The presence of antibiotic resistance genes in genetically modified bacteria raises a regulatory concern in the production of therapeutic proteins and additionally reduces the number of plasmids available for propagation in a cell. Cre recombinase from bacteriophage P1, involved in Cre/loxP mechanism is one of the widely used systems for selectable marker gene removal. We have overexpressed codon-optimized cre gene in pColdIV and pET28a(+) vector systems and purified His6-Cre recombinase by immobilized metal affinity chromatography. N-terminal His6 tagged Cre recombinase obtained was approximately 26 fold purified and promoted the site-specific recombination of two loxP sites of linearized pLox2+ vector allowing the excision of a re-circularized plasmid and a short stretch of DNA containing the recombined loxP site. The results of the expression using two vectors, purification and activity assessment of His6 tagged Cre recombinase is presented here.

中文翻译：

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密码子优化的cre重组酶在大肠杆菌中的表达和纯化。

转基因细菌中抗生素抗性基因的存在引起了治疗性蛋白质生产中的监管问题，并另外减少了可用于在细胞中繁殖的质粒的数量。参与Cre / loxP机制的噬菌体P1的Cre重组酶是广泛使用的选择性标记基因去除系统之一。我们已经在pColdIV和pET28a（+）载体系统中过表达了密码子优化的cre基因，并通过固定的金属亲和层析纯化了His6-Cre重组酶。将获得的N端His6标签的Cre重组酶纯化约26倍，并促进线性化pLox2 +载体的两个loxP位点的位点特异性重组，从而允许切除重新环化的质粒和包含重组loxP位点的短片段DNA。