水貂PIT-1基因多态性及遗传多样性的研究
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摘要
水貂是小型珍贵毛皮兽,水貂皮、狐皮和波斯羔皮早已成为国际裘皮贸易的三大支柱产品。当前,水貂产业正面临着良好的发展机遇和前景,因此,提高水貂的生长性能,将给水貂养殖业带来更高的经济效益。
本研究选择PIT-1基因作为候选基因,以大连金州水貂养殖场的五个品种的水貂为试验对象,采用克隆测序结合PCR-SSCP的方法,检测了PIT-1基因部分片段的SNPs,并分析了突变位点导致的多态性与生长性状之间的相关性,本研究采集了金州黑水貂、短毛黑水貂、蓝宝石貂、咖啡色貂和红眼白水貂5个品种的174个样本,应用ISSR标记对这5个品种的遗传结构进行了分析。主要研究结果如下:
1.克隆了水貂PIT-1基因外显子1部分序列,片段大小为117bp,与马属的PIT-1基因外显子1同源性为90%,与犬科、绵羊、牛、人的PIT-1基因外显子同源性均高于88%,这段序列在哺乳动物中有较高的保守性,经SSCP分析没有发现多态位点。
2.克隆了水貂PIT-1基因外显子4部分序列,片段大小为150bp,已提交到Genebank,序列号(GU362205),经SSCP分析检测到一个突变位点:突变位点C43T导致的三种基因型对整个水貂群体体重、胸围、腹围有极显著影响(P<0.01),AA基因型个体体重、胸围、腹围最高,显著高于BB基因型个体(P<0.05),AA基因型为影响水貂体重、胸围、腹围的有利基因型。
3.克隆了水貂PIT-1基因外显子6部分序列,片段大小为164bp,已发布到Genebank,序列号(GU446673),经SSCP分析检测到一个突变位点:突变位点A143G导致的三种基因型对整个水貂群体体重、体长、体高、胸围、腹围有极显著的影响(P<0.01),DD基因型个体的体重、体长、胸围、腹围、屠体重显著高于CC基因型个体(P<0.05)。
4. PIT-1基因C43T突变位点和A143G突变位点产生的合并基因型对整个水貂群体的体重、胸围、腹围、屠体重有极显著的影响(P<0.01),对体长接近于极显著影响(P=0.015)。多重比较结果表明:AADD基因型个体的体重、胸围、屠体重显著高于BBCC基因型个体,BBDD基因型个体体长显著高于其他基因型个体(P<0.05)。
5. ISSR标记分析中,多态条带比率、Shannon多样性指数以及Nei基因多样性指数(h)均显示不同品种间具有较为丰富的遗传多样性。采用Nei遗传距离对五个居群进行分析,其中咖啡貂与蓝宝石水貂的遗传距离最近,相应的一致度最高;而蓝宝石水貂和金州黑水貂的遗传距离最远,遗传一致度最低;本文采用Gst基因分化系数方法对5个群体进行居群分析,结果表明水貂各个品种间遗传分化程度不高,遗传变异主要来自种群内。
Mink is small precious fur-bearing animal, and the furs of mink, fox, and persian lamb are already the three key products in the international fur market. Now the mink industry is facing good development opportunity and prosect, so improving the growth performance of mink will bring more economic benefits to the mink industry.
The minks bred in Dalian Jinzhou base were chosen as our research object, the SNPs in the coding regions of the PIT-1 were studied by cloning and sequencing combined with PCR-SSCP technique, finally, the correlation between polymorphism caused by growth traits was analyzed. This study made foundation of marker assisted selection using marker genes related growth performance, and provided way to breed mink, improve growth rate, fur quality of mink at molecular level, five species which were standard-pitchy mink, black mink, sapphire mink, brown mink, white mink were collected in this research, and applied ISSR marker to analyzing their genetic relationship. Main research results as follows:
1. The sequence (117bp) of the exon1 in PIT-1 gene, whose homology with Equus caballus PIT-1 gene exon1 was 90%,and the homology with Canis lupus, Ovis aries, Bos taurus ,Homo sapiens were achieve to 88% through Blast of NCBI, was cloned and submit to the genebank. The sequence was conserved in mammal animals. There no SNPs in this sequence by SSCP.
