Reverse Transcription

Product description This product, based on the classic reverse transcription reagents, has optimized the reaction time. The total reaction time can be as short as less than 6 minutes, while maintaining stable detection rates, specificity, and yield. It is suitable for reverse transcription of RNA templates that are complex, present in low amounts, or represent low-copy genes. The reverse transcription product of this product can be used for downstream PCR or qPCR applications. The kit provides two types of cDNA synthesis primers: Random Primers N6 and Oligo (dT)18. Users can choose according to their needs. Specifications Catalog Number N132064E N132064S Specifications 10 T 100 T Components Catalog Number Component Name N132064E N132064S N132064-A 4×Fast cDNA Synthesis Mix (No Dye) 50 μL 500 μL N132064-B 5×gDNA Digester Mix 20 μL 200 μL N132064-C Random Primers（50 μM） 20 μL 200 μL N132064-D Oligo d(T)18 Primers (50 μM) 20 μL 200 μL N132064-E RNase-free H2O 200 μL 2×1 mL Storage Store at -25 to -15℃. Valid for 1 year. Notes 1. All operations should be carried out on ice, and RNase contamination should be avoided during the process. 2. For your safety and health, please wear a lab coat and disposable gloves when operating. 3. For Research Use Only. Instructions 1. Reverse Transcription with gDNA Removal Step 1) gDNA Digestion Prepare the following mixture in an RNase-free centrifuge tube and gently mix by pipetting. Incubate at 42°C for 2 minutes. Component Usage Amount 5×gDNA Digester Mix 2 μL Total RNA or mRNA Total RNA：10 pg-5 μg mRNA：10 pg-500 ng RNase-free H2O Up to 10 μL 【Note】It is recommended that the input amount of Total RNA does not exceed 2 µg. If the expression level of the target gene is low, the input amount can be increased up to a maximum of 5 µg. 2) Reverse Transcription Reaction Setup Component Usage Amount The reaction mixture from the previous step 10 μL 4×Fast cDNA Synthesis Mix (No Dye) 5 μL Oligo d(T)18 Primers（50 μM) or Random Primers（50 μM） 2 μL RNase-free H2O Up to 20 μL 【Notes】 a. The amount of primer can be adjusted according to the amount of template input. If the downstream experiment is qPCR, Random Primers can be added to the reaction mixture at the recommended amount. b. It is recommended to first add the 4×Fast cDNA Synthesis Mix (No Dye) and mix thoroughly before adding the reverse transcription primer to ensure that the primer is not affected by the gDNA Digester. 3) Reverse Transcription Procedure Settings Temperature Time 55℃ 5 min 85℃ 5 sec 2. Reverse Transcription without gDNA Removal Step 1) Reverse Transcription Reaction Setup Component Usage Amount 4×Fast cDNA Synthesis Mix (No Dye) 5 μL Oligo d(T)18 Primers（50 μM) or Random Primers（50 μM） 2 μL Total RNA or mRNA Total RNA：10 pg-5 μg mRNA：10 pg-500 ng RNase-free H2O Up to 20 μL 2) Reverse Transcription Procedure Settings Temperature Time 55℃ 5 min 85℃ 5 sec 【Notes】 a. It is recommended that the input amount of Total RNA does not exceed 2 µg. If the expression level of the target gene is low, the input amount can be increased up to a maximum of 5 µg. b. The amount of primer can be adjusted according to the amount of template input. If the downstream experiment is qPCR, Random Primers can be added to the reaction mixture at the recommended amount. c. For templates with high GC content or complex structures, the reverse transcription temperature can be increased to 60°C. d. This product can synthesize cDNA sequences up to 14 kb in length. If longer cDNA products are required, the reverse transcription time can be appropriately extended.

