用PCR-SSP方法检测中国恒河猴Mamu-DRB*W101和Mamu-DRB*W201基因
Detection of Mamu-DRB*W101 and Mamu-DRB*W201 in Chinese Rhesus Monkeys by PCR-SSP
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摘要: 利用序列特异引物聚合酶链反应（polymerase chain reactionsequence-specific primers，PCR-SSP）方法扩增中国恒河猴的主要组织相容性复合体II类基因Mamu-DRB*W101、- DRB*W201，初步了解中国恒河猴中Mamu-DRB*W101、- DRB*W201基因的阳性率。采集中国恒河猴静脉血，用DNA提取试剂盒提取全血DNA，分别用Mamu-DRB*W101、- DRB*W201特异引物PCR扩增Mamu-DRB*W101、- DRB*W201基因的第二外显子区域，并对扩增出的阳性条带进行测序，与已知序列对比验证序列是否正确。共检测了来自136只中国恒河猴的样本，PCR检测出Mamu-DRB*W101阳性个体10只，Mamu-DRB*W201阳性个体也是10只，其阳性个体所占比率均为7.35%。测序结果表明，PCR扩增产物的核苷酸序列与基因库中的序列完全一致。本研究表明中国恒河猴中存在Mamu-DRB*W101、- DRB*W201基因阳性个体，为中国恒河猴在AIDS研究中的应用及进一步分析中国恒河猴MHC II类基因提供了基础。Abstract: The aim of this study was to obtain preliminary data about the frequency of 2 MHC class II genes, Mamu-DRB*W101 and Mamu-DRB*W201 among Chinese rhesus macaques. Blood was obtained, and DNA was extracted using QIAamp DNA Blood Mini Kit according to the manufacturer’s instruction. PCR was performed using sequence specific primers to amplify the 2nd exon of Mamu-DRB*W101 and Mamu-DRB*W201. The nucleotide sequences of the amplicons were confirmed by sequencing. In the 136 samples tested in this study, 10 samples were found positive by PCR for Mamu-DRB*W101 and 10 samples positive for Mamu-DRB*W201. The frequency of positive individuals among the Chinese macaques tested was 7.35% for both Mamu-DRB*W101 and Mamu-DRB*W201. The nucleotide sequences of the amplicons were identical to the published sequences. This study demonstrated the existence of both Mamu-DRB*W101 and Mamu-DRB*W201 among Chinese rhesus macaques and provided the basis for large scale typing of these two alleles and studies using Chinese rhesus macaques in SAIDS.
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