[Cloning of DNA fragment related to salt tolerance in Sinorhizobium meliloti 042B].
Total DNA partially digested by EcoR I was prepared for S. meliloti 042B, in which 15-25 kb DNA fragments were collected. Vector pLAFR I was purified and digested by EcoR I, and then the various DNA fragments of 042B were ligated with pLAFR I by T4DNA ligase. Gene library of S. meliloti 042B was constructed with pLAFR I using E. coli S17-1 as recipient. The number of bacterial recombinants obtained was about 8,000 and 95% of them contained foreign DNA fragments. Using NTG, 042B was mutated on FY plates and 12 sensitive strains were screened at 0.5 mol/L NaCl from 2,000 colonies. One of them was named GZ17 and selected as a recipient strain. By biparental mating the foreign DNA fragments were introduced from gene library of strain 042B into recipient strain GZ17 which is sensitive to 0.5 mol/L NaCl. Then the transconjugants were grown on FY plates containing tetracycline (20 micrograms/mL) and 0.5 mol/L NaCl. A 7 kb inserted DNA fragment related to salt tolerance was obtained. In subcloning experiment, a 4 kb DNA fragment related to salt tolerance was obtained.
Chen
X
X
Department of Microbiology, College of Biological Sciences, China Agricultural University, Beijing 100094.
Zhang
H
H
Gao
W
W
Zhu
K
K
Kan
F
F
Yang
S
S
D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics; D005798:Genes, Bacterial; D015698:Genomic Library; D010957:Plasmids; D016962:Sinorhizobium meliloti / Q000187:drug effects / Q000235:genetics*; D012965:Sodium Chloride / Q000494:pharmacology*
D004269:DNA, Bacterial; D012965:Sodium Chloride