[Construction of the recombinant plasmid FasAD-pTYB2 capable of Fas activation domain fusion expression].
OBJECTIVE
To construct the recombinant plasmid FasAD-pTYB2 for the fusion expression of Fas activation domain (FasAD).


METHODS
FasAD cDNA was cloned by semi-nested reverse transcriptase-polymerase chain reaction (RT-PCR), and then inserted into the universal vector of pGEM-T for the identification of its DNA sequence. The target DNA fragment was inserted into the prokaryote vector pTYB2 that expressed intein to construct the plasmid recombinant FasAD-pTYB2 which was induced by isopropylthio-beta-D-galactoside for the expression of the fusion protein.


RESULTS
DNA sequence analysis confirmed sequence of FasAD cDNA previously cloned, and expression of the fusion protein by the recombinant plasmid was achieved, the product recognizable by rabbit anti-human Fas polyclonal antibody as demonstrated by Western blotting analysis.


CONCLUSIONS
The recombinant plasmid FasAD-pTYB2 constructed in this study is capable of expressing the fusion protein of FasAD and intein, which may be significant for further study of the epitope and function of Fas antigen.
Feng
Xiao-Qin
XQ
Department of Pediatrics, Nanfang Hospital, First Military Medical University, Guangzhou 510515, China.
Wang
Xiao-Ning
XN
Ning
Yun-Shan
YS
Zhou
Shu-Yun
SY
D017209:Apoptosis / Q000235:genetics; D001665:Binding Sites / Q000235:genetics; D015153:Blotting, Western; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D015870:Gene Expression; D010957:Plasmids / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D019014:fas Receptor / Q000235:genetics* / Q000378:metabolism
D018076:DNA, Complementary; D011993:Recombinant Fusion Proteins; D019014:fas Receptor