2000, 19: 551-557
Kiviharju E, Tuominen U, Tormala T. The effect of explant material on somatic embryogenesis of Cyclamen persicum Mill. Plant Cell, Tissue and Organ Culture, 1992, 28(2): 187-194
Kiyosue T and Shinozaki K. Cloning of a carrot cDNA for a member of the family of ADP- ribosylationfactors (ARFs) and characterization of the binding of nucleotides by its product after expression in e- coli. Plant Cell Physiol, 1995, 36: 849-856
Kiyosue T, Kamada H and Harada H. Induction of Somatic embryogenesis from carrot Seeds by Hypochlorite Treatment Plant Tissue Culture Letters, 1989, 6(3): 138-143
Kiyosue T, Kamada H, Harada H. Induction of somatic embryogenesis by salt stress in carrot. Plant Tissue Cult Lett, 1989, 6: 162-164
Kiyosue T, Nakajima M, Yamaguchi I, et al. Endogeneous levels of abscisic acid in embryogenic cells, non-embryogenic cells and somatic embryos of carrot (Daucus carota L.). Biochem Physiol Pflanzen, 1992b, 188: 343-347
Kiyosue T, Takano K, Kamada H, Harada H. Induction of somatic embryogenesis in carrot by heavy metal ions. Can J Bot, 1990, 68: 2021-2033
Kiyosue T, Yamaguchi-Shinozaki K, Shinozaki K, et al. Partial amino-acid sequence of ECP31, a carrot embryogenic-cell protein, and enhancement of its accumulation by abscisic acid in somatic embryos. Planta, 1992a, 186: 337-342
Kiyosue T, Yamaguchi-Shinozaki K, Shinozaki K, et al. Isolation and characterization of a cDNA that encodes ECP31, an embryogenic-cell protein from carrot. Plant Mol Biol, 1992, 19: 239-249
Kong L, Yeung EC. Effects of silver nitrate and polyethylene glycol on white spruce (Picea glauca) somaticembryo development: enhancing cotyledonary embryo formation and endogenous ABA content. Physiol Plant, 1995, 93: 298-304
Kochba J , Spiegel-Roy P, Saad S, et al. Stimulation of embryogenesis in Citrus tissue culture by galactose. Naturwissenschaften, 1978, 65: 261-262
Kopertekh LG and Butenko RG. Naturally occurring phytohormones in wheat explants as related to wheat morphogenesis in vitro. Rus J Plant Physiol., 1995, 42: 488-491
Kreuger M, Postma E, Brouwer Y et al. Somatic embryogenesis of Cyclamen persicum in liquid medium. Physiol Plant, 1995, 94: 605-612
Kuklin AD, Denchev PD, Atanassov AI, Scragg AH. Alfalfa embryo production in airlift vessels via direct somatic embryogenesis. Plant Cell, Tissue and Organ Culture, 1994, 38: 19-23
Landschulz WH, Johnson PF, Maknight SL. The leucine zipper: a hypothetical structure common to a new class of DNA binding proteins. Science, 1988, 240(4860): 1759-1764
Laurent L, Philippe V, Francoise C, et al. Effect of abascisic acid and high concentrations of PEG on Hevea brasiliensis somatic embryos development. Plant Sci, 1997, 124(2): 183-191
Leal I, Misra S. Molecular cloning and characterization of alegumin-like storage protein cDNA of Douglasfir seeds. Plant Mol Biol, 1993,21: 709-715
Lee KP and Lee DW. Somatic embryogenesis and plant regeneration from seeds of wild Dicentra spectabilis (L.) LEM. Plant Cell Rep, 2003, 22: 105-109
Libbenga KR, Mennes AM. Plant hormones. In: Davies PJ (ed). Physiology, Biochemistry andMolecular Biology. Dordrecht, the Netherlands: Kluwer Academic Pub, 1995, 270-275
Li BC and Wolyn DJ. Abscisic acid and ancymidol promote conversion of somatic embryos to plantlets and secondary embryogenesis in Asparagus officinalis L. In Vitro Cell Dev. Biol.-Plant, 1996b, 32: 223-226