2. The sequence (150bp) of the exon4 in PIT-1 gene was cloned and the accession number was GU362205. A SNP in the exon4 was detected in the research by SSCP, the least square analysis showed that the genotypes of mutation had significant association with Body Weight (BW), Chest Circumference (CC), Abdomen Circumference (AC) (P<0.01), AA genotype mink had a significant higher Body Weight (BW), Chest Circumference (CC), Abdomen Circumference (AC) than BB genotype mink in the whole mink group. AA genotype were the beneficial genotype for the Body Weight (BW), Chest Circumference (CC), Abdomen Circumference (AC) in mink.
3. The sequence (164bp) of the exon6 in PIT-1 gene was cloned and the accession number was GU446673. A SNP in the exon 6 was detected in the research by SSCP, the least square analysis showed that the genotypes of mutation had significant association with Body Weight (BW), Body Longth (BL), Body Height (BH), Chest Circumference (CC), Abdomen Circumference (AC) and Carcass Weight (CW) in the whole mink group (P<0.01). DD genotype mink had a significant higher Body Weight (BW), Body Longth (BL), Chest Circumference (CC), Abdomen Circumference (AC) than CC genotype mink in the whole mink group.
4. The interaction of C43T mutation and A143G mutation in PIT-1 gene was discussed through combination genotype analysis, the least square analysis showed that the combined genotype had significant association with Body Weight (BW), Chest Circumference (CC), Abdomen Circumference (AC) and Carcass Weight (CW) in the whole mink group. Results of multiple comparison revealed that AADD genotype minks had a higher Body Weight (BW), Chest Circumference (CC) and Carcass Weight (CW) than AABB genotype minks; BBDD genotype minks had a higher Body Longth (BL) the other genotypes minks.
5. In ISSR molecular marker analysis , Polymorphic bands percent , Shannon's Information index, Nei's (1973) gene diversity showed high genetic diversity. Nei's genetic distance was used to analyze five populations, the results showed that the genetic distance between sapphire mink and brown mink was the nearest among the populations, and the genetic distance between standard-pitchy mink and sapphire mink was the longest. The coefficient of gene differentiation were used to analyze population genetic differentiation and the results showed that the genetic differentiation degree between populations was lower and the hereditary variation primarily came from within population.
本研究选择PIT-1基因作为候选基因,以大连金州水貂养殖场的五个品种的水貂为试验对象,采用克隆测序结合PCR-SSCP的方法,检测了PIT-1基因部分片段的SNPs,并分析了突变位点导致的多态性与生长性状之间的相关性,本研究采集了金州黑水貂、短毛黑水貂、蓝宝石貂、咖啡色貂和红眼白水貂5个品种的174个样本,应用ISSR标记对这5个品种的遗传结构进行了分析。主要研究结果如下:
1.克隆了水貂PIT-1基因外显子1部分序列,片段大小为117bp,与马属的PIT-1基因外显子1同源性为90%,与犬科、绵羊、牛、人的PIT-1基因外显子同源性均高于88%,这段序列在哺乳动物中有较高的保守性,经SSCP分析没有发现多态位点。
2.克隆了水貂PIT-1基因外显子4部分序列,片段大小为150bp,已提交到Genebank,序列号(GU362205),经SSCP分析检测到一个突变位点:突变位点C43T导致的三种基因型对整个水貂群体体重、胸围、腹围有极显著影响(P<0.01),AA基因型个体体重、胸围、腹围最高,显著高于BB基因型个体(P<0.05),AA基因型为影响水貂体重、胸围、腹围的有利基因型。
3.克隆了水貂PIT-1基因外显子6部分序列,片段大小为164bp,已发布到Genebank,序列号(GU446673),经SSCP分析检测到一个突变位点:突变位点A143G导致的三种基因型对整个水貂群体体重、体长、体高、胸围、腹围有极显著的影响(P<0.01),DD基因型个体的体重、体长、胸围、腹围、屠体重显著高于CC基因型个体(P<0.05)。
4. PIT-1基因C43T突变位点和A143G突变位点产生的合并基因型对整个水貂群体的体重、胸围、腹围、屠体重有极显著的影响(P<0.01),对体长接近于极显著影响(P=0.015)。多重比较结果表明:AADD基因型个体的体重、胸围、屠体重显著高于BBCC基因型个体,BBDD基因型个体体长显著高于其他基因型个体(P<0.05)。
5. ISSR标记分析中,多态条带比率、Shannon多样性指数以及Nei基因多样性指数(h)均显示不同品种间具有较为丰富的遗传多样性。采用Nei遗传距离对五个居群进行分析,其中咖啡貂与蓝宝石水貂的遗传距离最近,相应的一致度最高;而蓝宝石水貂和金州黑水貂的遗传距离最远,遗传一致度最低;本文采用Gst基因分化系数方法对5个群体进行居群分析,结果表明水貂各个品种间遗传分化程度不高,遗传变异主要来自种群内。
Mink is small precious fur-bearing animal, and the furs of mink, fox, and persian lamb are already the three key products in the international fur market. Now the mink industry is facing good development opportunity and prosect, so improving the growth performance of mink will bring more economic benefits to the mink industry.
