重组犬γ干扰素的分泌表达及抗病毒活性鉴定  

Soluble Expression Of Recombinant Canine Ifn-γ And Detection Of Antiviral Activity

作　　者：姜艳平[1] 吴煜[1] 张希[1] 姜新鹏[1] 唐丽杰[1] 乔薪瑗[1] 崔文[1] 李一经[1] 

机构地区：[1]东北农业大学动物医学学院,黑龙江哈尔滨150030

出　　处：《中国兽医杂志》2015年第6期30-33,共4页Chinese Journal of Veterinary Medicine

基　　金：黑龙江省高校科技创新基金(2011TD001);"十二五"农村领域国家科技计划课题(2012AA101304-3)

摘　　要：为了研究犬γ干扰素在抗病毒方面的应用价值,依据乳酸菌密码子的偏好性和原核表达载体p AMJ399的特点,在不改变编码蛋白的基础上,将原有的犬γ干扰素序列重新优化,构建重组乳酸乳球菌p AMJ399-Ca IFNγ/PSM565,获得了分泌表达的犬γ干扰素蛋白,将培养p AMJ399-Ca IFNγ/PSM565的培养上清液进行5倍浓缩后与MDCK细胞进行作用24 h后,接种VSV病毒,通过观察病变、MTT法检测活细胞数及荧光定量PCR检测方法,均证实该蛋白能够抑制VSV的复制,具有明显的抗病毒活性。为进一步研究犬γ干扰素在临床上的抗病毒应用和作为细胞因子佐剂的研究奠定了基础。In order to study the antiviral application of IFN-γin canine(Ca IFNγ),we optimized the condon of the IFN-γsequence based on the characters of Codon preference and the prokaryotic expression vector in p AMJ399. All of these could not change the sequence of IFN-γ protein.The gene of IFN-γwas cloned into the vector of p AMJ399,and the recombinant Lactococcus lactis of p AMJ399-Ca IFNγ/PSM565 was constructed,which expressed the secretory protein of Ca IFNγ.The supernatant of the p AMJ399-Ca IFNγ/PSM565 culture was concentrated for five times,which was co-cultured with MDCK cells and VA virus for 24 hours. Cell activity was determined by CPE and MTT methods. q RT-PCR was used to test the amount of virus. All the results showed that the Ca IFNγ had remarkable antiviral effects in the form of Inhibiting of virus replication.The study laid the foundation for the application of Ca IFNγ in the clinical trials and for the research on its use as a chemokine adjuvant.

关 键 词：犬γ干扰素 重组乳酸乳球菌 分泌表达 活性检测 

分 类 号：S858.292[农业科学—临床兽医学]