刺槐Genomic-SSR与EST-SSR遗传差异性分析
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摘要
为在刺槐育种研究中合理选用不同来源的SSR分子标记,选取12个刺槐个体,试剂盒提取DNA后分别利用9对Genomic-SSR引物和9对EST-SSR引物进行扩增,并采取毛细管电泳检测扩增产物。利用所得条带信息及相关软件对2种SSR分子标记进行多态性、遗传相似系数相关性以及聚类等方面的比较分析。结果显示,刺槐Genomic-SSR平均检测到的条带数为6、Shannon多样性指数为1.3833、观测杂合度为0.5749、期望杂合度为0.6832;EST-SSR平均检测到的条带数为5.1、Shannon多样性指数为1.2711、观测杂合度为0.5648、期望杂合度为0.6526。由Genomic-SSR计算得到的个体间的遗传相似系数以及聚类结果与2种SSR标记综合计算得到的遗传相似系数和聚类结果更为相似。结果表明,刺槐Genomic-SSR与EST-SSR存在一定的遗传差异性,但差异并不显著;刺槐Genomic-SSR能更加准确地揭示基因型之间的遗传关系;刺槐EST-SSR具有相对较强的保守性。
The study aims at selecting the SSR molecular markers reasonably in the breeding of black locust.12 individuals of black locust were used as the materials, the total genomic DNA was extracted by kit. 9 Genomic-SSR and 9 EST-SSR primers were used for PCR amplification and the products were separated by capillary electrophoresis. Comparative analysis of polymorphism, correlation of genetic similarity coefficients and cluster analysis between Genomic-SSR and EST-SSR were conduct by related software according to the information of the electrophoretic bands. The average number of bands detected by Genomic-SSR was 6.0,Shannon index was 1.3833, observed heterozygosity was 0.5749 and expected heterozygosity was 0.6832. The average number of bands detected by EST-SSR was 5.1, Shannon index was 1.2711, observed heterozygosity was 0.5648 and expected heterozygosity was 0.6526. Genetic similarity coefficients and results of cluster analysis between individuals calculated from Genomic-SSR were more similar to the comprehensive results calculated from the two kinds of SSR markers. There are genetic differences between Genomic-SSR and ESTSSR of black locust, but the differences are not significant. Genomic-SSR of black locust can more accurately reflect genetic relationship of different genotypes. EST-SSR of black locust is relatively more conservative.
The study aims at selecting the SSR molecular markers reasonably in the breeding of black locust.12 individuals of black locust were used as the materials, the total genomic DNA was extracted by kit. 9 Genomic-SSR and 9 EST-SSR primers were used for PCR amplification and the products were separated by capillary electrophoresis. Comparative analysis of polymorphism, correlation of genetic similarity coefficients and cluster analysis between Genomic-SSR and EST-SSR were conduct by related software according to the information of the electrophoretic bands. The average number of bands detected by Genomic-SSR was 6.0,Shannon index was 1.3833, observed heterozygosity was 0.5749 and expected heterozygosity was 0.6832. The average number of bands detected by EST-SSR was 5.1, Shannon index was 1.2711, observed heterozygosity was 0.5648 and expected heterozygosity was 0.6526. Genetic similarity coefficients and results of cluster analysis between individuals calculated from Genomic-SSR were more similar to the comprehensive results calculated from the two kinds of SSR markers. There are genetic differences between Genomic-SSR and ESTSSR of black locust, but the differences are not significant. Genomic-SSR of black locust can more accurately reflect genetic relationship of different genotypes. EST-SSR of black locust is relatively more conservative.
引文
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[4]茹桃勤,李吉跃,张克勇,等.国外刺槐(Robinia pseudoacacia)研究[J].西北林学院学报,2005,20(3):102-108.
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[6] Tuskan G A, Gunter L E, Yang Z K, et al. Characterization of microsatellites revealed by genomic sequencing of Populus trichocarpa[J]. Canadian Journal of Forest Research, 2004, 34:85-93.
[7] Varshney R K, Graner A, Sorrells M E, et al. Genomicmicrosatellite markers in plants:Features and applications[J].Trends in Biotechnology, 2005, 23(1):48-55.
[8] Yin T M, Tuskan G A, Gunter L E, et al. Genome structure and emerging evidence of an incipient sex chromosome in Populus[J].Genome Research, 2008, 18:422-430.
[9] R?der M S, Korzun V, Wendehake K, et al. A microsatellite map of wheat[J]. Genetics,1998,149(4):2007-2023.
[10]周延清.DNA分子标记技术在植物研究中的应用[M].北京:化学工业出版社,2005.