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Fast RT-gDNA Digestion OneShot SuperMix (for qPCR) Product description This product is the brand's highly recommended high-performance cDNA synthesis reagent, featuring faster reverse transcription speed, higher yield, and strong resistance to residual inhibitors. It is suitable for reverse transcription reactions of RNA from animal, plant, and microbial sources. Additionally, the included gDNA Digester Mix can digest up to 1000 ng of DNA, effectively preventing contamination from residual DNA in the RNA template and ensuring more reliable downstream results. The reverse transcription product of this product can be used for downstream qPCR applications. It is recommended to use in combination with Universal Multiplex qPCR Probe Premix (Cat#N132041), Universal qPCR Dye Premix (Cat#N132031), or Traceable qPCR Dye Premix (Cat#N132034, Cat#N132035, Cat#N132036) for high-performance gene expression analysis. Specifications Catalog Number N132063E N132063S Specifications 10 T 100 T Components Catalog Number Component Name N132063E N132063S N132063-A 4×cDNA OneShot Synthesis Mix 50 μL 500 μL N132063-B gDNA Digester Mix 10 μL 100 μL N132063-C RNase-free H2O 1 mL 2×1 mL Storage Store at -25 to -15℃. Valid for 1 year. Notes 1. All operations should be carried out on ice, and RNase contamination should be avoided during the process. 2. For your safety and health, please wear a lab coat and disposable gloves when operating. 3. For Research Use Only. Instructions 1. Reaction System Preparation Thaw Components A, B, and C on ice. Mix each component thoroughly by vortexing. Then, prepare the following reaction system in an RNase-free centrifuge tube: Component Usage Amount 4×cDNA OneShot Synthesis Mix 5 μL gDNA Digester Mix 1 μL Total RNA or mRNA Total RNA：10 pg-5 μg mRNA：10 pg-500 ng RNase-free H2O Up to 20 μL 【Note】It is recommended that the input amount of Total RNA does not exceed 2 µg. If the expression level of the target gene is low, the input amount can be increased up to a maximum of 5 µg. 2. Reverse Transcription Protocol Setup Temperature Time 37℃ 5 min 85℃ 30 sec 【Notes】 1) This process includes both reverse transcription and gDNA digestion. 2) The reverse transcription product can be directly used for downstream qPCR detection. If the downstream experiment will not be performed immediately, the product can be stored at -20°C. For long-term storage, it is recommended to aliquot and store at -80°C to avoid repeated freeze-thaw cycles.

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Product description This product is the brand's highly recommended high-performance cDNA synthesis reagent, featuring faster reverse transcription speed, higher yield, and strong resistance to residual inhibitors. It is suitable for reverse transcription reactions of RNA from animal, plant, and microbial sources. Additionally, the included gDNA Digester Mix can digest up to 1000 ng of DNA, effectively preventing contamination from residual DNA in the RNA template and ensuring more reliable downstream results. The reverse transcription product of this product can be used for downstream qPCR applications. It is recommended to use in combination with Universal Multiplex qPCR Probe Premix (Cat#N132041), Universal qPCR Dye Premix (Cat#N132031), or Traceable qPCR Dye Premix (Cat#N132034, Cat#N132035, Cat#N132036) for high-performance gene expression analysis. Specifications Catalog Number N132062E N132062S Specifications 10 T 100 T Components Catalog Number Component Name N132062E N132062S N132062-A cDNA Fast Synthesis SuperMix 60 μL 600 μL N132062-B gDNA Digester Mix 20 μL 200 μL N132062-C RNase-free H2O 200 μL 2×1 mL Storage Store at -25 to -15℃. Valid for 1 year. Notes 1. All steps for adding samples and preparing solutions should be carried out on ice whenever possible. 2. Before use, each component should be thoroughly mixed by inverting up and down, followed by a brief low-speed centrifugation to collect the contents at the bottom of the tube. 3. For your safety and health, please wear a lab coat and disposable gloves when operating. 