| Products/Services Used | Details | Operation |
|---|---|---|
| Gene Synthesis | The vectors and primers used are listed in Tables 1 and S1, respectively. The hEGF gene was synthesized by GenScript, Inc., USA (www.genscript.com) and was constructed into the integration vector pDMT1 [28] to generate pDMT1-hEGFH using XhoI and BamHI, and the C-terminal of hEGF was fused with a His-tagged sequence to facilitate purification. Twenty-four SP sequences were amplified from the chromosome of B. subtilis using polymerase chain reaction (PCR) with specific primers. The aprE SP was removed from the vector pDMT1-hEGFH by NruI and XhoI, and the PCR products of SP sequences were separately cloned into the digested vector to generate 24 various integration vectors bearing different SPs. These vectors were transformed as described previously [26] and integrated into the B. subtilis PT5 chromosome using the double-crossover homologous recombination method to generate rhEGF-producing strains. | Get A Quote |
Human epidermal growth factor (hEGF) is a crucial factor in wound healing, and therefore has wide applications in the pharmaceutical and cosmetics industries. To improve the production of hEGF protein for the industrial scale, we established an hEGF gene construct for recombinant secretion by Bacillus subtilis. Examination of the secretion abilities of various signal peptides (SPs) revealed six SPs that could effectively guide hEGF secretion from B. subtilis into the culture medium. The highest production level was obtained when the hEGF gene was fused with the xynDSP (lipo type). Two copies of the hEGF expression cassette had the same effect on improving production yield as isopropyl-β-d-1-thiogalactopyra... More
