猪RPL36A基因对PK15细胞增殖过程的影响

doi:10.16431/j.cnki.1671-7236.2023.08.001

摘要/Abstract

摘要： 【目的】探究核糖体蛋白L36A(ribosomal protein L36A,RPL36A)基因对PK15细胞增殖过程的影响,为解析马身猪和大白猪生长速度差异的生理机制奠定基础。【方法】采用脂质体法将RPL36A基因干扰和过表达载体转染至PK15细胞中,通过实时荧光定量PCR和Western blotting技术检测RPL36A基因表达效率及细胞增殖标志基因(PCNA、Ki67、Cyclin B、CDK4)的表达变化,并通过划痕试验、CCK-8和EdU法检测细胞增殖情况。【结果】过表达RPL36A基因后,PK15细胞中PCNA、Ki67、CDK4基因mRNA表达量均极显著升高(P＜0.01),Cyclin B基因mRNA表达量显著升高(P＜0.05);PCNA蛋白表达量显著升高(P＜0.05);PK15细胞在48 h的细胞数量极显著高于空载组(P＜0.01),细胞增殖速度升高;阳性细胞数极显著升高(P＜0.01)。干扰RPL36A基因后,PK15细胞中PCNA、Cyclin B基因mRNA表达量均极显著降低(P＜0.01),Ki67、CDK4基因mRNA表达量均显著降低(P＜0.05);PCNA蛋白表达量显著降低(P＜0.05);PK15细胞在48 h的细胞数量显著低于对照组(P＜0.05),细胞增殖速度降低;阳性细胞数极显著降低(P＜0.01)。【结论】在PK15细胞中过表达和干扰RPL36A基因后,细胞增殖关键基因PCNA、Ki67、Cyclin B、CDK4的表达量及48 h的细胞数量和阳性细胞数均有显著变化,RPL36A基因可影响PK15细胞的增殖过程。

关键词: 猪; RPL36A基因; PK15细胞; 过表达; 干扰; 细胞增殖

Abstract: 【Objective】 The purpose of this study was to investigate the effect of ribosomal protein L36A (RPL36A) gene on PK15 cells proliferation,and lay the foundation for the analysis of the physiological mechanism of the difference in growth rate between Mashen and Large White pigs.【Method】 siRNA and overexpression vectors of RPL36A gene were constructed and transfected into PK15 cells by liposome transfection method.The expression efficiency of RPL36A gene and the expression of genes (PCNA,Ki67,Cyclin B and CDK4) related cell proliferation in PK15 cells were detected by Real-time quantitative PCR and Western blotting.The scratch tests,CCK-8 and EdU methods were used to detect cell proliferation.【Result】 After overexpression of RPL36A gene in PK15 cells,the expression of PCNA,Ki67 and CDK4 genes mRNA were extremely significant increased (P＜0.01),the expression of Cyclin B gene mRNA was significantly increased (P＜0.05),and the expression of PCNA protein was significantly increased (P＜0.05).The number of PK15 cells at 48 h was extremely significantly higher than control group (P＜0.01),and the rate of proliferation was increased.The number of positive cells of PK15 cells were extremely significantly increased (P＜0.01).After interference of RPL36A gene in PK15 cells,the expression of PCNA and Cyclin B genes mRNA were extremely significant decreased (P＜0.01),the expression of Ki67 and CDK4 genes mRNA were significantly decreased (P＜0.05),and the expression of PCNA protein was significantly decreased (P＜0.05).The number of PK15 cells at 48 h was significantly lower than siNC group (P＜0.05),and the rate of proliferation was decreased.The number of positive cells was extremely significantly decreased (P＜0.01).【Conclusion】 After overexpression and interference of RPL36A gene in PK15 cells,the expression of PCNA,Ki67,Cyclin B and CDK4 genes,which were key genes for cell proliferation,as well as the number of PK15 cells and the number of positive cells,were significantly altered.The proliferation of PK15 cells was affected by RPL36A gene.

Key words: pigs; RPL36A gene; PK15 cells; overexpression; interference; cell proliferation

中图分类号:

S828

王首元, 贠红梅, 史明月, 秦云梦, 李熊, 陈军舟, 周琛帛, 曹果清. 猪RPL36A基因对PK15细胞增殖过程的影响[J]. 中国畜牧兽医, 2023, 50(8): 3035-3044.

WANG Shouyuan, YUN Hongmei, SHI Mingyue, QIN Yunmeng, LI Xiong, CHEN Junzhou, ZHOU Chenbo, CAO Guoqing. Effect of Porcine RPL36A Gene on PK15 Cell Proliferation[J]. China Animal Husbandry and Veterinary Medicine, 2023, 50(8): 3035-3044.

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