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RT-PCR

Power cDNA Synthesis Kit 

Cat.No Capacity Inquire
25011 30 rxn. Inquire
PRODUCT INFORMATION
Description

Manual type product for convenient and easy first cDNA synthesis

  • • Simple step
  • • Rapid reaction time : within 1 hour
  • • Ready-to-use : all components are included for synthesis
  • • Efficacy : optimal experiment condition for cDNA synthesis
  • • High purity : no contamination by RNase and DNase

Power cDNA Synthesis Kit is used to synthesize first-strand cDNA from total RNA or mRNA template. It contains all components necessary for cDNA synthesis such as AMV reverse transcriptase and RNase inhibitor etc. The central dogma (expression from DNA to protein) is very important for the understanding of life phenomena. In the past, when these expression studies were carried out on proteins. However in recent years, research on mRNAs, transcription factors for protein expression, has became ore active. This is because analysis of expression genes is more accurate and easier than protein analysis.

Therefore RNA research is preferred along with advantages of cost saving such as reagents and time. What made it possible is reverse transcriptase from retroviruses, and finally the RNA-dependent DNA polymerase can make complementary DNA to RNA. These reverse transcriptase enzymes have ribonuclease H activity, which only degrades RNA in the RNA-cDNA hybrid state after the reverse transcription termination. As a result, cDNA remains only.  

In reverse transcription reactions, the purity and amount of the RNA template determines the efficiency of the reaction. Therefore, should be careful to avoid the incorporation of RNase into RNA samples, and make sure not to be mixed by proteins, poly-anions (eg, heparin), salts, EDTA, ethanol, phenol and other solvents. The Power cDNA Synthesis Kit uses the high-efficiency AMV reverse transcriptase, it is efficient for obtaining full-length cDNA. And it is useful to use for the expression of low copy gene because it was developed with optimal first cDNA synthesis conditions. In addition, it is very easy to synthesize the best cDNA and ensures optimal results when performing PCR.

Applications
  • 01Standard RT
  • 02Real-time PCR
  • 03Microarray analysis
  • 04RNA sequencing
  • 05cDNA library
Kit Contents
Contents 25011(30 rxn)
AMV Reverse Transcriptase(10U/μl) 15 μl x 1 tube
RT Buffer(5X) 120 μl x 1 tube
Oligo [dT]15 Primer(0.2 mM) 30 μl x 1 tube
Random Primer (1 mM) 30 μl x 1 tube
RNase inhibitor(10U/μl) 30 μl x 1 tube
dNTP Mixture(10 mM; each 2.5 mM) 60 μl x 1 tube
DTT(0.1M) 60 μl x 1 tube
DNase/ RNase-free sterile water 1ml x 1 tube
Technical Data

Observation of sensitivity according to the type of primer

 

Lane M, 100 DNA Marker Solution ; lane N, no template control; lane 1~7: template RNA 3-0 ~3-6

Application for various samples


 

After extraction of total RNA from virus and cell using Viral Gene-spinTM Viral DNA/RNA Extractio Kit (Cat. No. 17151) and easy-BLUETM Total RNA Extraction Kit (Cat. No. 17061), first strand cDNA was synthesized by Power cDNA Synthesis Kit then PCR performed. In result, it was varified that RT reaction is possible with templates from 20bp to 3Kb.

Lane M1, 100bp Ladder DNA marker; lane M2, 1kb Ladder DNA Marker; lane 1, Newcastle Disease Virus HN(120 bp); lane 2, Infectious Bursal
Disease Virus VP2(450 bp); lane 3, Bfl-1(570 bp),Bcl-2 family; lane 4, Human β-Action(890 bp); lane 5, hnRNP(900 bp), alternative splicing factor; lane 6, Bcl-2(3kb), Bcl-2 family

TroubleShooting Guide
QIt is the first time to experiment with cDNA synthesis. The manual says 'DNase/RNase-free sterile water'. Can I use the third distilled water?
ABasically, DEPC treated water is used for RNA experiments to reduce RNase contamination. You can think ‘DNase/RNase-free sterile water’ in manual is DEPC treated water.
QPreMix products have specific synthesized volume, so I would like to synthesize cDNA with Manual type. Is there any product to provide all necessary components for RT reaction?
AWe recommend ‘Power cDNA Synthesis Kit’. This product is used to synthesize first-strand cDNA from total RNA or mRNA template. It contains all components necessary for cDNA synthesis such as AMV reverse transcriptase and RNase inhibitor etc.
QAMV Reverse Transcriptase is included in product content. Can I replace it with M-MLV Reverse Transcriptase?
A It can not be exchanged with M-MLV. If you want M-MLV, M-MLV is sold separately, so please purchase it.
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