文献类型：期刊文献

中文题名：鸡PD-1及其配体实时荧光定量RT-PCR检测方法的建立与应用

英文题名：Establishment and application of real time RT -PCR method for detecting programmed death - 1 （ PD - 1 ） and its ligands in chickens

作者：孙国鹏[1];王爱国[1,2];李博文[1,2];岳锋[1];张艳芳[1];朱艳平[1];杨媛[2];郭东光[1];王选年[1,2]

第一作者：孙国鹏

机构：[1]新乡学院生物技术研究中心生命科学与技术系,河南新乡453003;[2]河南科技学院动物科学学院,河南新乡453003

第一机构：新乡学院生命科学技术学院

年份：2014

期号：8

起止页码：139-142

中文期刊名：黑龙江畜牧兽医

外文期刊名：Heilongjiang Animal Science And veterinary Medicine

收录：北大核心:【北大核心2011】；

基金：国家自然科学基金项目（31272539）

语种：中文

中文关键词：鸡;程序性死亡因子-1(PD—1);PD—L1;PD—L2;实时荧光定量RT—PCR

外文关键词：chicken ; programmed death - 1 （ PD - 1 ） ; pmgammcd death ligand 1 （ PD - L1 ） ; programmed death ligand 2 （ PD - L2 ） ; real - time fluorescence quantitative RT - PCR

摘要：为了建立并应用鸡相关免疫抑制性受体及其配体的检测方法，试验根据GenBank中程序性死亡因子-1（PD—1）及其配体PD—Ls（PD—L1、PD—L2）的基因序列分别设计特异性引物，建立这3种基因的SYBRGreenI实时荧光定量RT—PCR检测方法，并对鸡法氏囊病毒（IBDV）感染雏鸡7d的外周血单核细胞（PBMC）中3种基因mRNA表达水平进行检测。结果表明：建立的检测方法在1×10^1～1×10^8copies／μL模板范围内具有良好的线性关系，相关系数均大于0．990，重复性试验的组内及组间变异系数均小于3％；应用所建立的方法对临床样品进行检测，IBDV感染组PBMC中PD-1、PD—L1和PD—L2mRNA表达量与对照组相比均显著升高（P〈0．05）。说明所建立的检测方法具有良好的敏感性、特异性和重复性。
To establish and apply a method for detecting of an immunesuppression - associated receptor and its ligands in chickens, the specific primers were designed according to the gene sequences of the programmed death - 1 （ PD - I ） and its ligands （ PD - L1 and PD - L2 ） published in the Genbank, a SYBR Green I real - time fluorescence quantitative RT - PCR assay was established to detect the mRNA expression levels of these three genes in chicken peripheral blood mononuclear cells （PBMC） on the 7th day after infectious bursal disease virus （IBDV） infection. The results showed that the established method had a linear dynamic range from 1× 10^1 copies/μL to 1 ×10^8 copies/μL,and all the correlation coefficients were greater than 0. 990, and the intra - and inter - assay coefficients of variation in the repeatability tests were less than 3%. Further more, the results of clinical sample detection also demonstrated the mRNA expression levels of PD -1, PD -L1 and PD -L2 in the IBDV - infected group were significantly increased （ P 〈 0.05 ） compared with the control group, respectively. The results indicate that the method has good specificity, sensitivity and repeatability.