Li BC and Wolyn DJ. Temperature and genotype affect asparagus somatic embtyogenesis. In Vitro Cell Dev. Biol.-Plant, 1996a, 32: 136-139
Li MH, Steven EU, Andris K, et al. Improvement of anther culture methods or doubled haploid production in barley breeding. Plant Cell Rep, 1993, 12: 328-334
Lindzen E, Choi JH. A carrot cDNA encoding an atypical protein kinase homologous to plant calcium-dependent protein kinases.Plant Mol Biol, 1995, 28: 785-797
Liu CQ, Xia XL, Yin WL, Huang LC, Zhou JH. Shoot regeneration and somatic embryogenesis from needles of redwood (Sequoia sempervirens (D.Don.) Endl.) Plant Cell Rep, 2006, 25(7): 621 -628
Lo Shiavo F, Filippini F, Cozzani F, et al. Modulation of auxin-binding proteins in cell suspension I. Different responses of carrot embryo cultures.Plant Physiol, 1991, 97: 60-64
Lyngved R, Renaut J, Hausman JF, et al. Embryo-specific Proteins in Cyclamen persicum Analyzed with 2-D DIGE. J Plant Growth Regul, 2008, 27: 353-369
Mahesh-Wari SC, Tyagi AK, Malhotra K. Induction of haploidy from pollen grains in angiosperms the current status. Theor Appl Genet, 1980, 58: 193-206
Mamiya K, Sakamoto Y. Effects of sugar concentration and strength of basal medium on conversion of somatic embryos in Asparagus officinalis L. Sci Hort, 2000, 84: 15-26
Manickam A, vanDamme EJM, Kalaiselvi K, et al. Isolation and cDNA cloning of an Em-like protein from mungbean (Vigna radiata) axes. Physiol Plantarum, 1996, 97(3): 524-530
Manisha Vand BansalYK. Somatic embryogenesis and plantlet regeneration in semul (Bombax ceiba). Plant Cell, Tissue and Organ Culture, 2004, 79: 115-118
Mathew MM and Philip VJ. Somatic embryogenesis versus zygotic embryogenesis in Ensete superbum. Plant Cell, Tissue and Organ Culture, 2003, 72: 267-275
MedicagoT. Root-forming and embryogenic cultures. Plant Physiol, 2003, 133(1): 218-230.
Meskaoui AE, Desjardins Y, Tremblay FM. Kinetics of ethylene biosynthesis and its effects during maturation of white spruce somatic embryos. Physiol Plant, 2000, 109: 333-342
Michalczuk L, Cooke TJ, Conen JD. Auxin levels at different stages of carrot somatic embryogenesis. Phytochemistry, 1992, 31: 1097-1103
Michalczuk L, Ribnicky DM, Cooke TJ, et al. Regulation of indole-acetic acid biosynthetic pathway in carrot cell cultures. Plant Physiol, 1992, 100: 1346-1353
Michler CH, Lineberger RD. Effects of light abscisic levels in carrot suspension cultures. Plant Cell Tiss Org Cult, 1987, 11: 189-207
Miku?a A, Rybczyński JJ, Skierski J,et al. Somatic embryogenesis of Gentiana genus IV: Characterisation of Gentiana cruciata and Gentiana tibetica embryogenic cell suspensions. In: Hvolsef-Eide AK and Preil W, eds. Liquid Culture Systems for In Vitro Plant Propagation, Netherlands: Springer, 2005. 345-358
Mithila J, Hall JC, Victor JMR, Saxena PK. Thidiazuron induces shoot organogenesis at low concentrations and somatic embryogenesis at high concentrations on leaf and petiole explants of African violet (Saintpaulia ionantha Wendl.). Plant Cell Rep, 2003, 21: 408-414
Mohanna AM, Karam NS. In vitro propagation of wild Cyclamen persicum Mill from seedling tissue. Acta Horticilturae, 2000,(530): 243-250
