畜牧兽医学报2013，Vol.44Issue(1)：78-86,9.
神经氨酸酶基因联合抗黏液病毒基因的抗病毒作用
Antiviral Activities of Neuraminidase Gene Combining with Chicken Mx against Newcastle Disease Virus
摘要
Abstract
This experiment was conducted to build the neuraminidase (NA) gene and anti-mucus virus (Mx) dual eukaryotic gene co-expression plasmid,and detect the gene expression in trans-fected mouse fibroblasts (NIH-3T3) cells, and to investigate the influence of the recombinant plasmid on the chicken fibroblasts (CEF ) cells. The cDNA fragment of NA and mutant Mx gene were derived from pcDNA3. 0-NA, pcDNA3. 0-Mx plasmid by PCR, respectively. NA cDNA fragment and Mx cDNA fragment were inserted into the multiple cloning sites of pVITRO2 to construct the eukaryotic co-expression plasmid pVITRO2-Mx-NA. The recombinant plasmid was confirmed by restriction endonuclease treatment and sequenced, then the mouse fibroblasts (NIH-3T3) cells were transfected. The expression of genes in pVITRO2-Mx-NA were identified by RT-PCR and indirect immunofluorescence assay (IFA). Then the recombinant plasmid was transfected into CEF cells,RT-PCR and the micro-cell inhibition test were used to test the antiviral activity for NDV (Newcastle disease virus). The restriction endonuclease digestion and sequencing results suggested that co-expression vector pVITR02-Mx-NA was constructed successfully, the expression of Mx and NA could be detected in both NIH-3T3 and CEF cells. Recombinant proteins of Mx and NA protected CEF cells from NDV infection during 0-72 hours of incubation, the mutagenesis Mx or NA protein protected CEF cells from NDV infection during 0-48 hours of incubation; Compared with single-gene transfection group,co-transfection group significantly delayed the NDV infection time of CEF cells, the difference was statistically significant (P <0. 05). The recombinant eukaryotic expression vector pVITRO2-Mx-NA was constructed and expressed in CEF cell successfully, which would contribute to delaying the infection of NDV in cell level, it revealed the co-transfection of the combined genes is more powerful than single one, they had synergistic effects.关键词
NA/Mx/双基因表达载体/抗病毒活性/抗NDVKey words
NA/ Mx/ dual eukaryotic gene co-expression vector/ anti-viral activity/ anti-NDV分类
生物科学引用本文复制引用
傅德智,张亚妮,陈昊,李伟,郑蒙蒙,王丹,张振韬,施青青,李碧春..神经氨酸酶基因联合抗黏液病毒基因的抗病毒作用[J].畜牧兽医学报,2013,44(1):78-86,9.基金项目
江苏省高校自然科学重大基础研究项目资助(08KJA230001) （08KJA230001）
高等学校博士学科点专项 （）
"六大人才高峰"资助项目 （）
