卡氏肺孢子虫MSG抗原基因片段的克隆与分析

目的建立卡氏肺孢子虫动物模型,体外扩增卡氏肺孢子虫MSG基因部分片段,方法取肺孢子虫肺炎大鼠肺组织制备卡氏肺孢子虫(Pc)DNA,并以此模板扩增MSG基因片段,将PCR产物与pGEM-T-easy载体连接,转化E.coli DH5a,在含氨苄青霉素(Amp)的LB培养基上筛选出重组子,提取重组质粒,并通过PCR、酶切及测序验证,并进行序列比较分析;结果PCR扩增获得长度为554bp的序列,测序结果与报道的基因序列相比,碱基序列完全相同。结论本研究从卡氏肺孢子虫DNA中成功获得MSG基因,MSG基因的克隆,为进一步开展MSG蛋白致病机制、基因工程疫苗等研究奠定基础。

Molecular Cloning on the gene fragment of MSG Antigen from Pneumocystis carinii

Objective To establish animal model of Pneumocystis carinii and clone a gene fragment of MSG antigen from Pneumocystis carinii(Pc).Methods Pneumocystis carinii DNA was extracted from lung tissues of rats infected with P.carinii,and used it as template to amplify the MSG gene by PCR.Meanwhile,the PCR products were purified and inserted to pGEM-T Easy vector.E.coli DH5a were transformed in LB culture medium and the recombinant clones were selected.The positive clones of recombinants were identified by PCR and digested with enzymes.Then they were sequenced and analyzed.Results A gene fragment with 554 bp was obtained by PCR.The nucleotides sequence of the cloned gene was the same as that of the gene reported.Conclusion MSG gene fragment was successfully obtained from pneumocystis carinii DNA.Cloning on MSG gene would provide basis for the further studies on the pathologic mechanism,the gene engineering vaccine and so on.