Expression of simian virus 40 small t antigen in Escherichia coli and purification of the antigen.
A simian virus 40 (SV 40) DNA fragment encoding small t antigen was cloned in expression vector pUC8 for the purification of the antigen. The SV40 Hind III B fragment was inserted into the Hind III site of pUC8. A plasmid having the lacZ' and small t antigen genes in the same orientation was designated as pSVt. pSVt encodes the entire small t antigen and an extra 18 amino acids at the amino terminus of the antigen. E. coli transformed with pSVt produced hybrid small t antigen (22 kDa) which comprised about 6% of the total protein. The hybrid small t antigen reacted in immunoblot analysis with SV40-induced tumor-bearing hamster serum. Hybrid small t antigen was extracted from E. coli, and purified by preparative SDS-PAGE. The antigen was extracted from the gel using formic acid solution with high-yield. The gel-purified antigen showed the same antigenic reactivity as crude antigen.
Ikeda
S
S
Department of Biochemistry, Cancer Institute, Okayama University Medical School, Japan.
Akiyama
K
K
Mitsunobu
F
F
Wada
T
T
Hatsushika
M
M
Watanabe
S
S
Tsutsui
K
K
Seki
S
S
Oda
T
T
D000595:Amino Acid Sequence; D000957:Antigens, Viral, Tumor / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification; D001483:Base Sequence; D004279:DNA, Viral / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics / Q000276:immunology*; D005786:Gene Expression Regulation; D009693:Nucleic Acid Hybridization; D010957:Plasmids; D013539:Simian virus 40 / Q000235:genetics / Q000276:immunology*
D000957:Antigens, Viral, Tumor; D004279:DNA, Viral