| Products/Services Used | Details | Operation |
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| Peptide Synthesis | According to our previous study [31], a gene fragment encoding a ι-carrageenase without a signal peptide from M. thermotolerans (CGIA, accession No. AB439728) [14] was analyzed and redesigned, including the optimization of the rare codons used in B. choshinensis and the elimination of the negative cis-acting elements. Subsequently, this optimized ι-carrageenase gene (CGIOP) (Accession No. MF459059) with two restriction sites (BamHI and KpnI) and a 6×His tag was synthesized by Genscript (Nanjing, China) (Supplementary Fig. S1), and then was ligated into the BamHI/KpnI site of the integrative vector pBCGA to construct the recombinant plasmid pBCGA-CGIOP (Fig. 1). As a control, the gene CGIOP was also ligated into the BamHI/KpnI site of the replicative plasmid pNCMO2 (Takara Bio Inc., Dalian, China), resulting in a recombinant plasmid pNCMO2-CGIOP. | Get A Quote |
In this study, a new integrative vector (pBCGA) was developed and used for the high-level expression of an optimized ι-carrageenase gene (CGIOP) in Brevibacilluschoshinensis. The pBCGA vector allowed multiple copies of the gene CGIOP to be stably integrated into the genomic DNA of B. choshinensis. The recombinant strain I24 could produce an extracellular ι-carrageenase activity of 38.9 U/mL within 72 h, which remained stable after five sequential inoculations and cultivations under the antibiotic-free culture conditions. Furthermore, the strain I24 was applied to 10-L fermentation under the antibiotic-free culture condition, resulting in the highest observed ι-carrageenase activity of 182.4 U/mL with... More
