用实时荧光定量PCR选择急性呼吸窘迫综合征患者血清microRNA内参照基因  

Selection of internal reference genes serum microRNA in acute respiratory distress syndrome patients by real-time quantitative RT-PCR detection

作　　者：李建春[1] 毛璞[2] 李希[1] 潘莹[1] 吴苏龙 庞晓清[1] 许智恒 黄勇波[1] 刘晓青[1] 黎毅敏[1] 

机构地区：[1]广州医科大学附属第一医院重症医学科,广州510120 [2]广州医科大学附属第一医院医院感染管理科,广州510120

出　　处：《临床检验杂志》2015年第3期166-169,共4页Chinese Journal of Clinical Laboratory Science

基　　金：国家自然科学基金(81000005)

摘　　要：目的探讨用实时荧光定量PCR检测急性呼吸窘迫综合征(ARDS)患者血清微小RNA(microRNA,miRNA)的表达并选择最优内参照基因。方法用实时荧光定量PCR检测40例ARDS患者及20例体检健康者血清中5个miRNA候选内参照基因miR-16-5p、miR-93-5p、miR-101-3p、miR-191-5p和U6,并用ge Norm法、Norm Finder法、bestkeeper法和相对△Ct法4种算法分析内参照基因稳定性,通过对4种算法稳定值进行排序并计算几何均数的方法分析综合稳定值。结果比较5个候选内参照基因在ARDS患者和体检健康者的表达水平,差异均无统计学意义(P均>0.05)。4种算法综合分析显示,miR-16-5p以最小的稳定值1.32成为综合稳定性最高的内参照基因,其次是miR-101-3p、U6、miR-93-5p和miR-191-5p。结论实时荧光定量PCR法确定ARDS患者血清miRNA最优的内参照基因为miR-16-5p。Objective To investigate the expression of serum microRNA in acute respiratory distress syndrome（ ARDS） patients by real-time quantitative PCR,and select the optimal internal reference genes. Methods Five candidate miRNA reference genes,including miR-16-5p,miR-93-5p,miR-101-3p,miR-191-5p and U6,in serum samples from 40 ARDS patients and 20 healthy volunteers were detected by real-time quantitative PCR. Then,4 algorithms including ge Norm,Norm Finder,Bestkeeper and comparative delta Ct were used to calculate the values for the stability of internal reference genes. Last,a comprehensive value was calculated based on ranking the values of four algorithms and calculation of geometric mean. Results There was no significant difference in the expressions of 5 candidate reference genes between ARDS patients and healthy volunteers（ P〉0.05）. The comprehensive analysis of 4 algorithms showed that miR-16-5p was the most stable internal reference gene with the smallest value of 1. 32,followed by miR-101-3p,U6,miR-93-5p and miR-191-5p. Conclusion miR-16-5p is the optimal internal reference gene for serum miRNA detection of ARDS patients by real-time quantitative PCR.

关 键 词：急性呼吸窘迫综合征 实时荧光定量PCR 血清微小RNA 内参照基因 

分 类 号：R563.8[医药卫生—呼吸系统]