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Construction of signature-tagged mutant library in Mesorhizobium loti as a powerful tool for functional genomics.


ABSTRACT: Rhizobia are nitrogen-fixing soil bacteria that establish endosymbiosis with some leguminous plants. The completion of several rhizobial genome sequences provides opportunities for genome-wide functional studies of the physiological roles of many rhizobial genes. In order to carry out genome-wide phenotypic screenings, we have constructed a large mutant library of the nitrogen-fixing symbiotic bacterium, Mesorhizobium loti, by transposon mutagenesis. Transposon insertion mutants were generated using the signature-tagged mutagenesis (STM) technique and a total of 29,330 independent mutants were obtained. Along with the collection of transposon mutants, we have determined the transposon insertion sites for 7892 clones, and confirmed insertions in 3680 non-redundant M. loti genes (50.5% of the total number of M. loti genes). Transposon insertions were randomly distributed throughout the M. loti genome without any bias toward G+C contents of insertion target sites and transposon plasmids used for the mutagenesis. We also show the utility of STM mutants by examining the specificity of signature tags and test screenings for growth- and nodulation-deficient mutants. This defined mutant library allows for genome-wide forward- and reverse-genetic functional studies of M. loti and will serve as an invaluable resource for researchers to further our understanding of rhizobial biology.

SUBMITTER: Shimoda Y 

PROVIDER: S-EPMC2575893 | biostudies-other | 2008 Oct

REPOSITORIES: biostudies-other

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Construction of signature-tagged mutant library in Mesorhizobium loti as a powerful tool for functional genomics.

Shimoda Yoshikazu Y   Mitsui Hisayuki H   Kamimatsuse Hiroko H   Minamisawa Kiwamu K   Nishiyama Eri E   Ohtsubo Yoshiyuki Y   Nagata Yuji Y   Tsuda Masataka M   Shinpo Sayaka S   Watanabe Akiko A   Kohara Mitsuyo M   Yamada Manabu M   Nakamura Yasukazu Y   Tabata Satoshi S   Sato Shusei S  

DNA research : an international journal for rapid publication of reports on genes and genomes 20080725 5


Rhizobia are nitrogen-fixing soil bacteria that establish endosymbiosis with some leguminous plants. The completion of several rhizobial genome sequences provides opportunities for genome-wide functional studies of the physiological roles of many rhizobial genes. In order to carry out genome-wide phenotypic screenings, we have constructed a large mutant library of the nitrogen-fixing symbiotic bacterium, Mesorhizobium loti, by transposon mutagenesis. Transposon insertion mutants were generated u  ...[more]

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