中间球海胆与光棘球海胆的杂交及分子遗传学研究
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摘要
本文以中间球海胆(Strongylocentrotus intermedius)、光棘球海胆(S. nudus)及其杂交的F_1代为材料,通过分子标记、构建遗传连锁图谱及基因表达差异等分子遗传学技术对两种海胆杂交产生的包括杂交优势在内的遗传变异进行了相关的研究,为海胆杂交育种提供了遗传学依据。主要内容包括:
1杂交海胆的分子鉴定及生物学性状特征。
1.1从24对SSR引物中筛选出一对,能对父母本和杂交种的基因组DNA进行有效扩增,并且扩增条带清晰稳定。利用杂交海胆中同时拥有父母本的互补带型,对杂交海胆进行了分子鉴定。
1.2杂交海胆的外部形态特征介于双亲之间,正反交的F_1代壳色、棘色等形态特征接近,都有两种表型(一种壳为紫褐色,棘发白、尖部略紫;另一种壳和棘均为紫色),并且两种表型所占比例不同,壳径在1cm左右时正交F_1代(中间球海胆♀×光棘球海胆♂)中为74.0%和26.0%,反交F_1代(光棘球海胆♀×中间球海胆♂)中各占68.1%和31.9%;壳径3cm左右时正交F_1代为77.2%和22.8%,反交F_1代各占70.5%和29.5%。正交F_1代的生殖腺重、壳径、体重等平均值都比母本具有不同程度的优势,有的时间达到显著水平。有近10%的杂种生殖腺颜色不如母本鲜艳。
2杂交海胆(中间球海胆♀×光棘球海胆♂)群体多样性的AFLP分析及AFLP标记与数量性状的相关性分析。
2.1对光棘球海胆(NU)、中间球海胆(IN)及其杂交的F_1代(中间球海胆♀×光棘球海胆♂)(HYb,HYc两种表型)3个群体进行了AFLP分析,并根据遗传距离进行了个体聚类分析。结果表明:3个群体的多态位点比例分别是:81.99%,80.51%和95.95%。香农多样性指数分别为:0.2331±0.1273, 0.2005±0.1385和0.2625±0.1067。遗传相似度分别为: 0.6876±0.0523, 0.6501±0.0548和0.6552±0.0553。分子方差分析(AMOVA)分析结果表明,变异来源有25.39%来自群体间,有74.61%来自群体内。尽管杂交海胆在表型上可以明显分成两种类型,但是通过AFLP统计的遗传距离进行的个体聚类却聚在一起,不能分成两个群体。
2.2对杂交海胆的生殖腺、壳径、体重、棘长等数量性状与AFLP标记进行
Based on the materials of sea urchin Strongylocentrotus intermedius、S. nudus and their hybrids, the variations including heterosis were studied by the technologies of molecular genetics.The major results and conclusions are as follows:
1 The hybrids were identified by molecular marker technology. The biological characters of hybrids and parents were observed and measured.
1.1 One primer pair of SSR was filtered from 24 primer pairs, which can produced steady and clear amplification bands. The bands of hybrids were complementary with that of parents. This marker was used in the idenfication of hybrids and the parents.
1.2 Morphological characters are in between the parents. Hybrids by S.intermedius♀×S.nudus♂(IN♀×NU♂) and by S. intermedius♀×S. nudus♂(NU♀×IN♂) are similar in morphology. They all have two phenotypes, one is purple-brown test (shell) and white spine with purple tip; another is purple test and spine. The two phenotypes of hybrids have different ratios. The ratios of two phenotypes are 74.0% and 26.0% in IN♀×NU♂, 68.1% and 31.9% in NU♀×IN♂when the test diameter is about 1 centimeter; while 77.2% and 22.8% in IN♀×NU♂, 70.5% and 29.5% in NU♀×IN♂when the test diameter is about 3 centimeter.The hybrids (IN♀×NU♂) show heterosis in gonad weight, test diameter and body weight in average. Sometimes the difference between hybrids and S.intermedius shows remarkable. About ten percent of the hybrids (IN♀×NU♂) show inferior gonad color than female parents.
2. The genetic diversity of three populations (hybrids and their parents) was analysed by AFLP technology.
2.1 AFLP analyses of genetic diversity in the three populations of Strongylocentrotus nudus (NU), S. intermedius (IN) and F_1 progeny (IN♀×NU♂) (HYb and HYc represent two different phynotypes of F_1) indicate that the percent of polymorphic loci in three populations is 81.99%,80.51% and 95.95% respectively. Shannon diversity index is 0.2331±0.1273, 0.2005±0.1385 and 0.2625±0.1067 respectively. Genetic similarity is 0.6876±0.0523, 0.6501±0.0548 and 0.6552±0.0553 respectively. AMOVA analysis indicates that , 25.39% of variance is among populations and 74.61% of variance is within populations. Though hybrids can be classified two types by apparent characters, they cluster each other by UPGMA method according to their genetic distances.
