[Prokaryotic expression and identification of human papillomavirus type 16 E7 protein encoding gene].
OBJECTIVE
To construct the prokaryotic expression plasmid pET32/E7 and express the human papillomavirus type 16 E7 protein in E. coli.


METHODS
HPV16 E7 gene was amplified by PCR. The amplified E7 fragment was inserted into the plasmid pET32a (+) that was digested with BamH I and Hind III. The recombinant plasmid pET32/E7 was transformed into E. coli JM109 which was selected with ampicillin. The positive clones containing recombinant plasmid pET32/E7 were verified by BamH I and Xho I digestion, and then sequenced. HPV16 E7-TRX recombinant protein expression in the E. coli BL21(ED3) was identified by SDS-PAGE and Western blot.


RESULTS
The prokaryotic recombinant plasmid pET32/E7 was successfully constructed. The BL21(DE3) transformed recombinant plasmid pET32/E7 had expressed HPV16 E7-TRX recombinant protein effectively. Under the conditions of 1 mmol/L IPTG and 30 degrees C, the amount of HPV16 E7-TRX recombinant protein was about 30% of bacterial total proteins.


CONCLUSIONS
The construction of the prokaryotic recombinant plasmid pET32/E7 and the successful expression of the recombinant protein HPV16 E7-TRX would strongly promote the research of the biological properties and the transformational mechanism of the HPV16 E7 protein on the specific cells.
Shi
Qiao-Fa
QF
Department of Microbiology, West China School of Preclinical and Forensic Medicine, Sichuan University, Chengdu 610041, China.
Wang
Bao-Ning
BN
Li
Hong
H
Wei
Xiao-Li
XL
Zhang
Wei-Dong
WD
Cao
Kang
K
Jiang
Zhong-Hua
ZH
Li
Ming-yuan
MY
D000328:Adult; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005260:Female; D052162:Human papillomavirus 16 / Q000235:genetics / Q000302:isolation & purification*; D006801:Humans; D050725:Papillomavirus E7 Proteins / Q000096:biosynthesis* / Q000235:genetics; D010957:Plasmids / Q000235:genetics; D011387:Prokaryotic Cells / Q000378:metabolism*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics
D050725:Papillomavirus E7 Proteins; D011994:Recombinant Proteins