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杜氏利什曼原虫动基体DNA种林特异片段的克隆 被引量:2

CLONING OF THE SEQUENCES OF KINETOPLAST DNA SPECIFIC TO LEISHMANIA DONOVAN I SPECIES AND STRAINS
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摘要 作者在质粒pUC18中克隆了限制性内切酶AIuI消化的Leishmania donovani四川人分离株kDNA片段,筛选后获得能区别杜氏利什曼原虫山丘疫区分离株和平原疫区分离株的的克隆pLK1-10和对杜氏利什曼原虫四川人分离株特异的克隆pLK1-14,对杜氏利什曼原虫种特异的克隆PLK1-1、pLK1-2等。这些克隆将是鉴定杜氏利什曼原虫,区别鉴定山丘及平原疫区黑热病病原体较好的探针。 To identify the heterogeneous DNA sequences in the kinetoplast DNA (kDNA) mini-circles, we digested the kDNA from various species and isolates of Leishmania into fragments with several restriction endonucleases and those fragments were southern hybridized with whole kDNA. By this test, the AluI fragments were shown to possess the species-and strain-specific sequences. We inserted these kDNA (from L. d. Sichuan human isolate) fragments into Smal site of plasmid pUC 18. The blunt ligation reaction was carried out with T4 DNA ligase supplemented by T4 RNA ligase. Tens of recombinants were obtained and at least 16 recombinants were shown to have the sequences of kDNA by colony hybridization with whole kDNA. The inserted kDNA sequences can be cut off from vector by Bam HI and EcoR I digestion. The recombinant DNA had no homolgous sequences with human genomic DNA, Leptospiral DNA, Romanomermis DNA and L. donovani genomic DNA. The clone pLKl-14, which is the smallest one among all the clones obtained, onlyhybridized to the L.d. Sichuan human isolate from which it was originated. When pLKl-14was digested by BamHI and EcoRI, a 180bp fragment comprising about 20 bp of pUC 18sequence, could be produced which corresponded to the 120±30 bp fragment of kDNA digested by Alul and was shown to be isolate specific. The clone of pLKl-10 hybridized to L. donovani isolates from hill and desert foci, might be used as a specific probe in the distinction of L.d. isolates from hill foci and plain foci. The clones pLKl-1, pLKl-2, and pLKl-15 were present in all isolates of visceral Leishmania but not in L. major and lizard Leishmania tested. These sequences might be used as specific probes in the diagnosis of visceral leishmaniasis and might be useful in epidemiological studies for identification of vectors and reservoirs.
出处 《中国寄生虫学与寄生虫病杂志》 CAS CSCD 北大核心 1991年第4期269-273,T014,共6页 Chinese Journal of Parasitology and Parasitic Diseases
基金 国家教委博士点基金
关键词 利什曼原虫 动基体DNA 克隆 Leishmania donovani, kinetoblast DNA, clone.
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  • 1罗萍,地方病通报,1989年,4卷,61页
  • 2吕洪刚,华西医科大学学报,1988年,19卷,217页
  • 3吕洪刚,华西医科大学学报,1988年,19卷,222页

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