[Expression of mouse canstatin and its N-domain in E. coli BL21].
The mouse canstatin and its N-domain cDNA were amplified from total RNA of mouse liver by RT- PCR and cloned into vector pMD18-T for sequencing. Prokaryotic expression vectors pET/Can and pET/Can-N were constructed and expressed in E.coli BL21(DE3) with induction of IPTG.. Mouse canstatin cDNA is 684bp in length encoding 227 amino acids. The sequences of both cDNA and amino acids share high homology with human canstatin, with cDNA identity at 89% and amino acids identity at 96% to human canstatin. N-domain of mouse canstatin is the same amino acid sequence as that of human canstatin. In the present study, prokaryotic expression vector pET/Can and pET/Can-N were expressed in E.coli BL21 with amount of 35% and 18% of the total bacterial proteins after being induced by IPTG for 4h. The expressed products existed mainly as inclusion bodies. This work has laid down the basis for further study of its angiogenic activity and potential application for tumor dormancy therapy.
Hou
Wei-Hong
WH
Laboratory for Cell Biology, Zhengzhou University, Zhengzhou 450 052, China. whhou@zzu.edu.cn
Wang
Tian-Yun
TY
Chai
Yu-Rong
YR
Yuan
Bao-Mei
BM
Hou
Gui-Qin
GQ
Wang
Jian-Min
JM
Xue
Le-Xun
LX
D000595:Amino Acid Sequence; D020533:Angiogenesis Inhibitors / Q000096:biosynthesis* / Q000235:genetics; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D024141:Collagen Type IV / Q000096:biosynthesis* / Q000235:genetics; D018076:DNA, Complementary / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D005822:Genetic Vectors; D006801:Humans; D051379:Mice; D010446:Peptide Fragments / Q000096:biosynthesis* / Q000235:genetics; D010957:Plasmids; D017434:Protein Structure, Tertiary / Q000502:physiology; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D014162:Transfection
D020533:Angiogenesis Inhibitors; C502624:Col4a2 protein, mouse; D024141:Collagen Type IV; D018076:DNA, Complementary; D010446:Peptide Fragments; D011994:Recombinant Proteins