Motoike SY, Skirvin RM, Norton MA, et al. Somatic embryogenesis and long term maintenance of embryogenic lines from fox grapes. Plant Cell Tissue Org Cult, 2001, 66: 121-131
Murasaki K and Tsurushima H. Improvement on clonal propagation of cyclamen in vitro by the use of etiolated petioles. Acta Hort, 1988, 226: 721-724
Murashige T, Skoog F. A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol Plant, 1962, 15: 473-497
Nakagawa H, Saijyo T, Yamauchi N, et al. Effects of sugars and abscisic acid on somatic embryogenesis from melon (Cucumis melo L.) expanded cotyledon. Sci Hort, 2001, 90: 85-92
Nishiwaki M, FujinoK, KodaY, Masuda K, Kikuta Y. Somatic embryogenesis induced by the simple application of abscisic acid to carrot (Daucus carota L.) seedling in culture. Planta, 2000, 211: 756-759
Nitsch C, Norreel B. Effect d′u m choc thermique sue le pouvoir embryogene dupollen Datura innoxia culture dans I′anther ouisolede I′anthere. C R Acad Sci Paris, 1973, 276: 303-306
Nokta M, EatonD, Steinsland OS, Albrecht T. Ca 2+ responses in cytomegalovirus-infected fibroblasts of human origin. Virology, 1987, 157: 259-267
Nolan KE, Irwanto RR, Rose RJ. Auxin up-regulates MtSERK1 expression in both medicago truncatula root-forming and embryogenic cultures. Plant Physiol, 2003, 133(1): 218-230
Osuga K, Masuda H, Komamine A. Synchronization of somatic embryogenesis at high frequency using carrot suspension cultures: model systems and application in plant development. Methods in Cell Science, 1999, 21: 129-140
Overvoorde PJ, Grimes HD. The role of calcium and calmodulin in carrot somatic embryogenesis. Plant Cell Physiol, 1994, 35: 135-144
Owen HR, Miller AR. Haploid plant regeneration from anther cultures of three North American cultivars of strawberry ( Fragaria ananassa Duch). Plant Cell Rep, 1996, 15(12): 905-909
Padmanabhan K, Cantliffe DJ, Koch KE. Auxin-regulated gene expression and embryogenic competence in callus cultures of sweet potato, Ipomoeabatata(L.)Lam. Plant Cell Rep, 2001, 20: 187-192
Quiroz-Figueroa FR, Rojas-Herrera R, Galaz-Avalos RM, Loyola-Vargas VM. Embryo Production through somatic embryogenesis can be used to study cell differentiation in plants. Plant Cell, Tiss and Organ Culture, 2006, 86(3): 285-301
Rensing SA, Lang D, Schumann E, Reski R, Hohe A. EST sequencing from embryogenic Cyclamen persicum cell cultures identifies a high proportion of transcripts homologous to plant genes involved in somatic embryogenesis. J Plant Growth Regul, 2005, 24: 102-115
Ribas LLF, Guerra MP, Zanette F, Kulchetscki L. Somatic embryogenesis in Aspidosperma polyneuron Mull. Arg. In: Jain SM, Gupta PK, Newton RJ, eds. Somatic Embryogenesis in Woody Plants. Dordrecht, Boston, London: Kluwer Academic Publishers, 2000. 509-537
Rougtan JP, Latche A, Fallot J. Inhibition of ethylene production and stimulation of carrot somatic embryogenesis by salicylic acid. Biol Plant, 1990, 32: 273-278
Ruffoni B, Semeria L, Profumo P. Cyclamen persicum Mill. Somatic embryos developed in suspension cultures: histological analysis and conversion to plant. Acta Horticulturae, 2000, 520: 83-90
Ruffoni B. Embryogenic suspension cultures of cyclamen for the production of artificial seeds. Colture Protette, 1998, 27(11): 91-96
