Genetic transposition and insertional mutagenesis in Bacillus subtilis with Streptococcus faecalis transposon Tn917.
Genetic transposition and insertional mutagenesis in Bacillus subtilis with Streptococcus faecalis transposon Tn917.
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枯草芽孢杆菌与粪链球菌转座子 Tn917 的遗传转座和插入诱变。
DOI：
10.1073/pnas.80.8.2305
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发表时间：
1983
影响因子：
11.1
通讯作者：
Losick,R
中科院分区：
文献类型：
--
作者：
Youngman,PJ;Perkins,JB;Losick,R
通过用质粒pAM α 1：：Tn917转化感受态细胞将粪链球菌转座子Tn917引入枯草芽孢杆菌，并通过选择插入温和噬菌体SP β中来测试转座活性。插入获得的频率表明相对有效的移动的元素，和Southern杂交分析的一个特定的插入证实它是一个真正的转座事件的结果。将含有转座子序列的来自pAM α 1：：Tn917的限制性片段连接到温度敏感性质粒（pBD 95）中，并转座到B中。通过要求转座子耐药性保持在不允许质粒复制的温度下来选择枯草杆菌染色体。在许多染色体位点，包括产生不同种类的营养缺陷型的位点和产生各种不同的孢子形成缺陷型的位点，已经恢复了插入，这表明使用Tn917作为B中插入诱变的工具具有良好的前景。枯草杆菌。
The Streptococcus faecalis transposon Tn917 was introduced into Bacillus subtilis by transformation of competent cells with the plasmid pAM alpha 1::Tn917 and was tested for transposition activity by selection for insertions into the temperate phage SP beta. Insertions were obtained at a frequency indicating relatively efficient movement of the element, and Southern hybridization analysis of a particular insertion confirmed it to be the result of a genuine transposition event. A restriction fragment from pAM alpha 1::Tn917 containing the transposon sequences was ligated into a temperature-sensitive plasmid (pBD95), and transpositions into the B. subtilis chromosome were selected by requiring the transposon drug resistance to be maintained at temperatures nonpermissive for plasmid replication. Insertions have been recovered at many chromosomal sites, including ones that produced auxotrophy of different kinds and ones that produced various different sporulation-defective phenotypes, indicating good prospects for the use of Tn917 as a tool for insertional mutagenesis in B. subtilis.