*** 延伸时间根据PCR 产物大小确定，一般1kb/min 。Control 引物延伸时间45s 。c 、反应结束后，取3-5 μl PCR 产物进行琼脂糖凝胶电泳，确认PCR 反应产物。如果此PCR 产物需用于以后实验，应将PCR 产物冷冻保存。RNA control 反应时，退火温度55度，30个循环，扩增片段大小为500bp 。使用提示 1. RNA 模板可...
目的:比较荧光定量PCR( real-time PCR)与普通反转录酶-聚合酶链锁反应( RT-PCR)对人偏肺病毒( hMPV)的检测价值.方法:Real-time PCR和RT-PCR同时对500例急性下呼吸道感染( ALRTI)患儿鼻咽部分泌物进行hMPV检测,分析两种方法的特异性和灵敏性.结果:Real-time PCR和RT-PCR的检出率分别为16%及9.8%,RT-PCR...
1. Total RNA was extracted using a Trizol reagent (Invitrogen). RNA (2 μg) was converted into cDNA using the RevertAid First Strand cDNA Synthesis Kit (Thermo). QRT-PCR was accomplished using the FastStart Universal SYBR Green Master (Rox) (Roche) in the ABI PRISM® 7300 real-time PCR...
RT-PCR和real-time-PCR-原理及步骤.ppt,相对定量分析方法2:双标准曲线法 目标基因与内参基因扩增效率不同 待测样品目的基因浓度 待测样品内参基因浓度 F= 对照样品目的基因浓度 对照样品内参基因浓度 样品 ●DNA纯度:OD260/OD280=1.8,1.6 ~ 2.0之间 ●DNA用量:0.05 ng
Hepatitis C virus (HCV) RNA values measured with two real-time PCR methods (Cobas Ampliprep/Cobas TaqMan, CAP/CTM, and the Abbott real-time PCR test, ART) vary among patients with genotype 1. We investigated HCV RNA values measured by two real-time PCR assays during pegylated interferon pl...
1、RT-PCR和real-time-PCR-原理及步骤聚合酶链反应(Polymerase Chain Reaction)简称,是一项在短时间内大量扩增特定的片段的分子生物学技术。多次循环后的Taq DNA聚合酶仍然具有活性,因此反应体系自动往复多次地进行对所需的DNA的片段的酶促合成,使反应产物按指数增长,所以命名为聚合酶链式反应。 10 10 扩增缓冲液扩...
综上所述,利用外标准曲线的实时荧光定量PCR是迄今为止定量最准确,重现性最好的定量方法,已得到全世界的公认,广泛用于基因表达研究、转基因研究,药物疗效考核、病原体检测等诸多领域〔10,11,12,13〕。 参考文献 1.Higuchi R, Fockler C, etal. Kinetic PCR analysis: real-time monitoring of DNA amplification ...
We introduce new real-time PCR tests for the verification of foliar nematodes and the burrowing nematodes in leaf, roots or soil samples.
1、2013-11-26实时荧光定量实时荧光定量PCR技术技术的原理及应用的原理及应用(Real Time Quantitative PCR) Contents一、实时一、实时荧光定量荧光定量PCRPCR的定义的定义二、二、RT-PCRRT-PCR技术的技术的原理及试验流程原理及试验流程三、三、RT-PCRRT-PCR技术的数据分析技术的数据分析四、四、RT-PCRRT-PCR技术...
(r=0.997)and there was a single peak with a 82.5±0 ℃ Tm in melting curve.The coeffecients of variation(CV%)was 4.26% for the intra-assay test and there was no significant difference(P0.05)in the inter-assay test.The real-time RT-PCR assay for duck IFN-γ mRNA has broad range,...