中华鳖微卫星座位的分离与鉴定

Isolation and characterization of microsatellite DNA Markers from softshell turtle Trionyx sinensis

将生物素杂交法与放射性同位素杂交法相结合，从中华鳖Trionyx sinensis基因组中分离出含有GA／CT和CA／GT重复类型的微卫星分子标记。共获得931个阳性克隆，从中选取286个进行测序，得到273个（95．45％）含有微卫星的序列，其中70．34％的重复数在10以上，55．51％为完美型。除探针中使用的CA和GA重复单元外，还观察到GC、TGTGT的重复序列。设计获得242对微卫星引物，合成40对，并对中华鳖群体遗传背景进行分析，结果表明：8对引物表现为单态，6对引物扩增带不是目的带，4对引物无扩增条带，其余22对引物扩增出多态性条带；平均等位基因数为3．48个，观测杂合度为0．1667～0．9620，期望杂合度为0．1528～0．7222，多态信息含量为0．1411—0．6800，平均多态信息含量为0．4382，大部分标记适用于中华鳖群体的遗传研究。

A library rich in CA/GT and GA/CT microsatellites was constructed from softshell turtle Trionyx sinensis. Nine hundred and thirty one positive clones were obtained through twice screen. From these clones, 272 microsatellite sequences （about 95.45% ） were isolated, in which 70. 34% microsatellites repeated more than 10 times and 55.51% microsatellites were perfect repeats. Primer pairs were designed in the flanking regions of 40 microsatelites and subjected to PCR amplification. In 3 populations of 18 individuals, 4 pairs failed to amplification, 6 ones did not amplify target bands, 8 ones were monomorphic and the rest were polymorphic with an average of 3.48 alleles per locus. Heterozygosities ranged between 0. 1528 and 0. 7222, homozygosties varied from 0. 1667 to 0. 9620, and PIC changed from 0. 1411 to 0. 6800 with an average of 0. 4382. The results indicated that most of the microsatellites isolated were qualified to be applied to the population genetic studies of the turtle.