RT SuperMix for qPCR (One-Step gDNA Remover) K7536-50
Remove gDNA, all reagent one tube, reduce configuration, reduce pollution
Item number：K7536-50
Product Description/Introduction
First-Strand cDNA Synthesis SuperMix for qPCR (One-Step gDNA Remover) contains 5×ready-to-use
master mix (5×SuperMix for qPCR) developed specifically for first-strand cDNA synthesis and gDNA remover reagents,
which removes residual genomic residues from the template while the reverse transcription reaction is performed.
The kit contains 5×No-RT Control SuperMix as a reverse-transcriptase-free control to determine whether the qPCR template is derived from cDNA.
The master mix contains all reagents required for the reverse transcription reaction (Script RT, RNase Inhibitor, Random Primer, Oligo(dT)18 Primer, dNTPs and the optimized buffer system).
Script RT is a mutant reverse transcriptase obtained by in vitro evolutionary screening and is characterized by high thermal stability and sustained synthesis.
The gDNA Remover contains a thermosensitive DNase that is 30-fold more active than DNase I and acts only on dsDNA and is susceptible to inactivation under high temperature conditions.
Storage and Shipping Conditions
Ship with wet ice; Store at -20℃, valid for 12 months.
Product Contents
Component Number | Component | G75363-50 | G75363-100 |
G7536-1 | 5×SuperMix for qPCR | 200 μl | 400 μL |
G7536-2 | gDNA Remover | 50 μL | 100 μL |
G7536-3 | Nuclease-Free Water | 1 mL | 1 mL |
G7536-4 | 5×No-RT Control SuperMix for qPCR | 20 μL | 40 μL |
Manual | One copy | One copy | |
Assay Protocol / Procedures
1. First-strand cDNA synthesis and gDNA removal
1. Reaction system preparation (20 μL reaction system recommended):
Component | Volume |
5×SuperMix for qPCR | |
gDNA Remover | 1 μL |
Total RNA/mRNA | 0.1 ng-5ug / 10 pg-0.5ug |
Nuclease-Free Water | Add to 20 μL |
2. Gently mix and centrifuge;
3. Perform reverse transcription using the recommened thermal conditions below:
Temperature | Time |
25℃ | 5 min |
42℃ | 15-30 min |
85℃ | 5 sec |
Note
1. Please wear disposable gloves when handling to avoid RNase contamination.
2. Be sure to place 5×SuperMix, 5×No-RT Control SuperMix, gDNA Remover on ice.
3. 5×SuperMix, 5× No-RT Control SuperMix, gDNA Remover contain high concentrations of glycerol and should be briefly centrifuged before use.
4. For high GC or complex templates, the RNA template, nuclease-free water can be premixed and held at 65°C for 5 min, then quickly cooled on ice.
Then add other reaction components.
5. If a negative control is required, just replace the equal volume of G7536-1 5×SuperMix for qPCR in the reaction system with
G7536-4 5×No-RT Control SuperMix for qPCR.
6. The reverse transcription products can be stored at -20℃ for a short time. If long-term storage is needed, it is recommended to store them at -80℃
after packing to avoid repeated freeze-thaw cycles.
7. If the subsequent qPCR primers are designed across exons, the genome removal step can be omitted.
For Research Use Only!
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