摘要
目的将干血斑样本用于HIV-1基因序列测定和亚型分析。方法采集20例静脉吸毒HIV-1感染者的EDTA抗凝静脉全血2 ml,80μl制备干血斑样本。QIAamp Blood Mini试剂盒,10％Chelex 100树脂分别提取全血和干血斑中细胞DNA。HIV-1 env基因Gp41区2对外侧和内侧引物,按照巢式PCR方法,扩增Gp41区,PCR产物用内侧正反向引物测序。MEGA 3．0等软件完成序列比对及亚型分析。结果18对符合要求的干血斑和全血样本进入研究。16对经序列分析归为中国流行的BC重组亚型,样本间的基因平均离散率为5．79％,与中国流行的BC重组亚型的基因离散率为4．91％,与C亚型代表序列基因离散率为8．89％,与其他几个国际亚型代表株的基因离散率在15％～20％之间。另2对样本分别归属于泰国B’亚型和欧美B亚型。结论干血斑样本与全血样本HIV-1 DNA PCR产物亚型分析一致,氨基酸序列比对同义和非同义替换比例、位置和类型基本一致,其易采集,便于运输,保存条件和生物危险性低,费用低廉的优点适合作为实验条件较差的偏远地区HIV-1分子流行病学研究的一种样本采集手段。
Objective To use dried blood spot （D13S） in studying the sequence and subtype analysis of HIV-1 genome. Methods 2 ml whole blood containing EDTA anticoagulant from 20 HIV-1 infected patients were collected, then 80 μl blood was used to propare DBS. QIAamp Blood Mini kit and 10% Chelex 100 resin extracted DNA genome from whole blood and DBS as well as nested PCR amplified specifically HIV-1 Gp41 region from the two kinds of DNA extraction. Software MAGE 3.0 was used to study the sequence and subtype of the PCR products. Results Eligible 18 paired samples were analysed to show that 16 of them belonged to C54A, 97CNGX-7F, 98CN006 subtypes. The other two samples might belong to B. CN._. RL42 and 13. US. 90WEAU160. Conclusion Data showed that there were parallel results between the whole blood and DBS samples including subtype analysis, position of mutation and types of amino acid sequencing. Since DBS itself facilitated the collection, transportation and storage,it could be used as a measure to collect blood sample in resource limited area and to develop molecular epidemiologic research as well as early diagnosis on infant exposed to HIV.
出处
《中华流行病学杂志》
CAS
CSCD
北大核心
2006年第9期798-802,共5页
Chinese Journal of Epidemiology
基金
国家"十五"科技攻关计划课题(2004BA719A03)
关键词
艾滋病病毒
亚型分析
干血斑
Human immunodeficiency virus
Subtype analysis
Dried blood spot
