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OneTaq® RT-PCR Kit

Catalog # Concentration Size List Price Quantity Your Price
E5310S 30 reactions $245.00
$220.50
Catalog # Size List Price Your Price
E5310S 30 reactions $245.00
$220.50
Catalog #
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*On-line ordering is for Canadian customers only. Web pricing is applicable only to orders placed online at www.neb.ca

Easy and efficient two-step RT-PCR

  • Optimized reaction mixes for cDNA synthesis and PCR amplification
  • Hot start formulation eliminates nonspecific amplification and does not require a separate activation step
  • OneTaq® DNA Polymerase delivers robust amplification across a wide range of templates
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    Avoiding RNase Contamination

OneTaq® RT-PCR Kit combines two powerful mixes, M-MuLV Enzyme Mix and OneTaq Hot Start 2X Master Mix with Standard Buffer for 2-step RT-PCR applications. The two mixes require minimal handling during reaction setup and yet offer consistent and robust RT-PCR reactions.

The first strand cDNA synthesis is achieved by using two optimized mixes, M-MuLV Enzyme Mix and M-MuLV Reaction Mix. M-MuLV Enzyme Mix combines M-MuLV Reverse Transcriptase and murine RNase Inhibitor while M-MuLV Reaction Mix contains dNTPs and an optimized buffer. The kit also contains two optimized primers for reverse transcription and nuclease-free water. An anchored oligo-dT primer [d(T)23VN] forces the primer to anneal to the beginning of the polyA tail. The optimized Random Primer Mix provides random and consistent priming sites covering the entire RNA templates including both mRNAs and non-polyadenylated RNAs.

The amplification step features a OneTaq Hot Start DNA Polymerase in a master mix format. OneTaq Hot Start DNA Polymerase offers higher fidelity than Taq and better amplification. RT-PCR product up to 6 kb can be generated (Figure 1).

Figure 1. First Strand DNA Synthesis Figure 1. First Strand DNA Synthesis

First strand cDNA synthesis was carried out in the presence of 1X M-MuLV Enzyme Mix at 42°C using 0.5 μg of human spleen total RNA in the presence of dT23VN (lanes 1, 4 and 7) or Random Hexamer Mix (lanes 2, 5 and 8). No-RT controls were lanes 3, 6 and 9. OneTaq Hot Start 1X Master Mix was used to amplify a 1.5 kb fragment of beta-actin gene, a 0.6 kb fragment of GAPDH gene, and a 5.5 kb fragment from p532 gene in 35 cycles. The marker lane (M) contains 1 kb Plus DNA Ladder (NEB #N3200 ).
Figure 2. First Strand DNA Synthesis Figure 2. First Strand DNA Synthesis


Reagents Supplied

The following reagents are supplied with this product:

NEB # Component Name Component # Stored at (°C) Amount Concentration
  M-MuLV Enzyme Mix M0342AVIAL -20 1 x 0.06 ml 10 X
  M-MuLV Reaction Mix M0343AVIAL -20 1 x 0.4 ml 2 X
  OneTaq® Hot Start 2X Master Mix with Standard Buffer M0484SVIAL -20 1 x 1.25 ml 2 X
  Random Primer Mix S1330AVIAL -20 1 x 0.07 ml 60 µM
  Oligo d(T)23 VN S1332AVIAL -20 1 x 0.07 ml 50 µM
  Nuclease-free Water B1502AVIAL -20 1 x 1.5 ml

Properties & Usage



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References
  • Liao, J. and Gong, Z. (1997). Biotechniques. 23, 368-370.
  • Van Gilst, M.R. et al. (2005). PLoS Biology. 3, 301-312.
  • Sambrook, J. and Russel, D.W. (2001). Molecular Cloning: A Laboratory Manual. Cold Spring Harbor: Cold Spring Harbor Laboratory Press. 3rd Ed..
  • Don, R.H. et al. (1991). Nucleic Acid Research. 19, 4008.
  • Aguila et al. (2005). BMC Molecular Biology. 6, 9.
Quality Control Assay
Quality Control tests are performed on each new lot of NEB product to meet the specifications designated for it. Specifications and individual lot data from the tests that are performed for this particular product can be found and downloaded on the Product Specification Sheet, Certificate of Analysis, data card or product manual. Further information regarding NEB product quality can be found here.
Specifications
The Specification sheet is a document that includes the storage temperature, shelf life and the specifications designated for the product. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]
Certificate of Analysis
The Certificate of Analysis (COA) is a signed document that includes the storage temperature, expiration date and quality controls for an individual lot. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]_[Lot Number]
Safety Data Sheets
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Products and content are covered by one or more patents, trademarks and/or copyrights owned or controlled by New England Biolabs, Inc (NEB). The use of trademark symbols does not necessarily indicate that the name is trademarked in the country where it is being read; it indicates where the content was originally developed. The use of this product may require the buyer to obtain additional third-party intellectual property rights for certain applications. For more information, please email busdev@neb.com.

This product is intended for research purposes only. This product is not intended to be used for therapeutic or diagnostic purposes in humans or animals.

New England Biolabs (NEB) is committed to practicing ethical science – we believe it is our job as researchers to ask the important questions that when answered will help preserve our quality of life and the world that we live in. However, this research should always be done in safe and ethical manner. Learn more.Nucleic acid-based aptamers for use with thermophilic DNA polymerases and reverse transcriptases are licensed exclusively by New England Biolabs, Inc. from SomaLogic, Inc. New England Biolabs, Inc. gives the Buyer/User a non-exclusive license to use the OneTaq® RT-PCR Kit for Research Use Only (RUO). Commercial use of this product may require a license from New England Biolabs, Inc. For additional information or to inquire about commercial use, please contact busdev@neb.com. ONETAQ®, PROTOSCRIPT® and NEW ENGLAND BIOLABS® are registered trademarks of New England Biolabs, Inc.
POLYA SPIN™ is a trademark of New England Biolabs, Inc.
IGEPAL® is a registered trademark of Rhodia Operations.
iCYCLER® is a registered trademark of Bio-Rad Laboratories, Inc.
PRIMERSELECT™ is a trademark of DNASTAR Inc.
TWEEN® is a registered trademark of Uniqema Americas LLC.

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