目的探讨随机引物PCR(Arbitrary primer PCR,AP-PCR)结合克隆测序及生物信息学分析等方法在研究幽门螺杆菌(Helicobacter pylori,H.pylori)菌株地域起源特征中的价值和意义。方法针对临床分离培养的H.pylori菌株的基因组DNA,采用一组10...目的探讨随机引物PCR(Arbitrary primer PCR,AP-PCR)结合克隆测序及生物信息学分析等方法在研究幽门螺杆菌(Helicobacter pylori,H.pylori)菌株地域起源特征中的价值和意义。方法针对临床分离培养的H.pylori菌株的基因组DNA,采用一组10 nt的寡核苷酸引物进行随机PCR扩增,选取相对保守的片段进行回收、克隆及测序,测序的基因序列提交GenBank数据库进行序列相似性的BLAST比对,收集BLAST比对得到的同源性较高不同地域来源螺杆菌的对应序列,用ClustalX软件进行排序,采用Mega 4.0软件中的邻位相连法(Neighbor-joining)和最大简约法(Maximum-parsimony)进行进化树分析。结果随机引物扩增及筛选克隆测序得到的基因产物为NADH脱氢酶G和H亚单位的部分编码序列,与27株不同地域来源H.pylori及1株猫科动物来源螺杆菌菌株的同源性均高达90%以上,表明H.pylori中NADH脱氢酶基因序列为保守结构,进化树分析显示:采用AP-PCR方法得到的H.pylori临床菌株的基因序列,显示出东亚菌株来源的遗传特征,与具有东亚菌株特征的美洲秘鲁Sat464和Shi470菌株、韩国的52、51菌株、日本的F57菌株遗传距离较近,与南亚、欧洲菌株距离较远,与非洲的SouthAfrica7菌株和猫科动物来源的Sheeba菌株的遗传距离最远。结论不仅某些特殊基因可以反映地域差异,随机定位相对保守的基因片段同样可以反映H.pylori的地域起源特征。AP-PCR、测序等技术方法与进化树分析相结合是探讨H.pylori地域起源特征的一种更为便捷有效的新方法。展开更多
The arbitrarily primed polymerase chain reaction(AP-PCR)technique was used to detect polymorphisms in genomic DNA from Oryza minuta, O. officinalis, and related rice species. The results show that the genetic distance...The arbitrarily primed polymerase chain reaction(AP-PCR)technique was used to detect polymorphisms in genomic DNA from Oryza minuta, O. officinalis, and related rice species. The results show that the genetic distance between O.minuta and O.officinalis is much higher than that between two known species,O.sativa and O. rufipogon, showing that O.minuto and O.officinalis should be classified as two different spedes in genus Oryza,and that the AP-PCR technique can be used as a powerful tool for studying taxonomic relationships among related spedes, and even subspecies.展开更多
文摘The arbitrarily primed polymerase chain reaction(AP-PCR)technique was used to detect polymorphisms in genomic DNA from Oryza minuta, O. officinalis, and related rice species. The results show that the genetic distance between O.minuta and O.officinalis is much higher than that between two known species,O.sativa and O. rufipogon, showing that O.minuto and O.officinalis should be classified as two different spedes in genus Oryza,and that the AP-PCR technique can be used as a powerful tool for studying taxonomic relationships among related spedes, and even subspecies.
