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Molecular Biology Reagents | Consumables
Nucleic Acid Amplification
Reverse Transcription Series
SweScript One-Step RT-PCR Kit (for Reverse transcription series)
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Categories : Biochemicals , 1. Chemical and Reagents , Buffers , Servicebio ,
Brand : Servicebio
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Product Information
Product Name | Cat. No. | Spec. |
SweScript One-Step RT-PCR Kit | G3335-50 | 50 rxns |
G3335-100 | 100 rxns |
Product Description/Introduction
The SweScript One-Step RT-PCR Kit adopts a one-step RT-PCR method, using RNA as a template and gene-specific primers to carry out reverse transcription and PCR amplification in the same system. There is no need to add reagents during the reaction, just add RNA template, primers for the target genes and water to replenish the system, which can effectively reduce the risk of contamination. This kit contains SweScript One-Step RT Mix, 2×One-Step PCR Master Mix, of which SweScript One-Step RT Mix contains SweScript Reverse Transcriptase and RNase Inhibitor. SweScript reverse transcriptase is a mutant obtained by in vitro evolutionary screening with reduced RNase H activity and strong elongation and thermal stability. he 2×One-Step PCR Master Mix includes PCR DNA polymerase, reaction buffer, dNTPs, mg2+ and RT-PCR reaction stabilizer, etc. The DNA polymerase has a calibration function, and the reaction product is flat-ended, which has been modified and optimized to greatly improve the amplification performance and specificity. Under the carefully optimized reaction system and conditions, the amplification product of one step RT-PCR of this kit is up to 8 Kb.
Storage and Shipping Conditions
Ship with wet ice; store at -20, valid for 12 months.
Product Contents
Component Number | Component | G3335-50 | G3335-100 |
G3335-1 | SweScript One-Step RT Mix | 50 uL | 100 μL |
G3335-2 | 2×One-Step PCR Master Mix | 1 mL | 2 X 1 ml |
G3335-3 | Nuclease-Free Water | 1 mL | 1 ml |
Manual | One copy | One copy | |
Assay Protocol / Procedures
1. RT-PCR reaction system preparation
Component | Volume |
SweScript One-Step RT Mix | 1 μL |
2×One-Step PCR Master Mix | 25 μL |
Gene-specific Forward Primer(10 μM) | 2 μL |
Gene-specific Reverse Primer(10 μM) | 2 μL |
Total RNA/mRNA | 0.1 ng-1μg / 10 pg-0.1 μg |
Nuclease-Free Water | Add to 50 μL |
2. Gently mix and centrifuge instantaneously.
3. RT-PCR reaction program settings
Temperature | Time | Number of Cycles |
50a | 15-30 min | 1 |
98 | 2 min | 1 |
98 | 15 s | 35-40 |
50-65b | 20 s | |
72 | 10 s/kb | |
72 | 5 min | 1 |
a: For high GC or complex templates, the reverse transcription temperature can be raised to 55.
b: The annealing temperature is adjusted according to the TM value of the primer. Generally, the annealing temperature is lower than 3-5 of the TM value.
Note
1. Please wear disposable gloves during operation to avoid RNase contamination.
2. Be sure to combine swescript one-step RT mix and 2× One step PCR master mix is placed on ice.
3. The Swescript one step RT mix contains a high concentration of glycerol, which should be centrifuged briefly before use.
4. When using this kit for reverse transcription reaction, specific reverse transcription primers must be used, and Random Primer and Oligo dt Primer cannot be used.
5. The amount of RNA used to amplify different target genes is different, and the amount of template needs to be adjusted according to the actual amplification situation.
6. It is suggested to use the RNA extracted by the kit as a template to amplify the target gene of more than 5 kb.
For Research Use Only!