4. For Research Use Only. Instructions 1. Reverse Transcription with gDNA Removal Step 1) DNA Digestion Prepare the following mixture in an RNase-free centrifuge tube and gently mix by pipetting. Incubate at 37°C for 2 minutes. Component Usage Amount gDNA Digester Mix 2 μL Total RNA or mRNA Total RNA：10 pg-2 μg mRNA：10 pg-500 ng RNase-free H2O Up to 14 μL 【Note】It is recommended that the input amount of Total RNA does not exceed 2 µg. If the expression level of the target gene is low, the input amount can be increased up to a maximum of 5 µg. 2) Reverse Transcription Reaction Setup (Example for a 20 µL Reaction Volume) Component Usage Amount The reaction mixture from the previous step 14 μL cDNA Fast Synthesis SuperMix 6 μL 3) Reverse Transcription Procedure Settings Temperature Time 50℃ 5 min 85℃ 5 sec 【Note】The reverse transcription product can be directly used for downstream qPCR detection. If the downstream experiment will not be performed immediately, the product can be stored at 4°C. For long-term storage, it is recommended to aliquot and store at -20°C to avoid repeated freeze-thaw cycles. 2. Reverse Transcription without gDNA Removal Step 1) Reverse Transcription Reaction Setup (Example for a 20 µL Reaction Volume) Component Usage Amount cDNA Fast Synthesis SuperMix 6 μL Total RNA or mRNA Total RNA：10 pg-2 μg mRNA：10 pg-500 ng RNase-free H2O Up to 20 μL 2) Reverse Transcription Protocol Setup Temperature Time 50℃ 5 min 85℃ 5 sec 【Note】The reverse transcription product can be directly used for downstream qPCR detection. If the downstream experiment will not be performed immediately, the product can be stored at 4°C. For long-term storage, it is recommended to aliquot and store at -20°C to avoid repeated freeze-thaw cycles.

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Product description This product utilizes the poly(A) tailing method to synthesize the first-strand cDNA of miRNA. The enzymes and buffer system included in the product have been meticulously optimized to ensure that both the Poly(A) tailing at the 3' end of miRNA and the reverse transcription process can be carried out efficiently and simultaneously. For downstream qPCR applications, this product requires only the design of specific forward primers. In combination with the universal reverse primer provided in the kit, it enables the detection of miRNA in the sample. Additionally, the kit includes universal forward and reverse primers for U6 that are applicable to human, rat, and mouse samples, allowing for the generation of good standard curves over a wide range. This product is recommended for use with the miRNA qPCR Dye Premix (Cat#N132039) to achieve optimal experimental results. Specifications Catalog Number N132068E N132068S Specifications 10 T 50 T Components Catalog Number Component Name N132068E N132068S N132068-A miRNA RT Enzyme Mix 17.5 μL 87.5 μL N132068-B 2×miRNA RT Buffer 50 μL 250 μL N132068-C RNase-free H2O 2 mL 10 mL N132068-D Universal Reverse Primer (10 μM) 800 μL 4 mL N132068-E U6 Forward Primer (10 μM) 100 μL 500 μL N132068-F U6 Reverse Primer (10 μM) 100 μL 500 μL 【Note】Component C of the 4×cDNA Synthesis SuperMix contains the gDNA Digester terminator. Storage Transport with ice packs. Store at -20℃. Valid for 12 months. Notes 1. Before use, thaw the components completely and gently mix them before use. 2. During the experiment, use RNase-free consumables to avoid unnecessary losses that may affect the experimental results. 3. Avoid repeated freeze-thaw cycles of the product. When preparing the mixture, protect it from strong light. 4. For your safety and health, please wear a lab coat and disposable gloves when operating. 5. For Research Use Only. Instructions 1. Reaction System Preparation Thaw Component A (miRNA RT Enzyme Mix) and Component B (2× miRNA RT Buffer) at room temperature. After thawing, gently invert to mix and place on ice. Then, prepare the reaction system according to the table below: Component Volume（μL） Final concentration 2×miRNA RT buffer 5 μL 1× miRNA RT enzyme mix 1.75 μL - RNA - X RNase-free H2O Up to 10 - 【Note】For RNA samples, the concentration range of total RNA and extracted miRNA should be between 10 pg and 2 μg. The minimum number of miRNA copies that can be synthesized is 60 copies. The input volume should not exceed 3.25 μL. 2. Reverse Transcription Procedure Gently mix the prepared reaction mixture using a pipette. Perform the reverse transcription reaction for miRNA according to the program in the table below: Temperature Time Remarks 37℃ 50 min miRNA Poly(A) Tailing and Reverse Transcription Process 85℃ 5 sec The process of enzyme inactivation 【Note】 The reverse transcription product can be directly used for downstream qPCR detection. To avoid inhibition of the qPCR reaction by the reverse transcription system, the product can be diluted 10-1000 times before use. If the downstream experiment will not be performed immediately, the product can be stored at -20°C. For long-term storage, it is recommended to aliquot and store at -80°C to avoid repeated freeze-thaw cycles.