[11] Mutegi S M, Muchugi A, Carsan S, et al. Genetic diversity of the African poplar(Populus ilicifolia)populations in Kenya[J]. Tree Genetics&Genomes,2016,12(4):1-7.
[12] Sun R, Lin F, Huang P, et al. Moderate Genetic Diversity and Genetic Differentiation in the Relict Tree Liquidambar formosana Hance Revealed by Genic Simple Sequence Repeat Markers[J].Frontiers in Plant Science,2016,7.
[13]胡晓辉,毛瑞喜,苗华荣,等.山东省46个花生品种SSR指纹图谱构建与遗传多样性分析[J].核农学报,2016,30(10):1925-1933.
[14]艾畅,徐立安,赖焕林,等.马尾松种子园的遗传多样性与父本分析[J].林业科学,2006,42(11):146-150.
[15]车卓,常宏,石菁,等.利用SSR分子标记鉴定玉米杂交种吉祥1号纯度[J].中国农学通报,2015,32(18):28-32.
[16]齐凤慧,孙宏冉,詹亚光.EST-SSR标记在水曲柳雌雄鉴定中的应用[J].西北植物学报,2015,35(3):472-479.
[17]吴静,成仿云,庞利铮,等.紫斑牡丹表型性状与SSR分子标记的关联分析[J].北京林业大学学报,2016,38(8):80-87.
[18]杨新泉,刘鹏,韩宗福,等.普通小麦Genomic-SSR和EST-SSR分子标记遗传差异及其与系谱遗传距离的比较研究[J].遗传学报,2005,32(4):406-416.
[19]常玮,赵雪,李侠,等.大豆EST-SSR标记开发及与Genomic-SSR的比较研究[J].中国油料作物学报,2009,31(2):149-156.
[20] Wen M, Wang H, Xia Z, et al. Development of EST-SSR and genomic-SSR markers to assess genetic diversity in Jatropha curcas L.[J]. Bmc Research Notes,2010,3(1):1-8.
[21] Lian C, Hogetsu T. Development of microsatellite markers in black locust(Robinia pseudoacacia)using a dual-supression-PCR technique[J]. Molecular Ecology Notes,2002,2(3):211-213.
[22] Lian C, Oishi R, Miyashita N, et al. High somatic instability of a microsatellite locus in a clonal tree, Robinia pseudoacacia[J].Theoretical and Applied Genetics,2004,108(5):836-841.
[23] Mishima K, Hirao T, Urano S, et al. Isolation and characterization of microsatellite markers from Robinia pseudoacacia L.[J].Molecular Ecology Resources,2009,9(3):850-852.
[24] Smith J M, Haigh J. The hitch-hiking effect of a favourable gene[J].Genetical Research,1974,23(1):391-403.
[25] Van K, Hwang E Y, Kim M Y, et al. Discovery of single nucleotide polymorphisms in soybean using primers designed from ESTs[J].Euphytica,2004,139(2):147-157.
[26]贾继增,张正斌.小麦21条染色体RFLP作图位点遗传多样性分析[J].中国科学:生命科学,2001,31(1):13-21.
[27]刘晨晨,栾明宝,陈建华,等.苎麻Genomic-SSR与EST-SSR分子标记遗传差异性分析[J].中国麻业科学,2015(2):57-63.
[28] Ayres N M, Mcclung A M, Larkin P D, et al. Microsatellites and a single-nucleotide polymorphism differentiate apparent amylose classes in an extended pedigree of US rice germ plasm[J].Theoretical and Applied Genetics,1997,94(6):773-781.
[29]韩宏伟.中国刺槐遗传多样性和抗寒性地理变异规律的研究[D].保定:河北农业大学,2007.
[30]袁存权.刺槐有性生殖过程及交配系统研究[D].北京:北京林业大学,2013.
[2] Bongarten B C, Huber D A, Apsley D K. Environmental and genetic influences on short-rotation biomass production of black locust(Robinia pseudoacacia L.)in the Georgia Piedmont[J]. Forest Ecology&Management, 1992, 55(1-4):315-331.
[3]王世绩,张敦伦.刺槐[M].北京:中国科学技术出版社,1993.
[4]茹桃勤,李吉跃,张克勇,等.国外刺槐(Robinia pseudoacacia)研究[J].西北林学院学报,2005,20(3):102-108.
[5]宋跃朋,江锡兵,张曼,等.杨树Genomic-SSR与EST-SSR分子标记遗传差异性分析[J].北京林业大学学报,2010,32(5):1-7.
