Construction of signature-tagged mutant library in Mesorhizobium loti as a powerful tool for functional genomics.
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.
Shimoda
Yoshikazu
Y
Kazusa DNA Research Institute, 2-6-7 Kazusa-kamatari, Kisarazu, Chiba 292-0818, Japan.
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
D020561:Alphaproteobacteria / Q000235:genetics*; D004251:DNA Transposable Elements; D015723:Gene Library; D016680:Genome, Bacterial; D023281:Genomics; D016254:Mutagenesis, Insertional; D010957:Plasmids / Q000235:genetics; D012230:Rhizobiaceae / Q000235:genetics
D004251:DNA Transposable Elements