CAJ | 학술논문

目的构建表达H5N1亚型流感病毒M2和NA基因的多种真核表达载体.方法以我国分离的首株人H5N1亚型禽流感病毒(A/Anhui/1/2005)作为研究对象,PCR扩增全长M2与NA基因.将M2基因克隆入真核表达载体pStar的IRES上游或下游的MCS,构建重组pStar-M2/和pStar-/M2.将NA基因克隆人pStar载体IRES上游或下游的MCS,构建重组pStar-NA/和pStar-/NA.将M2和NA基因先后克隆到pStar载体IRES序列的上游和下游MCS,分别构建双基因共表达重组pStar-M2/NA和pStar-NA/M2.将重组质粒转染293细胞,使用IFA方法检测各重组质粒相应外源基因的表达.结果通过酶切鉴定,各重组质粒构建正确.将其转染293细胞后,使用IFA方法检测到了各重组质粒中相应外源基因的表达.结论构建了多种表达H5NI亚型流感病毒M2和(或)NA基因的真核表达载体,为研究开发流感DNA疫苗奠定了基础.
목적 구건표체H5N1아형류감병독M2화NA기인적다충진핵표체재체.방법 이아국분리적수주인H5N1아형금류감병독(A/Anhui/1/2005)작위연구대상,PCR확증전장M2여NA기인.장M2기인극륭입진핵표체재체pStar적IRES상유혹하유적MCS,구건중조pStar-M2/화pStar-/M2.장NA기인극륭인pStar재체IRES상유혹하유적MCS,구건중조pStar-NA/화pStar-/NA.장M2화NA기인선후극륭도pStar재체IRES서렬적상유화하유MCS,분별구건쌍기인공표체중조pStar-M2/NA화pStar-NA/M2.장중조질립전염293세포,사용IFA방법 검측각중조질립상응외원기인적표체.결과 통과매절감정,각중조질립구건정학.장기전염293세포후,사용IFA방법 검측도료각중조질립중상응외원기인적표체.결론 구건료다충표체H5NI아형류감병독M2화(혹)NA기인적진핵표체재체,위연구개발류감DNA역묘전정료기출.
Objective To construct vectors expressing M2 and NA genes of H5N1 influenza virus. Methods Based on the human HSN1 avian influenza virus ( A/Anhui/1/2005) isolated in china, M2 and NA genes were amplified by PCR. M2 or NA gene was subcloned into pStar vector to construct recombinant pStar-M2/, pStar-/M2, pStar-NA/and pStar-NA/. Furthermore, both of the M2 and NA genes were subcloned into pStar to construct two genes co-expressing recombinant pStar-M2/NA and pStar-NA/M2. Expression of the genes were detected by IFA after transfection of 293 cells with the recombinant plasmids. Results Recombinant plasmids were constructed and identified by restriction endonuclease digestion. Expression of the genes cloned into the recombinant plasmids was confirmed by IFA. Conclusion Recombinant plasmids expressing M2 and/or NA genes of H5N1 influenza virus were constructed, which provided basis for development of influenza DNA vaccine.