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梁素娴, 孙效文, 白俊杰, 高俊生. 微卫星标记对中国引进加州鲈养殖群体遗传多样性的分析[J]. 水生生物学报, 2008, 32(5): 694-700.
引用本文: 梁素娴, 孙效文, 白俊杰, 高俊生. 微卫星标记对中国引进加州鲈养殖群体遗传多样性的分析[J]. 水生生物学报, 2008, 32(5): 694-700.
LIANG Su-Xian, SUN Xiao-Wen, BAI Jun-Jie, GAO Jun-Sheng. GENETIC ANALYSIS FOR CULTURED LARGEMOUTH BASS (M ICRO PTERUS SALMO ID ES) IN CHINA WITHM ICROSATELLITES[J]. ACTA HYDROBIOLOGICA SINICA, 2008, 32(5): 694-700.
Citation: LIANG Su-Xian, SUN Xiao-Wen, BAI Jun-Jie, GAO Jun-Sheng. GENETIC ANALYSIS FOR CULTURED LARGEMOUTH BASS (M ICRO PTERUS SALMO ID ES) IN CHINA WITHM ICROSATELLITES[J]. ACTA HYDROBIOLOGICA SINICA, 2008, 32(5): 694-700.

微卫星标记对中国引进加州鲈养殖群体遗传多样性的分析

GENETIC ANALYSIS FOR CULTURED LARGEMOUTH BASS (M ICRO PTERUS SALMO ID ES) IN CHINA WITHM ICROSATELLITES

  • 摘要: 为了解我国引进加州鲈养殖群体的种群遗传结构和种质资源现状,本文发展了加州鲈的微卫星标记。用磁珠富集法获得加州鲈微卫星标记267个,设计并合成了85对微卫星引物对加州鲈的基因组进行遗传多样性分析,从中筛选出18对并从网上查询获得3对共21对引物对加州鲈的基因组DNA进行扩增。PCR扩增产物在8%的聚丙烯酰胺凝胶中分离,硝酸银染色,经紫外成像后对电泳条带进行统计分析。计算得到三个群体的平均等位基因数分别为2.3、2.3和2.1;多态信息含量(PIC)为0.0906-0.7480;平均杂合度观测值分别为0.384、0.396和0.356;杂合度期望值为0.375、0.403和0.368。统计结果初步表明3个不同群体平均杂合度处于中等偏低水平,遗传多样性不高。

     

    Abstract: The Largemouth bass (M icropterus salmoides) , native in the North America,was introduced into China in.980 sand is becoming an important freshwater cultured fish with more than.00, 000 tons of p roduction annually in China.Formore than 0 years of aquaculture, the germp lasm quality and genetic diversitymay have declined due to neglecting to inspect the genetic diversity in Largemouth bass and the lack of the longrun and effective administer. In order to aid in investigation of the population genetic structure and marker assisted breeding,microsatellite enrichment by magnetic beadswas used to isolating molecular genetic markers.The genomic DNA was cut by the enzyme of Sau3A I, and targeted segments were collected with the size of 400-900 base pairs by centrifugation of sucrose density gradient.Then the segmentswere purified in a low melting agarose gel and ligated with a short linkers(0bp) , from which, the“genetic PCR library”was created.These genomic DNA fragments were hybridized with a biotinlabeled SRS (simp le repeat sequence) p robe(CA).5.The hybrid mixture was incubated with magnetic beads coated with strep tavidin.Afterwashing to remove the nonSRS fragments, the eluted singlestranded DNA contained the selected microsatellite DNA.The selected DNAswere amp lified using p rimers designed comp lementary to the tinkers, cloned into the pMD.8T vector and transformed into competentE.coli DH5a.As a result, 67 microsatelliteswere obtained and 85 microsatellite p rimerswere designed by software Primer3.0, and.8 loci showed polymorphism. In addition, 3 pairs of microsatellite p rimers were obtained from nucleotide websites.The. polymorphic lociwere used to estimate genetic diversity and genetic differentiation in 3 culture populations ofLargemouth bass, three of which were collected from Guangzhou (G,n=30) ,Daliang (D,n=8) and Nanshui (N,n=30).Genome DNA was extracted from 88 samp les for the three culture populations.Each fish as extracted from blood andthe PCR was performed in a 5 L reaction and isolated by electrophoresis on 8% polyacrylamide gel and visualized by silver staining.The data were calculated and analyzed by statistic method.The results indicated that: the number of genotypes per locus ranged from to 3,with allele size ranging from.7 bpto 397 bp; the highest numbers of alleleswere from locus JZL3, JZL68, JZL83, JZL84, JZL85 and Lma. (3) followed bythe others were only acquired alleles.The mean effective number of three culture populations from Guangzhou, Daliangand Nanshui were..67,..75 and..67, respectively.Polymorphism information content value (PIC) wasmore than 0.5 in locus JZL3, JZL85 and Lma.; the PIC valuewasmore than 0.5 and less than 0.5 in locus JZL3, JZL36, JZL37, JZL43, JZL48, JZL53, JZL59, JZL60, JZL68, JZL7,JZL83, JZL84 and Lar7; the PIC value was less than 0.5 in locus JZL., JZL40, JZL67, JZL7. andMdo7.The observed heterozygosity (Ho) and the expected heterozygosity (He) were calculated based on frequencies of genotypes and alleles of each microsatellite locus, the average Ho value was the highest with Daliang (Ho=0.396,He= 0.403)and followed by Guangzhou (Ho=0.384, He=0.375) and Nanshui (Ho=0.356,He=0.368).The genetic departure index( d value) was ranged from -0.763 to 0.47 d value of JZL67 which in Daliang population was -0.763, obviously, on thelow side, JZL3. in Nanshui population was 0.47 on the high side, it showed the loci JZL67 deviated from the equilibrium inDaliang population for heterozyosis deficiency and JZL3. in Nanshui population for heterozyosis excess.All of these indices indicated that the cultured populations of Largemouth bass in Guangdong Province have relativelylow genetic diversity, and inbreeding has occurred.The reasons may result from inbreeding while artificial p ropagation, andneglecting the genetic diversities and population quantity while introduced into China.The methods for prevention of Largemouth bass germplasm degeneration are also suggested.

     

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