RRC ID |
61165
|
著者 |
Kuroki A, Ohtani N, Tsuge K, Tomita M, Itaya M.
|
タイトル |
Conjugational transfer system to shuttle giant DNA cloned by Bacillus subtilis genome (BGM) vector.
|
ジャーナル |
Gene
|
Abstract |
The Bacillus subtilis GenoMe (BGM) vector was designed as a versatile vector for the cloning of giant DNA segments. Cloned DNA in the BGM can be retrieved to a plasmid using our Bacillus recombinational transfer (BReT) method that takes advantage of competent cell transformation. However, delivery of the plasmid to a different B. subtilis strain by the normal transformation method is hampered by DNA size-related inefficiency. Therefore, we designed a novel method, conjugational plasmid-mediated DNA retrieval and transfer (CReT) from the BGM vector, and investigated conjugational transmission to traverse DNA between cells to circumvent the transformation-induced size limitation. pLS20, a 65-kb plasmid capable of conjugational transfer between B. subtilis strains, was modified to retrieve DNA cloned in the BGM vector by homologous recombination during normal culture. As the plasmid copy number was estimated to be 3, the retrieval plasmid was selected using increased numbers of marker genes derived from the retrieved DNA. We applied this method to retrieve Synechocystis genome segments up to 90 kb in length. We observed retrieved plasmid transfers between B. subtilis strains by conjugation in the absence of structural alterations in the DNA fragment. Our observations extend DNA transfer protocols over previously exploited size ranges.
|
巻・号 |
399(1)
|
ページ |
72-80
|
公開日 |
2007-9-1
|
DOI |
10.1016/j.gene.2007.04.030
|
PII |
S0378-1119(07)00247-8
|
PMID |
17560740
|
MeSH |
Bacillus subtilis / genetics*
Cloning, Molecular / methods*
Conjugation, Genetic
Gene Transfer Techniques*
Genetic Vectors / genetics*
Genome, Bacterial / genetics
Plasmids / chemistry
Plasmids / genetics*
Synechocystis / genetics
|
IF |
2.984
|
リソース情報 |
原核生物(枯草菌) |
MBS794
MBS795
MBS796
MBS797
MBS798
MBS799
MBS800
MBS801
MBS802
MBS803
MBS804
MBS805
MBS806 |