2.2 Correlation analyses between AFLP markers and quantitative traits of gonad weight, test diameter, body weight and spine length indicate that 18 AFLP markers are remarkably correlated with gonad weight, 3 markers are strongly correlated and the biggest contribution ratio is 17.16%. Twenty-one AFLP markers are remarkably
1杂交海胆的分子鉴定及生物学性状特征。
1.1从24对SSR引物中筛选出一对,能对父母本和杂交种的基因组DNA进行有效扩增,并且扩增条带清晰稳定。利用杂交海胆中同时拥有父母本的互补带型,对杂交海胆进行了分子鉴定。
1.2杂交海胆的外部形态特征介于双亲之间,正反交的F_1代壳色、棘色等形态特征接近,都有两种表型(一种壳为紫褐色,棘发白、尖部略紫;另一种壳和棘均为紫色),并且两种表型所占比例不同,壳径在1cm左右时正交F_1代(中间球海胆♀×光棘球海胆♂)中为74.0%和26.0%,反交F_1代(光棘球海胆♀×中间球海胆♂)中各占68.1%和31.9%;壳径3cm左右时正交F_1代为77.2%和22.8%,反交F_1代各占70.5%和29.5%。正交F_1代的生殖腺重、壳径、体重等平均值都比母本具有不同程度的优势,有的时间达到显著水平。有近10%的杂种生殖腺颜色不如母本鲜艳。
2杂交海胆(中间球海胆♀×光棘球海胆♂)群体多样性的AFLP分析及AFLP标记与数量性状的相关性分析。
2.1对光棘球海胆(NU)、中间球海胆(IN)及其杂交的F_1代(中间球海胆♀×光棘球海胆♂)(HYb,HYc两种表型)3个群体进行了AFLP分析,并根据遗传距离进行了个体聚类分析。结果表明:3个群体的多态位点比例分别是:81.99%,80.51%和95.95%。香农多样性指数分别为:0.2331±0.1273, 0.2005±0.1385和0.2625±0.1067。遗传相似度分别为: 0.6876±0.0523, 0.6501±0.0548和0.6552±0.0553。分子方差分析(AMOVA)分析结果表明,变异来源有25.39%来自群体间,有74.61%来自群体内。尽管杂交海胆在表型上可以明显分成两种类型,但是通过AFLP统计的遗传距离进行的个体聚类却聚在一起,不能分成两个群体。
2.2对杂交海胆的生殖腺、壳径、体重、棘长等数量性状与AFLP标记进行
Based on the materials of sea urchin Strongylocentrotus intermedius、S. nudus and their hybrids, the variations including heterosis were studied by the technologies of molecular genetics.The major results and conclusions are as follows:
1 The hybrids were identified by molecular marker technology. The biological characters of hybrids and parents were observed and measured.
1.1 One primer pair of SSR was filtered from 24 primer pairs, which can produced steady and clear amplification bands. The bands of hybrids were complementary with that of parents. This marker was used in the idenfication of hybrids and the parents.
1.2 Morphological characters are in between the parents. Hybrids by S.intermedius♀×S.nudus♂(IN♀×NU♂) and by S. intermedius♀×S. nudus♂(NU♀×IN♂) are similar in morphology. They all have two phenotypes, one is purple-brown test (shell) and white spine with purple tip; another is purple test and spine. The two phenotypes of hybrids have different ratios. The ratios of two phenotypes are 74.0% and 26.0% in IN♀×NU♂, 68.1% and 31.9% in NU♀×IN♂when the test diameter is about 1 centimeter; while 77.2% and 22.8% in IN♀×NU♂, 70.5% and 29.5% in NU♀×IN♂when the test diameter is about 3 centimeter.The hybrids (IN♀×NU♂) show heterosis in gonad weight, test diameter and body weight in average. Sometimes the difference between hybrids and S.intermedius shows remarkable. About ten percent of the hybrids (IN♀×NU♂) show inferior gonad color than female parents.
2. The genetic diversity of three populations (hybrids and their parents) was analysed by AFLP technology.
2.1 AFLP analyses of genetic diversity in the three populations of Strongylocentrotus nudus (NU), S. intermedius (IN) and F_1 progeny (IN♀×NU♂) (HYb and HYc represent two different phynotypes of F_1) indicate that the percent of polymorphic loci in three populations is 81.99%,80.51% and 95.95% respectively. Shannon diversity index is 0.2331±0.1273, 0.2005±0.1385 and 0.2625±0.1067 respectively. Genetic similarity is 0.6876±0.0523, 0.6501±0.0548 and 0.6552±0.0553 respectively. AMOVA analysis indicates that , 25.39% of variance is among populations and 74.61% of variance is within populations. Though hybrids can be classified two types by apparent characters, they cluster each other by UPGMA method according to their genetic distances.
2.2 Correlation analyses between AFLP markers and quantitative traits of gonad weight, test diameter, body weight and spine length indicate that 18 AFLP markers are remarkably correlated with gonad weight, 3 markers are strongly correlated and the biggest contribution ratio is 17.16%. Twenty-one AFLP markers are remarkably
引文
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