uMRT First-Strand cDNA Synthesis Kit

The UltraMarathonRT® (uMRT) First-Strand cDNA Synthesis Kit contains everything required for efficient synthesis of first-strand cDNA from mRNA or total RNA templates. The kit is based on the group II intron-encoded reverse transcriptase, UltraMarathonRT (uMRT), which copies RNA end-to-end in a single pass. uMRT has powerful strand displacement activity, ultra-processivity, and natural helicase-like activity, enabling it to make full-length cDNA from RNA templates up to 30 kb.
The kit includes both oligo(dT)18 and random hexamer primers. Gene-specific primers may also be used with the kit to prime synthesis from a specified sequence. First-strand cDNA synthesized with this system can be directly used as a template in PCR or real-time PCR. 
A key advantage of this kit is that it eliminates a pervasive RT-qPCR artifact across RNA-targeting CRISPR systems. Most reverse transcription kits use MMLV retroviral reverse transcriptase to copy RNA into cDNA, but these enzymes lack strong strand displacement activity, so cDNA synthesis can be directly blocked by the tightly bound guide RNA (gRNA) itself. With bound gRNA preventing MMLV RTs from reading through the guide-binding region, RT-qPCR amplicons spanning or sitting just upstream of the gRNA site can be suppressed as a result, artificially indicating knockdown of the target gene. This pervasive effect is observed across RNA-targeting CRISPR systems, including Csm, PspCas13b, and CasRx.

Key Features and Advantages

• Accurate CRISPR KO/KD quantitation: UltraMarathonRT's intrinsic helicase reads straight through bound guide RNA, giving you accurate knockdown quantification regardless of system employed or amplicon location.
• Flexible Primer Design: Produces full-length, end-to-end cDNA in a single pass, so downstream qPCR primer pairs can be chosen anywhere along the target transcript without loss of signal — unlike non-processive retroviral RTs, which show widely different C_t values depending on primer position.
• Robust Performance: Exceptional sensitivity and efficiency, ideal for detecting low-abundance RNA, viral RNA, non-coding RNAs, long RNAs, and even highly structured RNAs.
• High Inhibitor Tolerance: Maintains full activity in the presence of inhibitory compounds potentially found in biological sample preparation, including ethanol (resistant to 10%), isopropanol (resistant to 5%), TRIzol (resistant to 0.7%), formalin (resistant to 0.0025%), ammonium acetate (resistant to 50 mM), bile salts (resistant to 0.05%), and hematin (resistant to 10 µM).
• Broad Dynamic Range: Reverse transcribes RNA input from 1 pg to 2 µg, providing flexibility in experimental design.
• Fast Reaction Time: Complete first-strand cDNA synthesis in as little as ~17 minutes.
• Preserves RNA Template Integrity: Intrinsic helicase activity unwinds RNA structure at room temperature, making the high-temperature reaction conditions required by other RTs unnecessary.
• Compatible with both dye- and probe-based qPCR systems.
  

Key Applications

• Evaluation of RNA-targeting CRISPR knockdown (Csm, PspCas13b, CasRx) — eliminates false-positive signal from guide-RNA-blocked RT
• Verification of RNAi and ASO knockdown/knockout
• Also supports RNA-Seq verification, alternative splice variant quantification, RNA degradation measurement, RNA editing quantification, and mutation detection/allele discrimination
  

Kit Components

• UltraMarathonRT
• 2x RT Buffer Q
• Nuclease-free Water
• dNTP Mix (10 mM each)
• Random Hexamers (200 µM)
• Oligo(dT)18 Primer (50 µM)

Catalog#:  R1009S (20 rxns), R1009M (50 rxns), R1009L (100 rxns)

For the product brochure, click here; for purchase orders, please contact us at sales@rnaconnect.com.

Reagents Required But Not Supplied

• RNA template: 1 pg – 2 µg total cellular RNA or 1 pg – 500 ng of mRNA are recommended for each reaction.
• Gene-specific RT primers (optional; UltraMarathonRT's full-length coverage works equally well with oligo-dT, random, or gene-specific priming).
• qPCR reagents for downstream quantification, such as RNAConnect's 2X qPCR Green Master Mix (Cat. Q1009).
  

Storage Conditions
For short duration of storage (< 3 months), UltraMarathonRT can be stored in a -20°C freezer. -80°C is recommended for long-term storage. All other kit components are stable in a -20°C freezer for long-term storage. UltraMarathonRT can endure 10 freeze-thaw cycles without losing noticeable activity; after 20 freeze-thaw cycles, it maintains 90% of its original activity.