抗菌肽牛乳铁多肽素（LfcinB）在毕赤酵母中的表达及活性鉴定

Expression and Identification of LfcinB Gene in Pichia pastoris

摘要:

利用巴斯德毕赤酵母(Pichia pastors)系统表达抗菌肽——牛乳铁多肽素（bovine lactoferricin，简称LfcinB），获得的分泌型表达产物具有较强的抗菌活性。首先将人工合成的LfcinB基因片段克隆到巴斯德毕赤酵母分泌型表达载体pPIC9K中，获得的重组质粒pPIC9K-LfcinB通过限制性内切酶SalⅠ酶切线性化，经电穿孔法转化入毕赤酵母细胞SMD1168内。G418抗性筛选，得到高拷贝转化子，经PCR检测LfcinB基因与毕赤酵母染色体稳定整合。阳性克隆经甲醇诱导表达LfcinB。 结果表明，抗菌肽牛乳铁多肽素基因已经整合到酵母细胞基因组中并获得表达，表达产物具有较强的杀菌作用。

Abstract:

In present study，bovine Lactoferricin was first secretly expressed in Pichia pastors yeast expression system.The synthesized LfcinB gene fragment was cloned into expression vector pPIC9K，and then obtained recombinant plasmid，designated as pPIC9K-LfcinB，was linearized and transformed into Pichia pastors strains SMD1168 by electroporation.The transformants were screened with Geneticin and multiply-copy colonies were harvested，in which LfcinB gene was verified to inserted into yeast chromosome stably.The positive recombinant Pichia strains were induced with methanol to express LfcinB in culture supernatant.Its expressive products has high activity of killing bacteria.We concluded that LfcinB gene was cloned and integrated into yeast chromosomes，and obtained expression peptide was tested to have high antibacterial activity.