犬防御素-3基因的真核表达及活性检测  

Eukaryontic Expression of Canineβ-defensin-3 and Detection of Its Antimicrobial Activity

作　　者：陈征兵 莫永正 张晓明[1] 陈瑞爱[1,2] 罗满林[1,2] 

机构地区：[1]华南农业大学兽医学院,广东广州510642 [2]广东大华农动物保健品股份有限公司,广东云浮527300

出　　处：《动物医学进展》2014年第10期85-88,共4页Progress In Veterinary Medicine

摘　　要：为了在毕赤酵母X-33中表达犬防御素-3,并检测其抗菌活性,根据毕赤酵母密码子偏好性,人工合成犬防御素-3基因,构建重组质粒PpiczɑC-CBD-3,线性化后电转至毕赤酵母菌X-33,经抗性筛选,阳性克隆用10mL/L甲醇诱导表达,表达产物经Tricine-SDS-PAGE电泳分析,并用琼脂空穴扩散法测定其抗菌活性。表达产物经Tricine-SDS-PAGE检测,在分子质量8.5ku左右有目的条带,与预期大小一致,表达的犬防御素经琼脂空穴扩散法检测显示出一定的抗菌活性。研究表明,犬防御素-3在毕赤酵母中成功分泌表达,为进一步研究奠定了基础。The rescarch aimeds at expressing the canineβ-defensin-3 gene in Pichia pastoris and determine-ing the antimicrobial activity of expressed product;According to the bias codon of Pichia pastoris ,the ca-nineβ-defensin-3 gene was synthesized,and the recombined plasmid PpiczɑC-cbd-3 was constructed ;the linearized recombined plasmid was transformed into Pichia pastors ;X-33 by electroporation,the positive clones were resistantly screened and induced by 1.0% methanol.The expressed product was identified by Tricine-SDS-PAGE,and determined for antibacterial activity by agarose diffusion methods.A protein band about 8.5 ku was detected,and consistent with the purpose bands.Agarose diffusion test showed the ca-nineβ-defensin-3 had relatively antibacterial abilities.It showed that the canineβ-defensin-3 was successful-ly expressed in Pichia pastoris ,which laid a foundation for further study on defensins.

关 键 词：犬防御素-3 毕赤酵母 抗菌活性 分泌表达 

分 类 号：S852.42[农业科学—基础兽医学]