[Construction of genomic library of L. interrogans serovar lai using lambda gt11 as the vector and a study of recombiant plasmid pDL121].
A genomic library of L. interrogans serovar lai strain 017 has been constructed using lambda gt11 as the vector. DNA was partially digested by two blunt-end restriction enzymes, then methylated with EcoR I methylase; after EcoR I linker was added to the DNA, the linker-ended DNA was ligated to the dephosphorylated EcoR I digested lambda gt11 arms. The recombined DNA was packaged in vitro, and used to transduct E. coli Y1090 for amplification. There were 2.1 x 10(6) recombinant bacteriophages as recognized by their ability to form white plaques plated on Lac host in the presence of both IPTG and X-Ga1. A positive clone, designated lambda DL12, was screened with a rabbit anti-serum against L. interrogans serovar lai from the genomic library. The DNA from lambda DL12 was subcloned into plasmid pUC18. A recombinant (designated as pDL121) was obtained. SDS-PAGE analysis indicated that a 23 kd was expressed in E. coli JM 103 harboring pDL121. Western blotting analysis showed that a specific protein band molecular weight of 23 kd could be recognized by the rabbit antiserum against L. interrogans serovar lai strain 017.
Liu
H
H
Research Group of West China University Medical Science, Chengdu.
Dai
B
B
Jing
B
B
Wu
W
W
Li
S
S
Fang
Z
Z
Zhao
H
H
Ye
D
D
Yan
R
R
Liu
J
J
Song
S
S
Yang
Y
Y
Zhang
Y
Y
Liu
F
F
Tu
Y
Y
Yang
H
H
Huang
Z
Z
Liang
L
L
Hu
L
L
Zhao
M
M
D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000235:genetics*; D004274:DNA, Recombinant / Q000096:biosynthesis*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005822:Genetic Vectors; D015698:Genomic Library; D007921:Leptospira interrogans / Q000235:genetics*; D010957:Plasmids; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis*; D014162:Transfection
D004269:DNA, Bacterial; D004274:DNA, Recombinant; D011993:Recombinant Fusion Proteins