恶性疟原虫基因文库构建及特异DNA探针的应用研究

将恶性疟原虫Ｆｃｃ－１／ＨＮ株基因组ＤＮＡ用ＨｉｎｄⅢＤＮＡ用ＨｉｎｄⅢ消化与ｐＢＲ３２２重组后导入大肠杆菌ＨＢ１０１，转化效率为１．５×１０＾６转化子／μｇ ＤＮＡ，建成基因库。用Ｐ．ｆ基因组ＤＮＡ原位杂交，证明了该库的有效性，又筛出特异克隆。克隆之一ｐＢＦ２ ＤＮＡ用＾３２Ｐ和光敏生物素标记作探针，可分别从间日疟原虫，恶疟原虫，伯氏疟原虫，食蟹猴疟原虫和人白细胞ＤＮＡ样本中特异地检出Ｐ，ｆ

Construction of Plasmodium Genomic Library and Applications of the Specific DNA Probe

DNA of Fcc1/HN gene was digested with Hind and recombined with pBR322 and then transferred into E.coli HB 101 for the construction of genomic library. The transfer rate was shown to be 1.5 106 clones per g DNA.This library was provedto be effective when P.f genomic DNA was used for hybridization with the specific clone screened. One of the clone pBF2 DNA was labelled with32P and lightsensitivebiotin and used as probe, which could specifically distinguishthe P.f DNA form DNA of human leukocyte and some other kindsof plasmodium DNA, such as BXP.v, P.b and P.c in which the probewas shown to have high specificity for P.f. The sensitivity ofthis probe in the detection of P.f genomic DNA was 5 g and aparasitemia as 0.001% . The results of the detection with theprobe labelled by the two methods was in agreement with the results with microscope,demonstrating a very promisingDapplicability of the probe.