SuperRT cDNA Synthesis Kit

Cat. No.: S665657

Storage

Store at -20°C,Avoid repeated freezing and thawing

Shipped In

Ice chest + Ice pads

Visão geral

This product is a cDNA first strand synthesis kit specially prepared for the first step experiment of two-step RT-PCR. The reverse transcriptase used in this kit is a novel and efficient reverse transcriptase that utilizes E. coli engineered bacteria for recombination and expression. It removes RNase H activity and enhances its thermal stability. It can synthesize cDNA first strands using extremely low amounts of total RNA or mRNA, with an initial sample size as low as pg level. SuperRT reverse transcriptase has strong affinity for RNA and can read RNA templates with high GC content and complex secondary structures, obtaining high yields of cDNA. This product contains all the reagents required for reverse transcription from RNA templates to cDNA first strand, including Super RT efficient reverse transcriptase, reaction buffer, primers, dNTP, etc. It is simple and convenient to use. This system has high compatibility with subsequent PCR and quantitative PCR experiments, and is suitable for various DNA polymerase reactions.

S665657	Component	100 T	Storage
S665657A	SuperRT, 200 U/μL	100 μL	-20℃. Avoid freeze/thaw cycle.
S665657B	5×SuperRT Buffer	500 μL	-20℃. Avoid freeze/thaw cycle.
S665657C	Primer Mix	240 μL	-20℃. Avoid freeze/thaw cycle.
S665657D	dNTP Mix, 2.5 mM Each	500 μL	-20℃. Avoid freeze/thaw cycle.
S665657E	RNase-Free Water	1 mL	-20℃. Avoid freeze/thaw cycle.

Product features:
·Efficient reverse transcription: It has a high affinity for RNA templates, with a reverse transcription efficiency of up to 90%, and can recognize pg level templates.
·Free response to complex templates: Even templates with high GC content and complex secondary structures can achieve good results without high-temperature denaturation.
Notes:
1. During the operation process, RNase contamination should be avoided to prevent RNA degradation or cross contamination during experiments. It is recommended to perform RNA operations in specialized areas, use specialized instruments and consumables, and have operators wear masks and disposable gloves, and frequently change gloves.
2. Disposable plastic containers should be used as much as possible for experiments. If glass containers are used, they should be treated with a 0.1% DEPC (diethyl pyrocarbonate) aqueous solution at 37 ℃ for 12 hours, and sterilized under high pressure at 120 ℃ for 30 minutes before use. Alternatively, glass containers should be sterilized under dry heat at 180 ℃ for 60 minutes before use. The sterile water used in the experiment should be treated with 0.1% DEPC and then subjected to high-pressure sterilization.
3. All reagents in this reagent kit should be gently mixed upside down before use, avoiding foaming as much as possible, and used after brief centrifugation. The enzymes involved should be returned to -20 ℃ as soon as possible after use to avoid repeated freeze-thaw cycles.
If the initial amount of RNA is less than 50 ng, it is recommended to add RNA enzyme inhibitors (RNAsin). This kit is not provided.

Usage:
Note: 1 ng -5 µ g of total RNA can establish a 20 µ l reaction system. If the total RNA amount is greater than 5 µ g, please expand the reaction system proportionally.
Steps for reverse transcription:
1. Dissolve the RNA template, Primer Mix, dNTP Mix, SuperRT Buffer, SuperRT, and RNase Free Water and place them on ice for later use.
2. Prepare a reaction system according to the following table, with a total volume of 20 μ L.

Reagent	20 μlReaction system	Final concentration
dNTP Mix，2.5 mM Each	4 μl	500 μM Each
Primer Mix	2 μl	/
RNA Template	X μl	50 pg-5 µg
SuperRT，200 U/μl	1 μl	/
RNase-Free Water	up to 20 μl	/

Attention:
1) If the initial amount of RNA is less than 50 ng, it is recommended to add RNA enzyme inhibitors (RNAsin). This kit is not provided.
2) Primer Mix is formulated from Oligo (dT) and Random Primer. Oligo dT Primer or Gene Specific Primer can be used according to experimental needs, with a recommendation of 20 μ The reaction system Oligo dT Primer is 50 pmol, or Gene Specific Primer is 2 pmol.
3. Vortex shake and mix well, briefly centrifuge to collect the solution on the pipe wall to the bottom of the pipe.
Incubate at 4.42 ℃ for 30-50 minutes and 85 ℃ for 5 minutes. After the reaction is complete, centrifuge briefly and cool on ice.
5. Reverse transcripts can be directly used for PCR reactions and fluorescence quantitative PCR reactions, or stored at -20 ℃ for a long time. Reagent 20 μ Final concentration of reaction system dNTP Mix, 2.5 mM Each 4 μ L 500 μ M Each Primer Mix 2 μ RNA Template X μ L 50 pg-5 µ g 5 x SuperRT Buffer 4 μ 1 x SuperRT, 200 U/ μ L 1 μ RNase Free Water up to 20 μ L
ii If the reverse transcription efficiency is low, or the RNA template secondary structure is complex and the GC content is high, the following steps are recommended:
1. Dissolve the RNA template, Primer Mix, dNTP Mix, SuperRT Buffer, SuperRT, and RNase Free Water and place them on ice for later use.
2. Configure the reaction system according to the following table, with a total volume of 15 μ L.

Reagent	20 μlReaction system	Final concentration
dNTP Mix，2.5 mM Each	4 μl	500 μM Each
Primer Mix	2 μl	/
RNA Template	X μl	50 pg-5 µg
RNase-Free Water	up to 15 μl	/

Note: Primer Mix is formulated from Oligo (dT) and Random Primer. Oligo dT Primer or Gene Specific Primer can be used according to experimental needs.

3. Incubate at 70 ℃ for 10 minutes and quickly ice bath for 2 minutes.
4. Centrifuge briefly to collect the solution on the tube wall to the bottom of the tube.
5. Continue to add the following reagents to the above reaction solution:

Reagent	20 μlReaction system	Final concentration
5×SuperRT Buffer	4 μl	1×
SuperRT，200 U/μl	1 μl	/

Note: If the initial amount of RNA is less than 50 ng, it is recommended to add RNA enzyme inhibitors (RNasins). This kit is not provided.

6. Incubate at 42 ℃ for 30-50 minutes and 85 ℃ for 5 minutes.
7. After the reaction is complete, centrifuge briefly and cool on ice.
8. Reverse transcripts can be directly used for PCR reactions and fluorescence quantitative PCR reactions, or stored at -20 ℃ for a long time.

Especificações

Condições de armazenamento de armazenamento

Store at -20°C,Avoid repeated freezing and thawing

Enviado em

Ice chest + Ice pads

Este produto requer transporte de cadeia fria. Serviços terrestres e outros serviços econômicos não estão disponíveis.

Product Line

Produtos biológicos

Perguntas frequentes

How should this product be stored?

Store at ?20 °C. Avoid repeated freeze–thaw cycles. Aliquot into single-use volumes before freezing.

How is this product shipped?

This product ships in an insulated container with ice pads. Unpack on arrival and transfer it to the storage condition stated above.

What documentation is provided?

Available product documentation, including Certificates of Analysis (COA), Safety Data Sheets (SDS), and specification sheets, is shown in the product document area. Document availability and access follow the current site policy.