安氏隐孢子虫ITS-1序列的PCR扩增、克隆及分析

PCR amplification,cloning and sequencing of ITS-1 rDNA from cryptosporidium andersoni

通过对国内三株安氏隐孢子虫(Cryptosporidium andersoni)即GD株、HN株和AH株的rDNA的内转录间隔区Ⅰ(ITS_1)序列进行PCR扩增、克隆、测序和序列分析,旨在确定ITS_1是否可作为C.andersoni分子分类的遗传标记。结果表明:GD株、HN株和AH株的ITS_1序列基本一致,仅AH株有三个碱基的差异;但与GenBank注册的C.muris和C.parvum存在种间差异,而且差异显著。说明ITS_1可作为C.andersoni种的遗传标记,从而为隐孢子虫属的种间鉴定以及进一步的分子流行病学调查和分子诊断学研究奠定了基础。

The aims of this study were to assess whether the first internal transcribed spacer （ITS-1） of ribosomal DNA （rDNA） could be used as genetic markers of Cryptosporidium andersoni from different geographic origins in China, and to establish the foundation for further studies on molecular epidemiology and diagnostics of cryptesporidiesis. The ITS-1 rDNA from GD, HN and AH strains of C. andersoni were amplified by PCR, cloned and sequenced. The result showed that the ITS-1 sequences of GD, HN,AH strains were basically identical,which were unequivocally different with the sequences of the C. muris and C. parvum registered in the GenBank; but the ITS-1 sequence of the AH strain differed at three bases compared with that of the other 2 strains.The results indicate that the ITS-1 sequences provide useful genetic markers for the identification and differentiation of C. andersoni species, and also lay down the foundation for further study on molecular epidemiology and diagnostics of cryptesporidiesis.