Expression of functional domain of chicken gizzard calponin.
A full-length cDNA of the function domain of wild-type chicken gizzard calponin was cloned into expression vector pAED4 and the recombinant function domain of wild-type calponin was expressed in an Escherichia coli expression system. The actin domain of calponin (CaP-B) can bind with actin and it is a requisite for its inhibition of ATPase and vasoconstriction of smooth muscle. In this study, the cDNA of CaP-B was inserted into vector pAED4 by direction-cloning method. The cDNA of CaP-B was obtained with PCR cloning technique. The recombinant DNA pAED4-Cap-B was transformed into E. coli BL21 (DE3) and identified with the restriction analysis. The bacterial clones containing transformants were induced to be highly expressed in E. coli BL21 (DE3). The target protein was detected and identified by Western Blot analysis. The content of target protein was as high as 10% of the whole protein after overnight (16 h) culture. The results confirmed that Cap-B was relatively highly expressed in E. coli.
Xiang
J
J
Key Laboratory of Pulmonary Diseases of National Health Ministry, Tongji Medical University, Wuhan.
Lu
Z
Z
Hu
B
B
Lin
X
X
Cao
S
S
Tang
D
D
D000818:Animals; D002135:Calcium-Binding Proteins / Q000096:biosynthesis* / Q000235:genetics; D002645:Chickens; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D005895:Gizzard, Avian / Q000378:metabolism; D008840:Microfilament Proteins / Q000096:biosynthesis* / Q000235:genetics; D009130:Muscle, Smooth / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D017434:Protein Structure, Tertiary / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis; D000096985:Calponins
D002135:Calcium-Binding Proteins; D008840:Microfilament Proteins; D011994:Recombinant Proteins; D000096985:Calponins