鹿结核病野毒株与卡介苗差异基因基因文库的构建

Construction of subtracted gene library for deer tuberculosis wild strain and BCG

应用四碱基限制性内切酶对鹿结核病流行株和卡介苗基因组分别酶切,以鹿结核流行株酶切产物为testerDNA,卡介苗酶切产物为driver DNA,testerDNA接头连接后与driver DNA进行抑制性消减杂交。将获得的消减PCR产物与pMD-18连接,JM109感受态细胞,进行蓝白斑筛选。结果表明,RsaI酶切产生的酶切产物在0.1～2.0 kb之间,将消减PCR产物克隆后,挑取208个转化子,构建了鹿结核病流行株与卡介苗的差异基因文库。结果表明,应用抑制性消减杂交技术,构建鹿结核病流行株与卡介苗基因组差减文库,可为鹿结核病自然感染和卡介苗免疫的鉴别诊断及鹿结核病的综合防治奠定基础。

By suppression subtractive hybridization, the construction of subtracted gene library for deer tuberculosis wild strain and BCG , for differential diagnosising of the deer tuberculosis nature infection and BCG immunization, and laying the foundation of comprehensive prevention and control of deer tuberculosis. Application of four base pairs of restriction endonucleases enzyme on deer tuberculosis wild strains and BCG genome , the enzyme products of deer tuberculosis wild strains as tester DNA , the enzyme products of BCG as driver DNA, After tester DNA joint with connection , suppression subtractive hybridization with driver DNA. Subtractive PCR products were inserted into pMD18-T vector and be transformed into the JM109, screened of blue and white clones of the transformants. Rsal enzyme products in 0.1-2.0 kb, after clone the subtractive PCR products, out of 208 transformants, construction of subtracted gene library for deer tuberculosis wild strain and BCG. By suppression subtractive hybridization, the successful construction of subtracted gene library for deer tuberculosis wild strain and BCG. By suppression subtractive hybridization, the construction of subtracted gene library for deer tuberculosis wild strain and BCG, for differential diagnosising of the deer tuberculosis nature infection and BCG immunization, and laying the foundation of comprehensive prevention and control of deer tuberculosis.