AI Article Synopsis
- The study aimed to create and purify a recombinant plasmid named pGEX-4T-1-GRP78 for the expression of human glucose regulated protein 78 (GRP78).
- The GRP78 gene was successfully amplified via PCR, inserted into a prokaryotic expression vector, and expressed in bacterial cells after induction with IPTG, followed by purification and verification through ELISA.
- The outcomes include successfully constructing the recombinant vector and obtaining purified GRP78, which shows potential for use in diagnosing hepatocellular carcinoma.
Article Abstract
Aim: To construct a recombinant plasmid pGEX-4T-1-GRP78, express it and purify human glucose regulated protein 78 (GRP78).
Methods: GRP78 gene was amplified by PCR from the recombinant vector constructed in our laboratory. The PCR product was inserted into pGEX-4T-1 prokaryotic expression vector to generate pGEX-4T-1-GRP78. The pGEX-4T-1-GRP78 was then transformed into BL21 and GRP78 was expressed on induction of IPTG. After purification, GRP78 was released by thrombin cleavage, and its antigenicity was identified by ELISA.
Results: The GRP78 prokaryotic expression vector was successfully constructed as confirmed by enzyme digestion and DNA sequencing. ELISA demonstrated the antigenicity of the purified GRP78 protein.
Conclusion: The recombinant prokaryotic expression vector pGEX-4T-1-GRP78 has been constructed successfully. The purified GRP78 has been obtained with good antigenicity, which will be used for GRP78-based serum diagnosis of hepatocellular carcinoma.
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