Effects of Different Promoters, WPRE and ITRs on Recombinant Baculovirus Expression of Newcastle Disease F Protein

doi:10.11924/j.issn.1000-6850.casb2020-0011

Abstract

Abstract:

The aims are to improve the expression ability of genetically engineered vaccines to the Newcastle disease virus F protein, and lay a foundation for developing highly efficient and genetically engineered vaccines. The expression differences of baculovirus with different promoters and expression elements to F protein in Chinese hamster ovary cell were detected by Western blot. The results of Western blot showed that the expression level of complex promoter containing CMV enhancer region and chicken actin promoter region (CBA promoter) was higher than that of Cytomegaoviyns promoter (CMV) by 24.52%-231.18%, and the expression level of wood-chuck hepatitis virus post-transcriptional regulatory element (WPRE) was increased by 2.9-8.6 times, but adeno-associated virus inverted terminal repeats (ITRs) only increased the expression level of 1.95%-9.27%. In conclusion, the adjustment ability of CBA promoter is much higher than that of CMV promoter, while group WPRE could get a positive result, but group ITRs does not change significantly.

Key words: Newcastle disease virus, baculovirus, WPRE, CBA

CLC Number:

S852.65

Cao Dechen, Ye Lei, Li Weiqing, Li Sainan, Gao Dongni, Ge Jingping. Effects of Different Promoters, WPRE and ITRs on Recombinant Baculovirus Expression of Newcastle Disease F Protein[J]. Chinese Agricultural Science Bulletin, 2020, 36(30): 91-97.

Figures/Tables 10

References 27

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连接反应体系成分	体积
10×ligation Buffer	1 μL
目的基因片段	4 μL (40 ng)
线性质粒载体	1 μL (10 ng)
T4 DNA Ligase	1 μL
ddH₂O	Up to 10 μL

连接反应体系成分	体积
10×ligation Buffer	1 μL
目的基因片段	4 μL (40 ng)
线性质粒载体	1 μL (10 ng)
T4 DNA Ligase	1 μL
ddH₂O	Up to 10 μL

病毒名称	滴度/(pfu/mL)
vYL-LM	2.2×10⁸
vYL-LM-I	3.0×10⁸
vYL-SD	2.8×10⁸
vYL-SD-I	3.2×10⁸
vYL-LM(-)	2.3×10⁸
vYL-SD(-)	2.7×10⁸

病毒名称	滴度/(pfu/mL)
vYL-LM	2.2×10⁸
vYL-LM-I	3.0×10⁸
vYL-SD	2.8×10⁸
vYL-SD-I	3.2×10⁸
vYL-LM(-)	2.3×10⁸
vYL-SD(-)	2.7×10⁸

组一	分子量	强度	组二	分子量	强度
泳道2 vYL-SD	57.48	1109.7	泳道5 vYL-LM	55.661	831.52
泳道3 vYL-SD-ITRs	56.563	1015.6	泳道6 vYL-LMI	55.661	815.61
泳道4 vYL-SD(-)	56.563	285.69	泳道7 vYL-LM(-)	54.772	86.264
泳道1 空白细胞对照	0	0