沉默SCAP基因对奶牛乳腺上皮细胞脂滴的影响

doi:10.11843/j.issn.0366-6964.2019.03.005

Abstract

Abstract:

The aim of this study was to investigate the effect of silencing SCAP gene on formation of lipid droplet in dairy mammary epithelial cells. SCAP short hairpin RNA lentivirus vector was constructed, the bovine mammary epithelial cells were packed and infected. SCAP gene silencing cell lines were selected by applying puromycin in culture medium. Effects of SCAP gene silencing on the expression of SCAP gene and protein were detected by Real-time PCR and Western blot, moreover, the effects of SCAP gene silencing and transient transfection of SCAP plasmid on lipid droplet formation were detected using Nile red and Oil red O staining. The results showed that lentivirus recombinant vector with SCAP gene silencing was constructed successfully. SCAP gene silencing stable cell line was obtained via screening of 5 mg·L^-1 puromycin. The expression of SCAP gene in all silent cell lines were significantly reduced(P<0.05 or P<0.01). Compared with control group, the expression of SCAP protein in RNAi-SCAP3 cell lines were decreased by 0.49 fold (P<0.05), the expression of SCD and FAS genes were significantly decreased (P<0.05) in RNAi-SCAP3 cells. Lipid droplet content in RNAi-SCAP3 cell lines was significantly lower than that in control cells (P<0.01). The number of small lipid droplets with size ≤ 2.0 μm were increased, while the number of large lipid droplets with size ≥ 2.5 μm were decreased. After transient transfection of SCAP eukaryotic expression vector into RNAi-SCAP3 gene silencing cell lines, overexpression of SCAP significantly decreased the percentage of small lipid droplets and increased the percentage of large lipid droplets (P<0.05). In this study, SCAP gene silencing cell lines were successfully constructed, and the size of lipid droplets were affected by the expression of SCAP.

CLC Number:

S823.2

XING Zhiyang, ZHANG Menglu, ZHANG Han, WANG Yueying, HAN Liqiang, GAO Tengyun, YANG Guoyu. Effect of Silencing SCAP Gene on Lipid Droplets in Dairy Mammary Epithelial Cells[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2019, 50(3): 507-516.

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Effect of Silencing SCAP Gene on Lipid Droplets in Dairy Mammary Epithelial Cells

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