腺病毒载体转染水貂耳缘成纤维细胞的研究

doi:10.16431/j.cnki.1671-7236.2020.01.005

Abstract

Abstract: The objective of this study was to investigate the feasibility and efficiency of transfection of adenovirus vector with enhanced green fluorescent protein (EGFP) into fibroblasts in the ear margin of mink.Firstly,ear fibroblasts of mink were isolated and cultured by tissue adherent method,and then the virus Ad-EGFP was packaged and the titer was detected by TCID₅₀ method.Two groups of different concentration gradients were used to infect mink fibroblasts,multiplicity of infection(MOI):0/100/300/500/700/900 and 0/10/30/50/70/90.Transfection efficiency (%) was estimated by looking at the ratio of positive cells to the total number of cells under a microscope.The results showed that primary cells could be grown in the ear tissue of mink cultured by tissue adherent method for 11 d,and the fusion degree of passage cells could reach 80% in 3-4 d.The titer of the packaged Ad-EGFP virus was 1.99×10¹¹ PFU/mL,the optimal MOI value of the infected mink fibroblasts was 30,and the instantaneous transfection efficiency was more than 90%.In conclusion,it was feasible to transfection mink fibroblasts with adenovirus vectors,which was an efficient transfection method and provided a basis for the preliminary validation test of mink genome editing.

Key words: American mink; adenovirus transfection; fibroblast; cell culture

CLC Number:

S855.3

LI Wen, HAN Yuping, ZHAO Xiangyuan, FAN Bingfeng, LI Xiaoxia, DIAO Yunfei, YANG Yifeng, XU Baozeng. American Mink Ear Fibroblasts Transfected by Adenovirus Vector[J]. China Animal Husbandry and Veterinary Medicine, 2020, 47(1): 37-43.

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American Mink Ear Fibroblasts Transfected by Adenovirus Vector

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