The minks bred in Dalian Jinzhou base were chosen as our research object, the SNPs in the coding regions of the PIT-1 were studied by cloning and sequencing combined with PCR-SSCP technique, finally, the correlation between polymorphism caused by growth traits was analyzed. This study made foundation of marker assisted selection using marker genes related growth performance, and provided way to breed mink, improve growth rate, fur quality of mink at molecular level, five species which were standard-pitchy mink, black mink, sapphire mink, brown mink, white mink were collected in this research, and applied ISSR marker to analyzing their genetic relationship. Main research results as follows:
1. The sequence (117bp) of the exon1 in PIT-1 gene, whose homology with Equus caballus PIT-1 gene exon1 was 90%,and the homology with Canis lupus, Ovis aries, Bos taurus ,Homo sapiens were achieve to 88% through Blast of NCBI, was cloned and submit to the genebank. The sequence was conserved in mammal animals. There no SNPs in this sequence by SSCP.
2. The sequence (150bp) of the exon4 in PIT-1 gene was cloned and the accession number was GU362205. A SNP in the exon4 was detected in the research by SSCP, the least square analysis showed that the genotypes of mutation had significant association with Body Weight (BW), Chest Circumference (CC), Abdomen Circumference (AC) (P<0.01), AA genotype mink had a significant higher Body Weight (BW), Chest Circumference (CC), Abdomen Circumference (AC) than BB genotype mink in the whole mink group. AA genotype were the beneficial genotype for the Body Weight (BW), Chest Circumference (CC), Abdomen Circumference (AC) in mink.
3. The sequence (164bp) of the exon6 in PIT-1 gene was cloned and the accession number was GU446673. A SNP in the exon 6 was detected in the research by SSCP, the least square analysis showed that the genotypes of mutation had significant association with Body Weight (BW), Body Longth (BL), Body Height (BH), Chest Circumference (CC), Abdomen Circumference (AC) and Carcass Weight (CW) in the whole mink group (P<0.01). DD genotype mink had a significant higher Body Weight (BW), Body Longth (BL), Chest Circumference (CC), Abdomen Circumference (AC) than CC genotype mink in the whole mink group.
4. The interaction of C43T mutation and A143G mutation in PIT-1 gene was discussed through combination genotype analysis, the least square analysis showed that the combined genotype had significant association with Body Weight (BW), Chest Circumference (CC), Abdomen Circumference (AC) and Carcass Weight (CW) in the whole mink group. Results of multiple comparison revealed that AADD genotype minks had a higher Body Weight (BW), Chest Circumference (CC) and Carcass Weight (CW) than AABB genotype minks; BBDD genotype minks had a higher Body Longth (BL) the other genotypes minks.
5. In ISSR molecular marker analysis , Polymorphic bands percent , Shannon's Information index, Nei's (1973) gene diversity showed high genetic diversity. Nei's genetic distance was used to analyze five populations, the results showed that the genetic distance between sapphire mink and brown mink was the nearest among the populations, and the genetic distance between standard-pitchy mink and sapphire mink was the longest. The coefficient of gene differentiation were used to analyze population genetic differentiation and the results showed that the genetic differentiation degree between populations was lower and the hereditary variation primarily came from within population.
引文
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