鸡毒霉形体HS株pMGA多基因族的研究  

Studies on the pMGA Multigene Family of Mycoplasma gallisepticum HS Strain

作　　者：翁长江[1] 毕丁仁[1] 沈青春[1] 李自力[1] 石德时[1] 许青荣[1] 刘梅[1] 程峰[1] 

机构地区：[1]华中农业大学畜牧兽医学院,湖北武汉430070

出　　处：《中国兽医学报》2003年第2期149-152,共4页Chinese Journal of Veterinary Science

基　　金：湖北省自然科学基金资助项目 (99J0 10 4)

摘　　要：鸡毒霉形体 (Mycoplasma gallisepticum ,GM)基因组中存在着编码细胞表面血凝粘附素蛋白 (protein of My-coplasma gallisepticum adhesin,p MGA)相关基因组的多基因族。为筛选出含有完整的 p MGA基因片段 ,并估算出鸡毒霉形体 HS株基因组中 p MGA多基因族的基因数 ,本试验以 p UC1 8为载体 ,用 Eco R 限制性内切酶构建了鸡毒霉形体 HS株的基因组文库。根据 p MGA基因引导序列的保守区和 p MGA基因间隔区的序列 ,人工合成了 2个寡核苷酸探针 (探针 、探针 ) ,经标记后 ,双筛选所建的基因文库 ,从 6 0 0个转化子中共得到的 38个含有 p MGA基因的阳性克隆子 ,选其中 1个 7.5 kb的片段 (17号阳性克隆子 ,W17) ,用 4种限制性内切酶 (Eco R ,Hind ,Pst ,Bgl )酶切消化 ,绘制了该片段的物理图谱。该片段酶切产物经电泳、转膜后与探针 和探针 杂交 ,发现 2个探针杂交图谱基本相同 ,根据杂交带及放射信号的强度值 ,估测出该片段含有 4个 p MGA基因 ,以这个片段所含的 p MGA基因数为标准 ,估算出鸡毒霉形体 HS株含有 39个 p MGA基因。The pMGA multigene family encodes variant copies of the cell surface haemagglutinin of Mycoplasma gallisepticum. In order to screen these fragments containing pMGA genes and estimate the number of the pMGA multigene family in genome of Mycoplasmas gallisepticum HS strain (MG HS), a genomic library of M.gallisepticum HS strain was constructed by cloning the EcoRⅠ restriction endonuclease fragments of MG HS DNA into plasmid pUC 18 . The synthetic oligonucleotide probe Ⅰ which complementary to a encoding sequence conserved in all characterized pMGA genes and the probe Ⅱ which hybridizes to an intergenic repeat motif found between all charactized pMGA genes were used to screen the genomic library and attained 38 positive clones from 600 positive recombinant plasmids. The physical map of one EcoRⅠ fragment(7.5 kb) from positive clones of M.gallisepticum HS strain was constructed by digested with combination restriction endonuclease EcoRⅠ, HindⅢ, PstⅠ, BglⅠ. These fragments digested with restriction enzyme were hybridized with probe Ⅰ and probe Ⅱ in order to estimate the number of the pMGA of this 7.5 kb EcoRⅠ fragment. The results indicated that the 7.5 kb fragment contained 4 pMGA genes. With the 7.5 kb fragment as a standard, 39 pMGA genes in genome of M.gallisepticum HS strain was estimated.

关 键 词：HS株 鸡毒霉形体 pMGA基因 多基因族 

分 类 号：S858.31[农业科学—临床兽医学] S852.62[农业科学—兽医学]