间日疟原虫特异性DNA片段的克隆和鉴定

CLONING AND IDENTIFICATION OF THE DNA FRAGMENTS SPECIFIC FOR PLASMODIUM VIVAX

本文从间日疟患者血液中分离出间日疟原虫,将其基因组 DNA 片段克隆到载体 pUC12质粒中,构建了间日疟原虫 DNA 文库。利用质粒的遗传标志初步筛选重组克隆;用^(32)P 标记的间日疟原虫DNA,人白细胞 DNA 和恶性疟原虫 DNA 为探针分别作菌落杂交,差异筛选出4株间日疟原虫特异的重组克隆,命名为 pVA1—4。然后酶切鉴定其插入片段,并用 Southern blot 分析,表明重组质粒 pVAl 含有间日疟原虫特异性 DNA 片段。

The erythrocytes infected with P.vivax were preliminary separared and purified from the pa-tient's blood by Percoll gradient centrifugation.For constructing a P.vivax DNA library,the genomic DNAfragments of P.vivax partially digested with Sau3A were ligated into the BamHI site of plasmid pUC12 beforetransformation of E.coli JM103 host.Transformed clones were chosen by plasmia hereditary characters anddifferentially screened by the colony hybridization,using P-labelled nick translated human DNA,P.vivaxDNA or P.falciparum DNA,respectively.Southern blot analysis showed that the insert of recombinant plas-mid pVA1 can hybridize to P.vivax DNA,but not to human DNA and P.falciparum DNA.