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[用假T-偶数噬菌体转导质粒抗生素抗性决定簇]

[Transduction of plasmid antibiotic resistance determinants with pseudo-T-even bacteriophages].

作者信息

Taniashin V I, Zimin A A, Shliapnikov M G, Boronin A M

机构信息

Skryabin Institute of Biochemistry and Physiology of Microorganisms, Russian Academy of Sciences, Moscow Oblast, Pushchino, 142290 Russia.

出版信息

Genetika. 2003 Jul;39(7):914-26.

Abstract

Transduction of antibiotic resistance determinants of the plasmid pBR322 with pseudoT-even bacteriophages RB42, RB43, and RB49 was studied. It is established that antibiotic resistance determinants of plasmid pBR322 from Escherichia coli recA(+)- and recA(-)-donor strains do not differ significantly in respect to the efficiency of transduction. Amber mutants RB43-21, RB43-33, and a double amber mutant RB43am21am33 were obtained. These mutants facilitated transduction experiments in some cases. Transduction of antibiotic resistance markers of the vector plasmid pBR325 and recombinant plasmid pVT123, containing a DNA fragment with hoc segE uvsW genes of phage T4, was studied. The frequency of appearance of transductants resistant to pseudoT-even bacteriophages used in transduction was determined, and the sensitivity of resistant transductants to 32 RB bacteriophages and also to phages lambda, T2, T4, T5, T6, T7, and BF23 was estimated. The efficiency of plating pseudoT-even bacteriophages RB42 and RB43 on strain E. coli 802 himA hip carrying mutations in genes that encode subunits of the Integration Host Factor (IHF) was shown to be higher than on isogenic strain E. coli 802. The growth of pseudoT-even bacteriophages limited in vivo by modification-restriction systems of chromosomal (EcoKI, EcoBI), phage (EcoP1I), and plasmid (EcoRI, EcoR124I, and EcoR124II) localization was analyzed. It was shown that these phages were only slightly restricted by the type I modification-restriction systems EcoBI, EcoR124I, and EcoR124II. Phage RB42 was restricted by systems EcoKI, EcoP1I, and EcoRI; phage RB43, by systems EcoKI and EcoRI; and phage RB49, by the EcoRI modification-restriction system.

摘要

研究了用类T-偶数噬菌体RB42、RB43和RB49转导质粒pBR322的抗生素抗性决定簇。已确定来自大肠杆菌recA(+)和recA(-)供体菌株的质粒pBR322的抗生素抗性决定簇在转导效率方面没有显著差异。获得了琥珀突变体RB43-21、RB43-33和双琥珀突变体RB43am21am33。在某些情况下,这些突变体促进了转导实验。研究了载体质粒pBR325和含有噬菌体T4的hoc segE uvsW基因的DNA片段的重组质粒pVT123的抗生素抗性标记的转导。测定了转导中使用的对类T-偶数噬菌体有抗性的转导子出现的频率,并估计了抗性转导子对32种RB噬菌体以及对噬菌体λ、T2、T4、T5、T6、T7和BF23的敏感性。结果表明,类T-偶数噬菌体RB42和RB43在编码整合宿主因子(IHF)亚基的基因中携带突变的大肠杆菌802 himA hip菌株上的平板接种效率高于同基因的大肠杆菌802菌株。分析了在体内受染色体(EcoKI、EcoBI)、噬菌体(EcoP1I)和质粒(EcoRI、EcoR124I和EcoR124II)定位的修饰-限制系统限制的类T-偶数噬菌体的生长。结果表明,这些噬菌体仅受到I型修饰-限制系统EcoBI、EcoR124I和EcoR124II的轻微限制。噬菌体RB42受到EcoKI、EcoP1I和EcoRI系统的限制;噬菌体RB43受到EcoKI和EcoRI系统的限制;噬菌体RB49受到EcoRI修饰-限制系统的限制。

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