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  7. Thermo Scientific™ Maxima H Minus Reverse Transcriptase (200 U/μL)

Thermo Scientific™ Maxima H Minus Reverse Transcriptase (200 U/μL)

Catalog No. FEREP0753
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Catalog No. FEREP0753 Supplier Thermo Scientific™ Supplier No. EP0753
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Description

Thermo Scientific Maxima H Minus Reverse Transcriptase (RT) was developed through in vitro evolution of M-MuLV RT. The enzyme possesses an RNA-dependent and DNA-dependent polymerase activity but lacks RNase H activity due to mutation in RNase H domain of M-MuLV RT.

Thermo Scientific Maxima H Minus Reverse Transcriptase (RT) was developed through in vitro evolution of M-MuLV RT. The enzyme possesses an RNA-dependent and DNA-dependent polymerase activity but lacks RNase H activity due to mutation in RNase H domain of M-MuLV RT. The engineered enzyme features dramatically improved thermostability, 50X higher processivity, robustness, and increased synthesis rate compared to wild type M-MuLV RT.

The eliminated RNase H activity enables the enzyme to produce very long RNA transcripts up to 20 kb. Due to its high thermostability, the enzyme maintains full activity during the entire reverse transcription reaction and generates high yields of cDNA. The reaction temperature can be increased to 65°C for efficient transcription of RNA regions with a high secondary structure or to improve specificity using gene specific primers. The extremely high processivity of Maxima H Minus enzyme results in increased resistance to common reaction inhibitors, such as guanidine, formamide, and ethanol. 

Features of Maxima H Minus Reverse Transcriptase include:
• Thermostable—90% active after incubation at 50°C for 60 minutes in a reaction mixture
• Active up to 65°C
• High yields of full-length cDNA up to 20 kb
• High sensitivity—reproducible cDNA synthesis from a wide range of starting total RNA amounts (1 pg to 5 μg)
• Efficient—complete cDNA synthesis in 15 to 30 minutes
• Increased resistance to common reaction inhibitors
• Incorporates modified nucleotides

Applications
• First strand cDNA synthesis for RT-PCR and RT-qPCR
• Reverse transcription at elevated temperatures to reduce effects of secondary structure
• Synthesis of cDNA for cloning and expression
• Generation of labeled cDNA probes for microarrays
• Analysis of RNA by primer extension

Related products
• Maxima H Minus Reverse Transcriptase (Cat. No. EP0752)
• Maxima H Minus First Strand cDNA Synthesis Kit (Cat. No. K1652)
• Maxima H Minus First Strand cDNA Synthesis Kit, with dsDNase (Cat. No. K1681)
• Maxima H Minus cDNA Synthesis Master Mix (Cat. No. M1661)
• Maxima H Minus cDNA Synthesis Master Mix, with dsDNase (Cat. No. M1681)

Order Info

Orders placed Monday through Wednesday ship next day; Thursday and Friday orders ship the following Monday.

Specifications

Concentration 200 U/μL
Content And Storage

• Maxima H Minus Reverse Transcriptase, 4 x 10,000 units (200 U/μL)
• 5X RT Buffer

Store at –20°C.
Format Stand-alone Enzyme
GC-Rich PCR Performance High
Reaction Speed 15 to 30 min.
Technique Reverse Transcription
Optimal Reaction Temperature 50°C to 55°C
Quantity 4 x 10,000 Units
Reverse Transcriptase Maxima H Minus
Ribonuclease H Activity Reduced
Shipping Condition Dry Ice
Final Product Type First-Strand cDNA
Reaction Format Separate components
Reagent Type Reverse Transcription
Size (Final Product) Up to 20 kb
Starting Material RNA
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Frequently Asked Questions (FAQs)

When should I choose regular RevertAid RT or Maxima RT vs. RevertAid H-minus RT or Maxima H-minus RT?

It is generally beneficial to minimize RNase H activity when aiming to produce long transcripts for cDNA cloning. RNase H degrades RNA from RNA-DNA duplexes, which can result in truncated cDNA during reverse transcription of long mRNA. We also recommended using RNase H-minus RTs for template-independent addition of C nucleotides.

