荧光实时定量PCR检测老年人外周血淋巴细胞β1和β2肾上腺素能受体基因mRNA表达临床医学.docVIP

荧光实时定量PCR检测老年人外周血淋巴细胞β1和β2肾上腺素能受体基因mRNA表达临床医学 目录 TOC \o 1-9 \h \z \u 目录 1 正文 1 文1：荧光实时定量PCR检测老年人外周血淋巴细胞β1和β2肾上腺素能受体基因mRNA表达临床医学 1 1材料与方法 2 文2：实时定量RTPCR检测急性早幼粒细胞白血病PMLRARa融合基因医学 7 参考文摘引言： 16 原创性声明（模板） 18 文章致谢（模板） 18 正文 荧光实时定量PCR检测老年人外周血淋巴细胞β1和β2肾上腺素能受体基因mRNA表达临床医学 文1：荧光实时定量PCR检测老年人外周血淋巴细胞β1和β2肾上腺素能受体基因mRNA表达临床医学 Abstract： ObjectiveTo establish a realtime quantatitive polymerase chain reaction (PCR) method for the detection of β1and β2AR mRNA expression in elderly patients’peripheral lymphocytes. MethodsThirty elderly patients with ASA gradeⅠ-Ⅱwere recruited. Their blood was collected and the lymphoctes were isolated. The expressio of β1and β2AR mRNA were detected with fluorescent quantitative realtime PCR and semiquantitive were expressio of β1 and β2AR mRNA in all the subjects with realtime PCR, and the latter was significally more abundant than the former (P). Also, there was no difference between different gende (P). With semiquantive PCR, the ratio of β2AR mRNA/β1AR mRNA decreased when the cycles increased (P).Conclusioemiquantatitive PCR is not perfect at the platform stage, but fluorescent realtime quantitative PCR is more accurate and seitive than semiquantitive PCR for the detection of β1 and β2AR mRNA expressio in elderly patients’peripheral lymphocytes. Key words： realtime polymerase chain reaction (PCR); βadrenergic receptor; lymphocyte; elderly patient 人外周血淋巴细胞与心肌细胞表面βAR密度成显著正相关［1］。以淋巴细胞为模型，间接反应各种病理生理状态下心肌细胞βAR表达变化，被多数研究所证实。然而，淋巴细胞βAR亚型分布存在较大争议：早期研究认为，淋巴细胞仅表达β2AR［2］。最新研究发现，人外周血淋巴细胞存在β1、β2和β3AR三种亚型，且β1和β2AR mRNA丰度相近［3］。既往检测淋巴细胞βAR基因表达多采用半定量PCR，该方法属终点检测法，比较目的基因起始拷贝量准确度较低。本研究采用荧光实时定量PCR，检测老年人外周血淋巴细胞β1和β2AR基因mRNA表达，并与半定量PCR结果比较。 1材料与方法 试验对象 随机选择拟全麻下行择期手术的老年患者30例，其中男、女各15例，年龄65~80岁（平均岁），ASAⅠ~Ⅱ级，既往无心血管系统及血液系统疾病。 主要试剂及仪器 人淋巴细胞分离液（美国Sigma公司）、 TrizolTMRNA Isolation Reagent(美国Invitrogen公司)，、Revee Tracription System (美国Promega公司)， SYbr Green (日本Takara公司)。BIORAD iCycler iQ Muhicolor Realtime PCR Detection System(