重复序列DNA探针分析我国恶性疟原虫虫株

Differentiation of Plasmodium falciparum isolates from China with a repetitive DNA probe

从人工培养的恶性疟原虫安徽Fcc/AH株、云南思茅株和海南Fcc_1/HN株中,分别提取基因组DNA,去除RNA和降解的DNA,精确定量,平行梯度稀释点样(100ng～0.05 ng),以^(32)P标记的PF rep20为探针,斑点杂交的检测水平安徽Fcc/AH株为0.5 ng、云南思茅株2 ng、海南Fcc_1/HN株3 ng.进一步用HindⅢ限制性内切酶完全酶切3株P.f.DNA,Southern印迹分析显示,PF rep20识别的3株P.f.靶DNA片段的大小、数量及信号强度明显不同,提示我国P.f.安徽Fcc/AH株、云南思茅株和海南Fcc_1/HN株DNA序列存在着差异.

The genomic DNA was extracted and purified from cultured P.falciparum Anhui, Yunnan and Hainan isolates respectively removing RNA and degradated DNA, quantitated accurately and diluted gradiently (100 ng-0.05 ng), then discriminated by dot blot hybridization with 32P PF rep20. The levels of detection were Anhui Fcc/AH isolate 0.5 ng, Yunnan Simao isolate 2 ng and Hainan Fcc1 /HN isolate 3 ng. The genomic DNA of three P. falciparum isolates, which was digested with restriction enzyme Hind Ⅲ , was further analysed by Southern blot hybridization with 32P PF rep20. The results showed that there were obviously diversities in size, number and radiointensity of target DNA fragments of three isolates which were recognized with 32P PFrep20. It is suggested that there is differences in DNA repetitive sequence of P. falciparum Anhui Fcc/AH, Yunnan Simao and Hainan Foc1 /HN isolates from China.