Method and application of high efficient preparation of procalcitonin standby reference material based on protein recombination technology
10.3760/cma.j.cn114452-20211216-00780
- VernacularTitle:蛋白重组技术在制备视黄醇结合蛋白质控品和候选参考物质中的方法研究
- Author:
Chao ZHANG
1
;
Weiyan ZHOU
;
Jiangtao ZHANG
;
Jing WANG
;
Chuanbao ZHANG
Author Information
1. 北京医院 国家老年医学中心 国家卫生健康委临床检验中心 中国医学科学院老年医学研究院，北京100730
- Keywords:
Retinol-binding proteins;
Recombinant proteins;
Reference material;
Quality control product;
External quality assessment
- From:
Chinese Journal of Laboratory Medicine
2022;45(11):1131-1135
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To construct a prokaryotic expression vector for human retinol binding protein 4 (hRBP4) that allows technicians to obtain hRBP4 purified protein with low cost, high efficiency, high concentration and high purity.Methods:The hRBP4 coding sequence provided by National Center for Biotechnology Information was optimized by E. coli codons, and a synthetic DNA fragment was cloned into the PET-28A (+) prokaryotic expression vector to construct a recombinant hRBP4 expression plasmid. The recombinant protein was transformed into E. coli BL21, and the induced expression conditions (temperature, rotate speed and isopropyl β-d-thiogalactoside concentration) were optimized. The recombinant protein was purified by His fusion tag. Results:The recombinant hRBP4 prokaryotic expression plasmid was successfully constructed, and the expression concentration and induction temperature of the recombinant protein were optimized. The results of sodium dodecyl sulfate polyacrylamide gel electrophoresis showed that a band with a relative molecular weight of 26 000 daltons was clearly visible in the purified product. The purified hRBP4 protein could be detected clinically, and there was a good linear relationship between the dilution ratio and the detection concentration.Conclusions:The recombinant hRBP4 protein has high purity, high concentration, and short production cycle. It has the potential to become a candidate for reference materials for laboratory quality evaluations.
