| 太湖新银鱼微卫星位点的分离与序列特征分析 |
| Isolation and Sequence Characterization of Microsatellite loci in Neosalanx taihuensis |
| 投稿时间：2014-06-16 修订日期：2014-07-14 |
| DOI：10.15928/j.1674-3075.2015.02.009 |
| 中文关键词:太湖新银鱼 微卫星 磁珠富集法 |
| 英文关键词:Neosalanx taihuensis microsatellite enrichment by magnetic beads |
| 基金项目:国家自然科学基金项目(No.51109091，51310105036)；湖北省教育厅项目(No.B2013149)；武汉市市属高校教研课题（2014007）。 |
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| 中文摘要: |
| 为了开发太湖新银鱼（Neosalanx taihuensis）的微卫星分子标记，采用生物素标记探针(AC)12、(AAC)10、(AAG)10和(GATA)8对其微卫星位点进行了筛选，并对其序列特征进行了分析。共获得490个微卫星序列，筛选的总效率为78.09%。筛选出725个微卫星位点，其中完美型592个，占81.66%；混合型82个，占11.31%；非完美型51个，占7.03%。探针(AC)12富集到的微卫星重复次数大多集中在14～26次，最高44次；探针(AAC)10和探针(AAG)10富集得到的微卫星重复次数主要集中在8～20次，最高42次；探针(GATA)8富集得到的微卫星重复次数一般在6～15次，最高32次。探针(AC)12、(AAC)10、(AAG)10和(GATA)8的杂交效率分别为85.19%、60.19%、5.16%和10.64%。筛选得到的725个微卫星位点中，二碱基重复位点390个（53.79%），三碱基重复位点284个（39.17%），四碱基重复位点47个（6.48%），五碱基重复位点1个（0.14%）和六碱基重复位点3个（0.42%）。根据二碱基重复核心序列进行引物设计，对抚仙湖24尾太湖新银鱼样本进行PCR扩增，电泳结果显示部分微卫星位点有较高的多态性，同时本研究获得的三碱基、四碱基、五碱基和六碱基重复位点也可为长重复单元微卫星引物的开发提供基础。 |
| 英文摘要: |
| Neosalanx taihuensis is naturally distributed in the middle and lower reaches of the Yangtze River and its connected lakes. It is one of the most important economic fishes in these waters and has been widely and successfully introduced to other fresh water lakes and reservoirs. Studies on the species mainly focused on stock enhancement, but there is a lack of research on its genetics which could be important for resource management. To better understand the genetic diversity and genetic structure of Neosalanx taihuensis, microsatellite DNA markers were developed for N. taihuensis using the magnetic bead technique. (AC)12, (AAC)10, (AAG)10 and (GATA)8 oligonucleotide probes were used to screen the microsatellite sequences in six N. taihuensis specimens collected from Fuxian Lake and Taihu Lake in March, 2012. A total of 490 microsatellite sequences were identified with a total screening efficiency of 78.09%. A total of 725 microsatellite loci were detected, with 592 perfect repeat motifs accounting for 81.66%, 82 compound repeat motifs accounting for 11.31%, and 51 imperfect repeat motifs accounting for 7.03%. Among them, there were 390 di-nucleotide repeat loci, 284 tri-nucleotide repeat loci, 47 tetra-nucleotide repeat loci, 1 penta-nucleotide repeat locus and 3 hexa-nucleotide repeat loci, contributing, respectively, 53.79%, 39.17%, 6.48%, 0.14% and 0.42% to the total microsatellite loci. The number of microsatellite repeats enriched by probe (AC) 12 ranged mostly from 14 to 26 but with the highest number of 44. The number of microsatellites repeats enriched by probes (AAC)10 or (AAG)10 ranged mostly from 8 to 20 with the highest number of 42. While the microsatellites repeats enriched by probes (GATA)8 ranged mostly from 6 to 15 with the highest number of 32. The hybridization efficiency of probes (AC)12, (AAC)10, (AAG)10 and (GATA)8 was 85.19%, 60.19%, 5.16% and 10.64%, respectively. PCR amplification was then conducted on 24 samples of N. taihuensis collected from Fuxian Lake using the primer design obtained from the di-nucleotide repeat loci. The electrophoretic pattern of PCR products showed high polymorphism. The tri-nucleotide repeat loci and tetra-nucleotide repeat loci obtained will be useful for developing long repeat microsatellite loci for N. taihuensis.
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| 李存耀,刘红艳,熊飞.2015.太湖新银鱼微卫星位点的分离与序列特征分析[J].水生态学杂志,36(2):55-60. |
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