伤寒沙门菌质粒pR_(ST98)的限制性核酸内切酶分析

Molecular Analysis of Plasmid pR_(ST98) from Multidrug Resistant Salmonella typhi by Restriction Endonuclease Diagestion

目的 以我国特有的多重耐药伤寒沙门菌中分离的质粒pRST98为研究对象 ,对其除编码细菌抗药性外还能使宿主菌毒力增强这一独特性状产生的分子基础 ,用限制性核酸内切酶进行核酸酶谱分析。方法 将质粒pRST98用QIAGEN试剂盒提纯并精确定量后 ,选用常用的 9种限制性核酸内切酶消化 ,制作该质粒的酶切图谱。结果 用试剂盒提纯质粒 ,再用限制性内切酶消化后得到了清晰、准确的质粒酶切图谱 ,其中BglⅡ、EcoRⅤ、PstⅠ和SacⅡ是对该质粒进行分子生物学和分子流行病学研究的理想工具。结论 伤寒沙门菌多效性质粒 pRST98限制性核酸内切酶谱的建立 ,为分析该质粒的遗传特征、流行病学追踪和基因功能定位奠定了基础。

Objective To Characterize plasmid (pR ST98 ) encoding both resistant to antimicrobial agents and mediating virulence to its host bacteria in S.typhi by restriction endonuclease digestion.Methods Plasmid pR ST98 was isolated and purified by using QIAGEN kit. Nine restriction endonuc leases (BamHI, BglⅡ,EcoRI,EcoRⅤ,HindⅢ,PstⅠ,SacⅠ,SacⅡand SphⅠ) were selected to make the enzyme profile by digestion with pR ST98 DNA.Results The results showed that when using the kit we could get enough and purified plasmid DNA,which was very important in getting an identical restriction pattern. Endonucleases BglⅡ, EcoRⅤ, PstⅠand SacⅡ were especially suitable for mole cular analysis of pR ST98 and were very useful tools for further research.Conclusion The restriction endonuclease pattern of pR ST98 is of much help in character molecular markings, epidemiological surverillence and the location of the functional genes of the plasmid.