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Product description This product is a classic one-step cDNA synthesis reverse transcription ready-to-use premix from an RNA template. The core reverse transcriptase contained in the product has been effectively engineered to exhibit excellent thermal stability and template affinity, allowing it to tolerate reaction temperatures as high as 60°C. It is suitable for reverse transcription of RNA templates with complex secondary structures, low amounts of template, and low-copy genes. Additionally, this product contains a gDNA digestion component that can remove residual genomic DNA contamination from the RNA template, ensuring more reliable downstream results. The cDNA synthesized by this product is suitable for downstream qPCR applications. It is recommended to use it in combination with the Universal Multiplex qPCR Probe Premix (Cat#N132041), Universal qPCR Dye Premix (Cat#N132031), or the Traceable qPCR Dye Premix (Cat#N132034, Cat#N132035, Cat#N132036) for high-performance gene expression analysis. Specifications Catalog Number N132061E N132061S Specifications 10 T 100 T Components Catalog Number Component Name N132061E N132061S N132061-A RNase-free H2O 1 mL 2×1 mL N132061-B 5×gDNA Digester Mix 30 μL 320 μL N132061-C 4×cDNA Synthesis SuperMix 50 μL 500 μL 【Note】Component C of the 4×cDNA Synthesis SuperMix contains the gDNA Digester terminator. Storage Transport with ice packs. Store at -20℃. Valid for 18 months. Notes 1. Component B (5× gDNA Digester Mix) and Component C (4× cDNA Synthesis SuperMix) both contain high concentrations of glycerol. Before use, briefly centrifuge to collect the components at the bottom of the reaction tube. Then gently mix thoroughly by pipetting up and down, and accurately aspirate the required volume. 2. If the volume of the RNA template to be added is relatively large (e.g., more than 2 μL), ensure that the RNA is dissolved in water rather than TE buffer, as TE buffer may inhibit gDNA removal and reverse transcription. 3. The gDNA removal step can be omitted, and reverse transcription can be performed directly using Component C (4× cDNA Synthesis SuperMix). 4. The preparation of the reaction mixture should be carried out on ice to maintain stability. During the process, avoid contamination with RNase. 5. For your safety and health, please wear a lab coat and disposable gloves when operating. 6. For Research Use Only. Instructions 1. Removal of Residual Genomic DNA Prepare the following mixture in an RNase-free centrifuge tube and gently mix by pipetting. Incubate at 42°C for 2 minutes. Component Usage Amount RNase Free H2O To 15 μL 5×gDNA Digester Mix 3 μL Total RNA or mRNA RNA：10 pg-5 μg mRNA：10 pg-500 ng 【Note】For a 20 μL reverse transcription reaction system, it is recommended that the input amount of Total RNA does not exceed 1 μg. If the expression level of the target gene is low, up to 5 μg of Total RNA can be used. However, excessive RNA input may exceed the linear range of subsequent quantitative PCR. 2. Reverse Transcription Reaction Setup Directly add the 4× cDNA Synthesis SuperMix into the reaction tube from Step 1 and gently mix by pipetting. Component Usage Amount Step 1 Reaction Mixture 15 μL 4×cDNA Synthesis SuperMix 5 μL 3. Reverse Transcription Procedure Settings 1) Standard procedure Temperature Time 25℃ 5 min 55℃ 15 min 85℃ 5 min 【Note】The recommended reverse transcription temperature is 55℃. For templates with high GC content or complex structures, the reverse transcription temperature can be increased to 60℃. 2) Rapid Program [Suitable for templates with GC content ≤ 55% or simple templates] Temperature Time 55℃ 15 min 85℃ 5 sec 【Note】The reverse transcription product can be used immediately for qPCR reactions or stored short-term at -20°C. For long-term storage, it is recommended to aliquot and store at -80°C to avoid repeated freeze-thaw cycles.  

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