Gene cloning and fusion expression of nuclear antigen Sm B′in E. coli
- VernacularTitle:细胞核抗原Sm B′的基因克隆和原核表达
- Author:
Xiangyue YANG
;
Xiaopeng LAN
;
Jian LIAO
;
Zhiqiang ZHONG
;
Zhongyong ZHU
- Publication Type:Journal Article
- Keywords:
nuclear antigen;
cloning;
molecular;
gene fusion
- From:
Chinese Journal of Clinical Laboratory Science
2006;0(03):-
- CountryChina
- Language:Chinese
-
Abstract:
Objective To clone,express and identify the nuclear antigen Sm B′in E. coli to establish a new assay for detecting autoanti-body to Sm B′. Methods A full length cDNA of Sm B′was cloned from cell line HL-60 by RT-PCR. The PCR product was TA cloned and sequenced and inserted into the vector pGEX-5T. The recombinant plasmid was transformed into E. coli BL21. The positive clones were identified by restricted enzymes and induced by IPTG. The expression product was analyzed by SDS-PAGE and Western blot. Results The PCR product was about 700 bp in size which was in accordance with predicted 657 bp and sequencing result showed consistent with the sequence in GenBank. The pGEX-5T-Sm B′positive clone produced a 51 000 kD of fusion protein which was immunoreac-tive with anti-Sin B′confirmed by SDS-PAGE and Western blot. Conclusion The successful cloning and expression of nuclear antigen Sm B′laid a foundation for further research work.
