2.2. Construction and identification of recombinant human-like
collagen
The gene encoding RHC was cloned
into the pET-3c expression vector to generate a recombinant plasmid
named pET3c-hlcollagen, which was transformed into E. coliBL21(DE3 ). After screening for ampicillin resistance and
induction by IPTG, the best expression condition was selected. Lager
scale production of RHC was performed using a 50L fermenter. And RHC
protein was purified using affinity chromatography on a NI Sepharose 6
Fast Flow column combined with gel filtration Sephadex G-25. PCR,
western blot, and gel electrophoresis were used for identification of
RHC.