
Description
Product Highlights
- Efficient – high-target affinity, coupled with a novel TransAmp buffer system for improved yield of full-length cDNA
- Unbiased – optimized mix of random hexamers and anchored oligo dT primers for complete 5’ to 3’ RNA sequence representation
- Sensitive –lower Ct values from a broad range of input cDNA concentrations, enabling accurate detection of very low-copy targets
- Robust – reliable reverse transcription under challenging conditions, including complex templates and in the presence of inhibitors
- Fast – high-yield reverse transcription from a very broad of targets in as little as 5 minutes
Product Description
The SensiFAST™ cDNA Synthesis Kit provides a rapid and sensitive method for first-strand cDNA synthesis, which displays excellent linearity across a wide range of starting material. This gives the same relative representation in cDNA templates, regardless of gene abundance, making it excellent for use in qPCR studies.
SensiFAST cDNA Synthesis Kit contains a highly-pure reverse transcriptase and optimized TransAmp™ buffer system, which includes a unique blend of random hexamers and anchored oligo (dT) primers to deliver the highest quality qPCR ready cDNA. This makes the SensiFAST cDNA Synthesis Kit ideal for working with limited sample volumes, such as laser-micro dissected samples and tissue biopsies (down to 1 pg of input RNA), to reverse transcribe precious RNA into stable cDNA ready for accurate real-time quantification. The unique blend of random hexamer primers and anchored oligo dT in the TransAmp Buffer also ensure unbiased 3’ and 5’ coverage and reverse transcription of all regions.
SensiFAST cDNA Synthesis Kit can be used with SensiFAST Probe and SensiFAST SYBR® Kits for fast real-time RT-qPCR without compromising on quality, giving real-time results in less than an hour.
Applications
- Gene expression analysis
- Tissue biopsy analysis
- miRNA profiling / quantification
- RNA target detection
- Pathogen detection

SensiFAST cDNA Synthesis Kit Customer Review
SensiFAST cDNA Synthesis Kit Customer Review
Real-Time PCR Selection Chart
One-step Vs. Two-step real-time RT PCR
A discussion of the pros and cons of each detection strategy.ebiomall.com






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和RevertAid™HMinusFirstStrandcDNASynthesisKit,FirstStrandcDNASynthesisKit
大家又没有用过,这个公司的酶据说很好啊。
一、cDNA第二链的合成:
1. 第一链反应完成后,取2ul一链产物-20℃冰箱中保存,待电泳检测。其余的产物合并,混匀,然后顺序加入下列试剂(promega):
20ul 10×DNA Polymerase I buffer
6ul10mM dNTP(自己配制)
xuldd H2O
1ulRNase H(2U/ul)
10ul DNA Polymerase I(10U/ul)
总体系为200ul;
2. 混匀后,16℃反应2.5小时;
3. 70℃灭活10分钟;
4. 反应完成后,得到200ul cDNA第二链反应体系,将此体系置于冰上;
5.取2ul二链产物,同保存的一链产物一起电泳鉴定。同时上1kb ladder,确定双链的大小范围。
注:一链,二链的电泳图是smear,且二链稍比一链大一些。
二、双链cDNA末端补平:
1. 在第二链反应体系中,顺序加入下列试剂(promega):
6ul 10mM dNTP
2ul T4 DNA Polymerase(8.7U/ul)
2ul BSA(10mg/ml)
2. 稍微离心混匀反应物, 37℃反应至少30分钟,然后75℃灭活10分钟;
3. 加入等体积酚/氯仿/异戊醇,剧烈振荡后,常温下13000g离心5分钟;
4. 离心后,吸取上清于另一1.5ml eppendof管中,加入等体积氯仿,上下颠倒几次混匀后,常温下13000g离心5分钟;
5. 吸取上清至另一eppendof管,加入1/10V3M NaAc(PH5.2)和2.5V预冷的无水乙醇,混匀,-20℃放置过夜以沉淀双链cDNA;
6. 第二日,将昨日沉淀物在4℃,13000g离心60分钟以充分沉淀双链cDNA;
7.离心完毕,弃上清,加入1ml 70%乙醇洗涤沉淀,常温下13000g离心5分钟;
8.离心完毕,弃上清,干燥沉淀至无乙醇气味.
注:第3,第4步可以用PCR 纯化试剂盒代替。
PCR纯化试剂盒操作流程:
1.溶液PE使用前应加入适量体积95%-100%的乙醇,混匀。
2.向200ul二链补平产物中加入5倍体积的buffer PB,混匀。
3.加入spin column中,13000rpm离心1min。
4.加入0.75ml buffer PE,13000rpm离心1min。
5.13000rpm,再离心1min。
6.将spin column放入一新的离心管中,加入50ul buffer EB,静置10min。
7.13000rpm离心2min。
8.加入30ul buffer EB,静置10min。
9.13000rpm离心2min。
10.加入1/10体积3M的NaAc,2.5倍体积无水乙醇,混匀,-20℃沉淀过夜。
TAKARA的RR047A逆转录试剂盒,做逆转录程序设置错误本来应该37℃15min85℃5s,变成37℃15s,85℃
5s,请问再重新设置程序可以吗,85℃酶会全部失活吗?

