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RPL 36 A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RPL 36 A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RPL 36 A基因后,PK15细胞中PCNA、Ki 67、CDK 4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RPL 36 A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),Ki 67、CDK 4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RPL 36 A基因后,细胞增殖关键基因PCNA、Ki 67、Cyclin B、CDK 4的表达量及48 h的细胞数量和阳性细胞数均有显著变化,RPL 36 A基因可影响PK15细胞的增殖过程。
【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 RPL 36 A gene were constructed and transfected into PK15 cells by liposome transfection method.The expression efficiency of RPL 36 A 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 RPL 36 A gene in PK15 cells,the expression of PCNA,Ki 67 and CDK 4 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 RPL 36 A gene in PK15 cells,the expression of PCNA and Cyclin B genes mRNA were extremely significant decreased(P<0.01),the expression of Ki 67 and CDK 4 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 RPL 36 A gene in PK15 cells,the expression of PCNA,Ki 67,Cyclin B and CDK 4 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 af
作者
王首元
贠红梅
史明月
秦云梦
李熊
陈军舟
周琛帛
曹果清
WANG Shouyuan;YUN Hongmei;SHI Mingyue;QIN Yunmeng;LI Xiong;CHEN Junzhou;ZHOU Chenbo;CAO Guoqing(College of Animal Science,Shanxi Agricultural University,Taigu 030801,China;Shanxi Animal Husbandry Technology Extansion Service Center,Taiyuan 030001,China)
出处
《中国畜牧兽医》
CAS
CSCD
北大核心
2023年第8期3035-3044,共10页
China Animal Husbandry & Veterinary Medicine
基金
2021年国家级大学生创新创业训练计划项目(202110113003)
三晋学者支持计划专项经费(2016、2017)
山西省高等学校科学研究优秀成果培育项目
山西农业大学生物育种工程项目(YZGC128)
2022年度山西农业大学“特”“优”农业高质量发展科技支撑工程(TYGC-07)。
