Prokaryotic expression and bioactivity analysis of insulin-like growth factor-I from Hucho taimen
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Graphical Abstract
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Abstract
Insulin-like growth factor-I (IGF-I) plays a major role in the control of growth and development in fish. Total RNA was isolated from H. taimen liver tissue. The cDNA encoding insulin-like growth factor I (IGF-I) peptide was amplified by reverse transcription polymerase chain reaction (RT-PCR) strategy using isolated total RNA as template. Recombinant expression plasmid (IGF-I/pS) was constructed by prokaryotic expression vector pS and transformed into E. coli Rosetta. The expression of recombinant IGF-I protein of H. taimen was induced by IPTG. There was a clear target band with expected size between 35~50 kD in SDS-PAGE electrophoresis, and the recombinant protein existed in the form of inclusion body. Purified IGF-I protein was obtained after the inclusion body was denatured/renatured. ELISA results showed that target protein can identify commercial fish IGF-I antibody, indicating that H. salmon IGF-I protein with immune activity was obtained. Cell proliferation assay (MTT) results showed that recombinant IGF-I protein can significantly promote the propagation of CHSE-214, EPC and RTG-2, indicating that the recombinant IGF-I protein had biological activity. The study has laid a foundation for the research of IGF-I function in the growth and development of H. taimen.
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