长牡蛎17个fosmid-SSR标记的开发与分析
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大连海洋大学生命科学与技术学院,中国科学院海洋研究所,中国科学院海洋研究所,大连海洋大学生命科学与技术学院,中国科学院海洋研究所

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国家“九七三”重点基础研究发展计划(2010CB126402);国家自然科学基金重点项目(40730845);国家公益性行业(农业)科研专项(3-53)


Development and analysis of 17 SSR from Crassostrea gigas fosmid database
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Dalian Ocean University,Institute of Oceanology,Chinese Academy of Sciences,TsingTao,Institute of Oceanology,Chinese Academy of Sciences,TsingTao,Dalian Ocean University,Institute of Oceanology,Chinese Academy of Sciences,TsingTao

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    摘要:

    研究所用序列由长牡蛎fosmid文库末端测序获得。首先用TRF程序扫描序列获得一批重复单元为2碱基的候选SSR位点,然后在部分SSR序列两侧的保守区设计50对引物,对取自山东青岛的一个长牡蛎野生群体进行基因型分析。结果显示,共有17个SSR位点显示多态性,等位基因数(Na)平均为4,有效等位基因数(Ne)平均为2.82,平均杂合度观测值(Ho)和期望值(He)分别为0.395 9 和0.628 8。其中,11个位点的多态信息含量(PIC)值均大于0.5,共有49个等位基因,适合对长牡蛎群体遗传结构的分析;6个位点 0.25<PIC<0.5,为中度多态位点。χ2检验估计Hardy-Weinberg平衡,经Sequential Bonferroni校正后,除DEC_1以外,其他位点仍偏离平衡。将SSR附近10 000 bp内基因组预测的编码蛋白通过与NCBI的nr库blastp分析,共有13条微卫星序列获得连锁的相关基因功能注释。

    Abstract:

    Pacific oyster(Crassostrea gigas)is a widespread,delicious and nutritious species and is of high economic value.The codominant molecular microsatellite marker(also called simple sequence repeat,SSR)complies with Mendelian inheritance principles and is a powerful tool for analyzing population genetic structure,breeding and constructing genetic map because of its highly polymorphic and accurate genotyping.Sequences used in the study were obtained from end sequencing of fosmid library of the C.gigas.TRF software was used to exploit SSR loci with dinucleotide repeat,and 50 pairs of primers were acquired based on conserved SSR flanking sequence.We did preliminary screening in eight individuals and 17 of the above 50 primers were proved polymorphic.These primers were then screened and verified in a wild population in Qingdao,Shandong Province.Of these loci,17 showed polymorphism.The average number of allele(Na),number of effective alleles(Ne),observed heterozygosity(Ho),and expected heterozygosity(He)was 4,2.82,0.395 9,and 0.628 8,respectively.Among these loci,polymorphism information content(PIC)of 11 loci was more than 0.5,and 49 alleles in total were suitable for C.gigas population genetic structure analysis.The other 6 loci was moderately polymorphic and 0.25χ2 test)and Sequential Bonferroni calibration,we found that all loci except DEC_1 deviated equilibrium,suggesting that the growth and frequencies of genotypes of C.gigas changed.Blastp alignment analysis of 10 000 bp flanking sequences of SSR loci in nr database of NCBI showed that 13 linked gene sequences were development or metabolism related and can be used as molecular markers.The present results indicated that the development of SSR molecular marker based on the fosmid library can effectively remedy the predicament of lacking SSR markers of Pacific oyster to provide basis to protect and utilize the genetic diversity of this species.

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李慧娟,亓海刚,李莉,闫喜武,张国范.长牡蛎17个fosmid-SSR标记的开发与分析[J].水产学报,2011,35(10):1463~1468

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  • 收稿日期:2011-02-12
  • 最后修改日期:2011-05-06
  • 录用日期:2011-05-21
  • 在线发布日期: 2011-10-17
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