Prokaryotic expression,identification,primary purification of human prostate specific membrane antigen
- VernacularTitle:人前列腺特异性膜抗原cDNA的克隆、原核表达、鉴定和抗原初步纯化
- Author:
Hai HUANG
;
Jian HUANG
;
Tianxin LIN
;
Al ET
- Publication Type:Journal Article
- Keywords:
Prostatic neoplasms;
Carcinoma;
Prostate specific membrane antigen
- From:
Chinese Journal of Urology
2001;0(08):-
- CountryChina
- Language:Chinese
-
Abstract:
Objective To describe the cloning of cDNA of prostate specific membrane antigen (PSMA),construction of the recon of PSMA,induction of the expression of 6 ? His PSMA fusion protein from PSMS recon,and purification of the expression products. Methods The total RNA was extracted from prostate cancer tissues.The site of EcoR I lying in the middle of PSMA’s cDNA was used to perform the reverse transcriptase and polymerase chain reaction of PSMA in two different parts,which were then linked into the carrier of pEt 30(a),thus the full cDNA of PSMA and the pronucleus expression carrier pET 30(a) PSMA were obtained.The pET30a (+) PSMA recon was transformed into the Escherichia coli BL 21 and the engineering bacteria expressing protein PSMA was gotten. With the induction of IPTG, the recon was expressed. The expression products were purified with Ni NTA Agarose resin, then the purer protein PSMA was obtained. The antigenicity and specificity of the expressed PSMA were evaluated with Western blotting and SDS PAGE. Results The wholly right base sequence of PSMA’s cDNA was successfully cloned.PSMA protein was expressed and purified.This protein had better antigenicity and specificity. Conclusions This study provides experimental basis for the function study of PSMA and scanning of the phage antibody library.