Russinova E, Slater A, Atanassov AI, et al. Cloning novel alfalfa cyclin sequences a RACE- PCR approach. Cellular and Molecular Biology, 1995, 41: 703-714
Sandra LS, Linda WK, Kelly MY, et al. LEAFY COTYLEDON2 encodes a B3 domain transcription factor that induces embryo development. Plant Biol, 2001, 20(98): 11806-11811
Santos MDO, Romano E, Yotoko KSC, et al. Characterisation of the cacao somatic embryogenesis receptor-like kinase (SERK) gene expressed during somatic embryogenesis. Plant Sci, 2005, 168(3): 723-729
Sasaki K, Shimomura K, Kamada H, Harada H. IAA metabolism in embryogenic and non-embryogenic carrot cells. Plant Cell Physiol, 1994, 35: 1159-1164
Sato S, Toya T, Kawahara R, et al. Isolation of a carrot gene expressed specifically during early-stage somatic embryogenesis. Plant Mol Biol, 1995, 28: 39-46
Savona M, Ruffoni B, Falasca G, et al (2007). NCBI GenBank database, #EF661828, #EF672247, #DQ350614. http://www.ncbi.nlm.nih.gov
Schmidt ED, Guzzo F, Toonen MAJ, et al. A leucine-rich repeat containing receptor-like kinase marks somatic plant cell competent to form embryos. Development, 1997, 124(10): 2049-2062
Schmidt Th, Ewald A, Seyring M, Hohe A. Comparative analysis of cell cycle events in zygotic and somatic embryos of Cyclamen persicum indicates strong resemblance of somatic embryos to recalcitrant seeds. Plant Cell Rep, 2006, 25: 643-650
Schwenkel HG, Grunewaldt J. Somaclonal Variation in Cyclamen Persicum Mill after in Vitro Mass Propagation. Gartenbauwissenschaft, 1991, 56, 16-20
Schwenkel HG, Winkelmann T. Plant regeneration via somatic embryogenesis from ovules of Cyclamen persicum Mill.Plant Tiss Cult Biotechnol, 1998, 4: 28-34
Sengupta C, Raghavan V. Somatic embryogenesis in carrot cell suspension.Ⅰ. Pattern protein and nucleic acid synthesis. J Exper Bot, 1980, 30: 247-258
Seyring M, Hohe A. Induction of desiccation tolerance in somatic embryos of C. persicum MILL. J Hortic Sci Biotechol, 2005, 80: 65-69
Shah K, Gadella TW, Jrvan EH, et al. Subcellular localization and oligomerization of the Arabidopsis thaliana somatic embryogenesis receptor kinase1 protein. J Mol Biol, 2001, 309: 641-655
Sharp W R, Sondahl M R, Caldas L S, et al. The physiology of in vitro asexual embryogenesis. Hort Rev, 1980, 2: 268 -310
Shibli RA, Ajlouni MM. Somatic embryogenesis in the endemic black iris. Plant Cell Tiss OrgCult, 2000, 61: 15-21
Shimada T, Hirabayashi T, Endo T, et al. Isolation and characterization of the somatic embryogenesis receptor-like kinase gene homologue (CitSERK1)from Citrus unshiu Marc. Sci Hortic, 2005, 103(2): 233-238
Smeeken S. Sugar-induced signal transduction in plants. Annu Rev Plant Physiol Plant Mol Biol, 2000, 51: 49-81
Sofiari E, Raemakers CJJM, Kanju E, et al. Comparison of NAA and 2, 4-D induced somatic embryogenesis in Cassava. Plant Cell, Tissue and Organ Culture, 1997, 50: 45-56
Somleva MN, Schmidt EDL, Vries SC. Embryogenic cells in Dactylis glomerata L.(Poaceae) explants identified by cell tracking and by SERK expression. Plant Cell Rep, 2000, 19(1): 718-726
Songstad DD. Effect of aminocyclopropane carboxylic acid, silver nitrate and norbornadiene on plant regeneratin from maize callus cultures. Plant Cell Rep, 1988, 7: 262-265
Stange L, Osborne D J. Cell specificity in auxin and ethylene induced‘supergrowth’in Riella helicophylla. Planta, 1988, 175: 341 - 347.
