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目的赖氨酸、鸟氨酸脱羧酶阴性、精氨酸双水解酶阳性和阿拉伯糖发酵实验是区分河弧菌和其他非凝集弧菌的特征性化学指标。本研究对发现的1株精氨酸双水解酶阴性的河弧菌Ma-2531进行了分子遗传学分析。方法利用多对特异性引物对精氨酸双水解酶代谢通路的arc操纵子区基因簇进行普通聚合酶链反应(PCR)扩增,并对产物进行测序分析,同时比较Ma-2531和精氨酸双水解酶阳性菌株EF85003的生长曲线和培养液pH值的变化。结果针对arcBCA基因簇序列的3对引物arc-F/arc-R、arc-F/arc-rev和arc-ck-up/arc-R均为阴性扩增,而其上下游序列特异性的引物对arc1-up/arc1-dn和arc72695-up/arc75219-dn分别扩增出预期大小的目的条带。arc1-dnRev/arc72695-upRev引物对的扩增产物序列分析显示该扩增片段包含transposase IS4序列,进一步的对比分析表明包括精氨酸双水解酶系统arcBCAD基因簇及上游的天冬氨酸氨甲酰转移酶调节亚基和催化亚基编码基因在内的约8.6 kb的大片段发生了缺失。生长曲线和pH值测定显示菌株Ma-2531和EF85003没有明显差异。结论菌株Ma-2531中插入序列的转座导致了该菌株精氨酸双水解酶这一鉴定生化指标阴性的结果,但这种变化应只是随机和罕见的,使河弧菌生化代谢表现出多样性。
The purpose of lysine, ornithine decarboxylase negative, arginine double hydrolase positive and arabinose fermentation experiment is to distinguish between Vibrio River and other non-agglutinating Vibrio characteristic chemical indicators. In this study, a molecular genetic analysis was carried out on one of the arginine dihydrolase negative strains, Vibrio cholerae Ma-2531. Methods Multiple arc pairs of specific primers were used to amplify the arc operon region of arginine hydrolase metabolic pathway by polymerase chain reaction (PCR). The products were sequenced and compared. Meanwhile, Ma-2531 and arginine Growth curve of acid double hydrolase positive strain EF85003 and the change of pH value of culture medium. Results Three pairs of primers arc-F / arc-R, arc-F / arc-rev and arc-ck-up / arc-R directed against the arcBCA gene cluster were all negative amplification, while the upstream and downstream sequence- The expected size of the target band was amplified for arc1-up / arc1-dn and arc72695-up / arc75219-dn, respectively. Sequence analysis of the amplified product of the arc1-dnRev / arc72695-upRev primer pair revealed that the amplified fragment contained the transposase IS4 sequence. Further comparative analysis indicated that the arcBCAD gene cluster including arginine dihydrolase system and upstream aspartate ammonia methyl A large fragment of about 8.6 kb, including the acyltransferase regulatory and catalytic subunit genes, was deleted. Growth curves and pH measurements showed no significant difference between strains Ma-2531 and EF85003. Conclusion The transposition of the inserted sequence in strain Ma-2531 resulted in the negative result of arginine dihydrolase, a biochemical marker of this strain. However, this change should only be random and rare, Sex.