Product Class: Other

WarmStart® RTx Reverse Transcriptase
NEBU cloned at NEB recombinant 80 Heat

Need assistance designing LAMP primers? Use the NEB LAMP Primer Design Tool.

Product Introduction

  • Ideal for use in Loop-Mediated Isothermal Amplification (LAMP))
  • WarmStart® feature enables high-throughput applications, room temperature reaction setup, and increases the consistency and specificity of amplification reactions
Catalog # Size Concentration
M0380S 50.0 reactions 15000 units/ml
M0380L 250.0 reactions 15000 units/ml

Product Information

Description




WarmStart RTx Reverse Transcriptase is a unique in silico designed RNA-directed DNA polymerase coupled with a reversibly-bound aptamer that inhibits RTx activity below 40°C. This enzyme can synthesize a complementary DNA strand initiating from a primer using RNA (cDNA synthesis) or single-stranded DNA as a template. RTx is a robust enzyme for RNA detection in amplification reactions and is particularly well suited for use in LAMP (loop-mediated isothermal amplification). The WarmStart property enables high throughput applications, room temperature setup, and increases the consistency and specificity of amplification reactions. RTx contains intact RNase H activity.

Improved Speed and Sensitivity in RT-LAMPImproved Speed and Sensitivity in RT-LAMP

RT-LAMP reactions with Bst 2.0 WarmStart DNA Polymerase and the indicated reverse transcriptase were incubated at 65°C with 1 pg - 100 ng of Jurkat total RNA. Reactions were monitored with real-time fluorescence, and the resulting curves are shown on left, with the corresponding threshold times on right. WarmStart RTx provides faster reaction threshold times for improved consistancy and sensitivity with lower input RNA amounts. RT-LAMP reactions performed with AMV Reverse Transcriptase resulted in inconsistent detection, as indicated by wide variation at lower RNA input concentrations (blue curves).
Sensitive Detection with Reduced False Negatives
Total RNA WarmStart RTx AMV RT
100 ng 3/3 3/3
10 ng 3/3 3/3
1 ng 3/3 3/3
100 pg 3/3 2/3
10 pg 3/3 2/3
1pg 3/3 1/3

RT-LAMP reactions were performed with WarmStart RTx and AMV Reverse Transcriptase with total RNA at 6 input concentrations. Positive detection, indicated by fluorescent detection with a twenty-minute reaction time, was achieved down to 1 pg of total input RNA. Use of AMV resulted in false negatives when total RNA was 100 pg or lower.
WarmStart Control of WarmStart RTxWarmStart Control of WarmStart RTx

cDNA synthesis was performed for 10 minutes followed by qPCR analysis. Resulting Cts were normalized against both a "no RT" control for 0% activity and the fastest Ct for 100% activity. WarmStart RTx is inhibited by a reversibly-bound aptamer at temperatures below 40°C.
Robust RT-LAMP Across a Range of TargetsRobust RT-LAMP Across a Range of Targets

A variety of RT-LAMP targets were amplified using Bst 2.0 WarmStart DNA Polymerase and the indicated reverse transcriptase. Reactions with WarmStart RTx resulted in faster threshold times and more consistent reaction success across a range of targets and copy number. 
This product is related to the following categories:
RT-PCR Products,
Reverse Transcriptases & RT-PCR Products,
cDNA Synthesis & Reverse Transcriptases Products,
This product can be used in the following applications:
Strand Displacement Amplification & Nicking Enzyme,
cDNA Synthesis,
Reverse Transcription (cDNA Synthesis),
Loop-Mediated Isothermal Amplification,
RT-PCR & cDNA Synthesis,
RT-qPCR, RT-PCR and cDNA Synthesis,
Isothermal Amplification,
PCR

Reagents Supplied

Reagents Supplied

The following reagents are supplied with this product:

NEB # Component Name Component # Stored at (°C) Amount Concentration

Properties & Usage

Unit Definition

One unit is defined as the amount of enzyme that will incorporate 1 nmol of dTTP into acid-insoluble material in a total reaction volume of 50 μl in 20 minutes at 50°C using poly(rA)•oligo(dT)18 as template.

Reaction Conditions

1X Isothermal Amplification Buffer Pack

1X Isothermal Amplification Buffer Pack
20 mM Tris-HCl
10 mM (NH4)2SO4
50 mM KCl
2 mM MgSO4
0.1% Tween® 20
(pH 8.8 @ 25°C)

Storage Buffer

100 mM KCl
10 mM Tris-HCl
0.1 mM EDTA
1 mM DTT
50% Glycerol
pH 7.4 @ 25°C

Heat Inactivation

80°C for 10 minutes

Unit Assay Conditions

20 mM Tris-HCl (pH 8.8), 10 mM (NH4)2SO4, 50 mM KCl, 2 mM MgSO4, 0.1% Tween-20, 0.5 mM dTTP, 0.4 mM poly(rA)•oligo(dT)18.

Application Features

  • RT-LAMP
  • cDNA synthesis
  • RT reactions requiring room temperature setup

Product Notes

  1. Reaction Conditions: 1X Isothermal Amplification Buffer, template, primer, dNTPs and 0.25–0.5 µl of WarmStart RTx Reverse Transcriptase in a reaction volume of 25 µl. Incubate at 50–55°C for cDNA synthesis or directly at 65°C for One-step RT-LAMP.

Protocols, Manuals & Usage

Protocols

  1. Typical RT-LAMP Protocol
  2. Typical cDNA Synthesis Protocol

Tools & Resources

Web Tools

FAQs & Troubleshooting

FAQs

  1. What is LAMP and RT-LAMP?
  2. What advantages does WarmStart® RTx provide?
  3. How sensitive is RNA amplification with WarmStart® RTx?
  4. Can WarmStart® RTx be inactivated?
  5. Is WarmStart® RTx active in other buffers?
  6. How do I use WarmStart® RTx in RT-LAMP?
  7. What is the maximum length of cDNA product produced by WarmStart® RTx?
  8. How do I use WarmStart® RTx for cDNA synthesis?
  9. Does WarmStart® RTx have RNase H activity?
  10. What dNTPs do you recommend for use with WarmStart® RTx?
  11. Can DNA be used as a template for WarmStart® RTx?
  12. Does NEB have a master mix for LAMP or RT-LAMP reactions?
  13. What does WarmStart® mean?

Quality, Safety & Legal

Quality Assurance Statement

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 DataSheets

The following is a list of Safety Data Sheet (SDS) that apply to this product to help you use it safely.

Legal and Disclaimers

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.

Licenses

Nucleic acid-based aptamers for use with 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 WarmStart® RTx Reverse Transcriptase 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