Steward FC, Mapes MO, Hears K. Growth and organized development of cultured cells. II. Growth and division of freely suspended cells. Am.J.Bot, 1958, 45: 705-708
Stitn S, Jacobsen HJ. Marker proteins for embryogenic differentiation patterns in pea callus. Plant Cell Reports, 1987, 6: 50 -54
Suprasanna P, Desai NS, Nishanth G, et al. Differential gene expression in embryogenic, non-embryogenic and desiccation induced cultures of sugarcane. Sugar Technol, 2004, 6(4): 305-309
Takamura T, Miyajima I, Matsuo E. Somatic embryogenesis of Cyclamen periscum Mill. 'Anneke' from aseptic seedlings. Plant Cell Reports, 1995, 15(2): 22-25
Takamura T, Nagita Y and Tanaka M. Effects of temperature on in vitro plant regeneration through somatic embryogenesis in Cyclamen persicum Mill. Hot Res, 2003, 2(1): 25-28
Takamura T. and Tanaka M. Somatic embryogenesis from the etiolated petiole of cyclamen (Cyclamen persicum Mill). Plant Tissue Culture Letters, 1996, 13(1): 43-48
Tetteroo FAA, Bino RJ, Bergervoet JHW et al. Effect of ABA and slow drying on DNA replication in carrot (daucus carota) embryoids. Physiologia Plantarum, 1995, 95: 154-158
ThiL T, Pleschka E. Somatic embryogenesis of some Daucus species influenced by ABA. J ApplBot Food Qual, 2005, 79: 1- 4
Thomas C, Meyer D, Himber C, Steinmetz A. Spatial expression of a sunflower SERK gene during induction of somatic embryogenesis and shoot organogenesis. Plant Physiol Biochem, 2004, 1(42): 35-42
Thomas TL. Gene expression during plant embryogenesis and germination: An overview. Plant Cell, 1993, 5(10): 1401-1410
Thorpe TA, Murashige T. Some hischemical changes underlying shoot initiation in tabacco callus cultures. Can J Bot, 1970, 48: 277 -285
Tisserat B, Murashige T. Effects of ethephon, ethylene and 2, 4-dichlorophenoxyacetic acid on asexual embryogenesis in vitro. Plant Physiol, 1977, 60(3): 437-439
Toivonen L, Laakso S, Rosenqvist H. The effects of temperature on growth, indole alkaloid accumulation and lipid composition of Catharanthus roseus cell suspension cultures. Plant Cell Rep, 1992, 11: 390-394
TornéJM, Moysset L, Claparols I, et al. Photo control of somatic embryogenesis and polyamine content in Araujia sericifera petals. Physiol Plant, 1996, 98: 413-418
Tremblay L,Tremblay FM. Effect of gelling agents ,ammonium nitrate and light on the development of Picea mariana (Mill) B. S. P. (black spruce) and Picea rubens Sarg. (red spruce) somatic embryos. Plant Sci, 1991, 77: 233-242
Verhagen SA, Wann SR. Norway spruce somatic embryogenesis High frequency initiation from light cultured mature embryos. Plant Cell Tiss Org Cult, 1989, 16: 103-111
Verma DC, Dougall DK. Influence of carbohydrates on quantitative aspects of growth and embryo formation in wild carrot suspension cultures. Plant Physiol, 1977, 59: 81- 85
Vikrant and Rashid A. Direct as well as indirect somatic embryogenesis from immature (unemerged) inflorescence of a minor millet Paspalum scrobiculatum L. Euphytica, 2001, 120: 167-172
Von Aderkas P, Thompson RG, Zaki M, Benkrima L. Somatic embryogenesis of western larch (Larix occidentalis). In: Bajaj YPS ed. Biotechnology in Agriculture and Forestry Somatic Embryogenesis and Synthetic Seeds, Berlin: Springer-Verlag, 1995. 378-387
Wang YH and Bhalla PL. Somatic embryogenesis from leaf explants of Australian fan flower Scaevola aemula R. Br. Plant Cell Rep, 2004, 22: 408-414
