摘要
目的建立实时荧光定量(RT-qPCR)检测Wistar大鼠黄嘌呤脱氢酶/氧化酶(XDH/XO)基因转录水平的方法,以在转录水平上对XDH/XO基因进行定量检测。方法提取Wistar大鼠肝脏组织中总RNA,经逆转录得到cDNA,以10倍为稀释因子稀释为5个浓度梯度,使用设计的引物序列和内参基因进行RT-qPCR检测,得到XDH/XO基因表达的标准曲线。进而检测Wistar大鼠高尿酸血症动物模型中XDH/XO基因转录水平的变化。结果 RT-qPCR法检测得到的XDH/XO基因标准曲线溶解峰单一,R^2接近1,能检测出高尿酸动物模型中XDH/XO基因转录水平的变化。结论 RT-qPCR检测Wistar大鼠XDH/XO基因转录水平的方法具有定量准确,重复性好的特点,可应用于高尿酸血症的发病机理、新药研究等方面。
Objective To establish Quantitative Real-time PCR(RT-qPCR) method for detecting the transcription level of xanthine dehydrogenase/oxidase(XDH/XO) gene in Wistar rats,to quantitatively detect XDH/XO gene at the transcriptional level. Method Total RNA was extracted from the liver tissue of Wistar rats. The cDNA was obtained by reverse transcription, and diluted to 5 concentration gradients with 10 dilution factors. RT-qPCR was performed using the designed primer sequences and reference genes to obtain XDH/XO gene standard curve. Furthermore, changes in the transcription level of XDH/XO gene in the animal model of hyperuricemia in Wistar rats were examined. Result The XDH/XO gene standard curve obtained by RT-qPCR method has a single dissolution peak and R^2 is close to 1. It can detect the change of XDH/XO gene transcription level in animal model of high uric acid. Conclusion RT-qPCR is a quantitative and accurate method for detecting the XDH/XO gene transcription level in Wistar rats. It can be applied to the pathogenesis of hyperuricemia and new drug research.
作者
王陈芸
李哲丽
叶尤松
蔡发晶
肖涵
谢季平
唐东红
WANG Chenyun;LI Zheli;YE Yousong;CAI Fajin;XIAO Han;XIE Jiping;TANG Donghong(Institute of Medical Biology, Chinese Academy of Medical Science/Peking Union Medical College, Kunming 650118 , China;Yunnan University of Traditional Chinese Medicine, Kunming 650200, China;Kunming institute of science and technology intelligence, Kunming 650600, China)
出处
《实验动物科学》
2019年第1期26-30,共5页
Laboratory Animal Science
基金
中国医学科学院医学与健康科技创新工程重大协同创新项目(No.2016-I2M-2-006)