Do Thermo Scientific reverse transcriptases (RevertAid RT, RevertAid H-minus RT, Maxima RT, and Maxima H-minus RT) possess terminal deoxynucleotidyl (TdT) activity?

All Thermo Scientific reverse transcriptases possess intrinsic TdT activity although at varying degrees depending upon the reaction conditions. For addition of template-independent C nucleotides (as for SMART and RACE experiments), this specific TdT activity can be induced by Mn2+. We would recommend Maxima H- or RevertAid H- minus RTs for this purpose.
Is it preferable to use elevated temperature in the RT reaction when using Maxima reverse transcriptases?

cDNA synthesis at higher temperatures ensures successful transcription of RNA with high levels of secondary structure, reducing issues of primer access to template. Therefore, we do recommend to use RT enzymes with high thermostability, e.g. Maxima and Maxima H Minus Reverse Transcriptases, which provide higher yields of full-length cDNA, better sensitivity, and successful transcription of GC-rich templates.
Is Maxima H Minus Reverse Transcriptase (200 U/µL) (Cat. No. EP0753) available without the buffer?

No, this product is always provided with the buffer. Connect with us to explore bulk and custom formats.

What steps should I take while performing first strand cDNA synthesis using low purity template (e.g., inhibitors in RNA sample)?

Trace amounts of reagents used in RNA purification protocols may remain in solution and inhibit first-strand synthesis, e.g., SDS, EDTA, guanidine salts, phosphate, pyrophosphate, polyamines, spermidine. To remove trace contaminants, we recommend re-precipitating the RNA with ethanol and washing the pellet with 75% ethanol, or re-purifying the RNA.
For reverse transcription, how important is the quality of RNA template?

RNA purity and integrity are essential for synthesis and quantification of cDNA. Always assess the integrity of RNA prior to cDNA synthesis. Use freshly prepared RNA. Multiple freeze/thaw cycles of the RNA sample and synthesized cDNA is not recommended. Avoid RNase contamination and discard low quality RNA.
When should I choose regular RevertAid RT or Maxima RT vs. RevertAid H Minus RT or Maxima H Minus RT?

It is generally beneficial to minimize RNase H activity when aiming to produce long transcripts for cDNA cloning. RNase H degrades RNA from RNA-DNA duplexes, which can result in truncated cDNA during reverse transcription of long mRNA. We also recommended using RNase H Minus RTs for template-independent addition of C nucleotides. In contrast, reverse transcriptases with intrinsic RNase H activity are often favored in 1-step RT-qPCR applications.
What is the fidelity of RevertAid and Maxima reverse transcriptases?

The fidelity of RevertAid and Maxima reverse transcriptases is the same as that of wild-type M-MuLV RT, which is in the range of 1 error per 15,000-27,000 nucleotides synthesized.
Do Thermo Scientific reverse transcriptases (RevertAid RT, RevertAid H Minus RT, Maxima RT, and Maxima H Minus RT) possess terminal deoxynucleotidyl (TdT) activity?

All Thermo Scientific reverse transcriptases (RevertAid RT, RevertAid H Minus RT, Maxima RT, and Maxima H Minus RT) possess intrinsic TdT activity, although at varying degrees depending upon the reaction conditions. We recommend specialized SuperScript IV Template Switching RT Master Mix for high efficiency in applications requiring template switching RT.
Do your 'engineered' MMLV-derived reverse transcriptase enzymes (e.g., SuperScript IV RT and Maxima RT) retain the ability to add a few additional nucleotides to the 3' end of the newly synthesized cDNA strand?

Regarding the tailing/TdT activity, a recent comparative analysis by R&D has shown that SuperScript IV RT has a much better TdT activity than SuperScript III RT, and that the latter has lost almost all activity.

Regarding the length of fragments that can be synthesized, internal tests have shown that fragments of up to 13 kb can be synthesized using SuperScript IV RT. Larger fragments have not been tested yet. Maxima H- RT was successfully tested with fragments up to 20 kb and since it has a certain amount of TdT activity, this enzyme could also be an alternative.
For template switching, we would recommend using SuperScript IV RT in the first instance, but Maxima H- RT would also work.

For Research Use Only. Not for use in diagnostic procedures.