[6] Tuskan G A, Gunter L E, Yang Z K, et al. Characterization of microsatellites revealed by genomic sequencing of Populus trichocarpa[J]. Canadian Journal of Forest Research, 2004, 34:85-93.
[7] Varshney R K, Graner A, Sorrells M E, et al. Genomicmicrosatellite markers in plants:Features and applications[J].Trends in Biotechnology, 2005, 23(1):48-55.
[8] Yin T M, Tuskan G A, Gunter L E, et al. Genome structure and emerging evidence of an incipient sex chromosome in Populus[J].Genome Research, 2008, 18:422-430.
[9] R?der M S, Korzun V, Wendehake K, et al. A microsatellite map of wheat[J]. Genetics,1998,149(4):2007-2023.
[10]周延清.DNA分子标记技术在植物研究中的应用[M].北京:化学工业出版社,2005.
[11] Mutegi S M, Muchugi A, Carsan S, et al. Genetic diversity of the African poplar(Populus ilicifolia)populations in Kenya[J]. Tree Genetics&Genomes,2016,12(4):1-7.
[12] Sun R, Lin F, Huang P, et al. Moderate Genetic Diversity and Genetic Differentiation in the Relict Tree Liquidambar formosana Hance Revealed by Genic Simple Sequence Repeat Markers[J].Frontiers in Plant Science,2016,7.
[13]胡晓辉,毛瑞喜,苗华荣,等.山东省46个花生品种SSR指纹图谱构建与遗传多样性分析[J].核农学报,2016,30(10):1925-1933.
[14]艾畅,徐立安,赖焕林,等.马尾松种子园的遗传多样性与父本分析[J].林业科学,2006,42(11):146-150.
[15]车卓,常宏,石菁,等.利用SSR分子标记鉴定玉米杂交种吉祥1号纯度[J].中国农学通报,2015,32(18):28-32.
[16]齐凤慧,孙宏冉,詹亚光.EST-SSR标记在水曲柳雌雄鉴定中的应用[J].西北植物学报,2015,35(3):472-479.
[17]吴静,成仿云,庞利铮,等.紫斑牡丹表型性状与SSR分子标记的关联分析[J].北京林业大学学报,2016,38(8):80-87.
[18]杨新泉,刘鹏,韩宗福,等.普通小麦Genomic-SSR和EST-SSR分子标记遗传差异及其与系谱遗传距离的比较研究[J].遗传学报,2005,32(4):406-416.
[19]常玮,赵雪,李侠,等.大豆EST-SSR标记开发及与Genomic-SSR的比较研究[J].中国油料作物学报,2009,31(2):149-156.
[20] Wen M, Wang H, Xia Z, et al. Development of EST-SSR and genomic-SSR markers to assess genetic diversity in Jatropha curcas L.[J]. Bmc Research Notes,2010,3(1):1-8.
[21] Lian C, Hogetsu T. Development of microsatellite markers in black locust(Robinia pseudoacacia)using a dual-supression-PCR technique[J]. Molecular Ecology Notes,2002,2(3):211-213.
[22] Lian C, Oishi R, Miyashita N, et al. High somatic instability of a microsatellite locus in a clonal tree, Robinia pseudoacacia[J].Theoretical and Applied Genetics,2004,108(5):836-841.
[23] Mishima K, Hirao T, Urano S, et al. Isolation and characterization of microsatellite markers from Robinia pseudoacacia L.[J].Molecular Ecology Resources,2009,9(3):850-852.
[24] Smith J M, Haigh J. The hitch-hiking effect of a favourable gene[J].Genetical Research,1974,23(1):391-403.
[25] Van K, Hwang E Y, Kim M Y, et al. Discovery of single nucleotide polymorphisms in soybean using primers designed from ESTs[J].Euphytica,2004,139(2):147-157.
[26]贾继增,张正斌.小麦21条染色体RFLP作图位点遗传多样性分析[J].中国科学:生命科学,2001,31(1):13-21.
[27]刘晨晨,栾明宝,陈建华,等.苎麻Genomic-SSR与EST-SSR分子标记遗传差异性分析[J].中国麻业科学,2015(2):57-63.
[28] Ayres N M, Mcclung A M, Larkin P D, et al. Microsatellites and a single-nucleotide polymorphism differentiate apparent amylose classes in an extended pedigree of US rice germ plasm[J].Theoretical and Applied Genetics,1997,94(6):773-781.
[29]韩宏伟.中国刺槐遗传多样性和抗寒性地理变异规律的研究[D].保定:河北农业大学,2007.
[30]袁存权.刺槐有性生殖过程及交配系统研究[D].北京:北京林业大学,2013.
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