Method for isolation of insertion elements from coryneform bacteria
    9.
    发明授权
    Method for isolation of insertion elements from coryneform bacteria 失效
    从棒状细菌中分离插入元件的方法

    公开(公告)号:US5380657A

    公开(公告)日:1995-01-10

    申请号:US33320

    申请日:1993-03-18

    摘要: A method locating insertion elements (IS elements) or transposons in coryneform bacteria, a positive selection system suitable for the above, the IS elements found in this manner and their use, is disclosed. The method involves:(1) The construction of a non-self-transferrable vector mobilizable from an E. coli mobilizer strain which vector is composed of(a) A DNA segment containing a replicon functional in E. coli,(b) A second DNA segment containing the DNA fragment coding for the mobilization function (Mob site containing the oriT),(c) A third DNA segment which recombines homologously in Gram-positive bacteria and/or contains a replicon functional in coryneform bacteria,(d) A DNA segment from Bacillus subtilis containing the sacB gens,(2) Transfer of this vector by means of conjugative transfer into the coryneform recipient strains,(3) Cultivation of the transconjugants containing the vector in an .about.10% sucrose-containing nutrient medium,(4) Lysis of the sucrose-resistant clones, cleaving of the plasmids with restriction endonucleases and analysis of the fragments.

    摘要翻译: 公开了一种在棒状细菌中定位插入元件(IS元件)或转座子的方法,适用于上述的正选择系统,以这种方式发现的IS元件及其用途。 该方法包括:(1)从大肠杆菌动员菌株可移动的非自转型载体的构建,其载体由(a)在大肠杆菌中含有复制子功能的DNA片段组成,(b)第二 含有编码动员功能的DNA片段(含有oriT的Mob位点)的DNA片段,(c)在革兰氏阳性细菌中同源重组和/或含有棒状细菌功能性复制子的第三DNA片段,(d)DNA (2)通过共轭转移将该载体转移到棒状细胞受体菌株中,(3)在含有10%蔗糖的营养培养基中含有载体的转导结合体的培养物(4 )分解蔗糖抗性克隆,用限制性内切核酸酶切割质粒并分析片段。

    Method for location of insertion elements
    10.
    发明授权
    Method for location of insertion elements 失效
    插入元件位置的方法

    公开(公告)号:US5633154A

    公开(公告)日:1997-05-27

    申请号:US336069

    申请日:1994-11-04

    摘要: A method of locating insertion elements (IS elements) or transposons in coryneform bacteria, a positive selection system suitable for the above, the IS elements found in this manner and their use, is disclosed. The method involves:(1) The construction of a non-self-transferrable vector mobilizable from an E. coli mobilizer strain which vector is composed of(a) A DNA segment containing a replicon functional in E. coli,(b) A second DNA segment containing the DNA fragment coding for the mobilization function (Mob site containing the oriT),(c) A third DNA segment which recombines homologously in Gram-positive bacteria and/or contains a replicon functional in coryneform bacteria,(d) A DNA segment from Bacillus subtilis containing the sacB gene,(2) Transfer of this vector by means of conjugative transfer into the coryneform recipient strains,(3) Cultivation of the transconjugants containing the vector in an .about.10% sucrose-containing nutrient medium,(4) Lysis of the sucrose-resistant clones, cleaving of the plasmids with restriction endonucleases and analysis of the fragments.

    摘要翻译: 公开了一种在棒状细菌中定位插入元件(IS元件)或转座子的方法,适用于上述的正选择系统,以这种方式发现的IS元件及其用途。 该方法包括:(1)从大肠杆菌动员菌株可移动的非自转型载体的构建,其载体由(a)在大肠杆菌中含有复制子功能的DNA片段组成,(b)第二 含有编码动员功能的DNA片段(含有oriT的Mob位点)的DNA片段,(c)在革兰氏阳性细菌中同源重组和/或含有棒状细菌功能性复制子的第三DNA片段,(d)DNA (2)通过共轭转移将该载体转移到棒状细胞受体菌株中,(3)在含有10%蔗糖的营养培养基中含有载体的转导结合体的培养物(4 )分解蔗糖抗性克隆,用限制性内切核酸酶切割质粒并分析片段。