Weatherwax SC, Ong MS, Degenhardt J, et al. The interaction of light and abscisic acid in the regulation of plant gene expression. Plant Physiol, 1996, 111: 363-370
Welander M, Maheswaran G. Shoot regeneration from leaf explants of dwarfing apple rootstocks. J Plant Physiol, 1992, 140: 223-228
Wetherell DF, Dougall DK. Sources of nitrogen supporting growth and embryogenesis in cultured wild carrot issue. Physiol Plant, 1976, 37: 97-103
Wicart G, Mouras A, Lutz A. Histological study of organogenesis and embryogenesis in Cyclamen persicum Mill. Tissue cultures: evidence for a single organogenesis pattern. Protoplasma, 1984, 119(3): 159-167
Wilson SM, Thorpe TA . Somatic embryogenesis in Picea glauca (white spruce), P. engelmannii (Engelman spruce) and P. engelmannii complex (interior spruce). In: Mohan J S, Gupta P K, Newton R J eds. Somatic Embryogenesis in Woody Plants, Vol. 1. Dordrecht, The Netherlands: Kluwer Academic Publishers, 1995. 37-54
Winkelmann T, Heintz D, van Dorsselaer A, Serek M, Braun HP. Proteomic analysis of somatic and zygotic embryos of Cyclamen persicum Mill. reveal new insights into seed and germination physiology. Planta, 2006, 224: 508-519
Winkelmann T, Hohe A, Schwenkel HG. Establishing embryogenic suspension cultures in Cyclamen persicum 'Purple Flamed'. Advances in Horticultural Science, 1998, 12(1): 25-30
Winkelmann T, Meyer L, Serek M. Desiccation of somatic embryos of Cyclamen persicum Mill. J Horticult Sci Biotechnol, 2004, 79 (3): 479-483
Winkelmann T, Schwenkel H G, Hohe A. Development of somatic embryos of Cyclamen persicum Mill in liquid culture. Gartenbauwissenschaft, 2001, 66(5): 219-224
Winklemann T, Serek M. Genotypic differences in callus formation and regeneration of somatic embryos in Cyclamen persicum Mill. Euphytica, 2005, 144: 109-117
Xiang TH, Liang HM, Zhong HX. Physiological and biochemical changes during the establishment of embryogenic cell suspension cultures of rice (Oryza satira L.). II: changes of the contents of amino acid, polyamine and endohormone. ActaAgronomica Sinica, 1997, 23(3): 353-359
Yang H, Saitou T, Komeda Y, et al. Arabidopsis thaliana ECP63 encoding a LEA protein is located in chromosome 4. Gene, 1997, 184: 83-88
Yang H, Saitou T, Komeda Y, et al. Late embryogenesis abundant protein in Arabidopsis thaliana homologous to carrot ECP31. Physiol Plant, 1996, 98: 661-666
Yasuda H, Nakajiima M, Ito T, et al. Partial characterization of genes whose transcripts accumulate preferentially in cell clusters in the early stages of carrot somatic embryogenesis.Plant Mol Biol, 2001, 45: 705-712
Yasuda H, Sato T, Masuda H. Rapid and frequent embryogenesis from single cells of regenerated carrot plantlets. Biosci Biotechnol Biochem, 1998, 62: 1273-1278
Yeung EC, Stasolla C. Somatic embryogenesis-apical meristem formation and conversion. Korean J Plant Tiss Cult, 2000, 27: 253-258
Yeung EC, Stasolla C, Kong L. Apical meristem formation during zygotic embryo development of white spruce. Can J Bot, 1998, 76: 751-761
Zhu C, Kamada H, Harada H, et al. Isolation and characterization of a cDNA encoded an embryogenic cell protein-63 related to embryogenesis from carrot. Acta Bot Sin, 1997, 39: 1091-1098
Zimmerman JL. Somatic embryogenesis: a model for early development in higher plants. Plant Cell, 1993, 5: 1411-1423