The kit uses revertaid reverse transcriptase rt, which has lower rnase h activity compared to amv reverse transcriptase. The enzyme maintains activity at 4250c and is suitable for synthesis of cdna up to kb. The length of cdna achieved is verified by detection of a 9. Different amounts of total rna from k562 cells were reverse transcribed using random hexamer primers and the reaction conditions outlined in the pack insert of the kit. The goscript reverse transcription system is a convenient kit that includes a reverse transcriptase and an optimized set of. Firststrand cdna synthesis for quantitative rtpcr origene. Following first strand cdna synthesis, the sample can be used directly for in vitro amplification using pcr or for second strand synthesis using the gublerhoffman method. Sigmaaldrich offers roche11483188001, first strand cdna synthesis kit for rtreverse transcriptasepcr amv for your research needs. Biorad iscript cdna synthesis kit invitrogen superscript iii firststrand synthesis supermix quanta qscript cdna synthesis kit quanta qscript cdna supermix qiagen quantitect reverse transcription. For robust twostep rtpcr and rt qpcr of rna up to 14 kb. First strand cdna synthesis kit can be used to synthesize the first strand cdna by reverse transcription rt at higher temperatures than the wild type mmulv to reach higher cdna yields for.
Researchers can choose from among a wide variety of commercial cdna synthesis kits. The first is the difficulty in reaching the cap site in the firststrand synthesis with the reverse transcriptase rt, the first enzyme involved in the preparation of a cdna library. The thermo scientific reverse transcription portfolio offers robust and reliable reverse transcriptases in a variety of optimized formats for efficiency in rtpcr and rtqpcr applications. First strand cdna synthesis for qrtpcr realtime pcr. First strand cdna synthesis starts from the 3end cds. Transcriptor first strand cdna synthesis kit roche life science. However, this step can be omitted in many cases unpublished results. Thermo scientific revertaid first strand cdna synthesis kit is a complete system for efficient synthesis of first strand cdna from rna templates. The enzyme mix combines protoscript ii reverse transcriptase and murine rnase inhibitor, while the reaction mix contains dntps and an optimized buffer. Thermo scientific revertaid h minus first strand cdna synthesis kit is a complete system for efficient synthesis of first strand cdna from mrna or total rna reverse transcriptase, which has a point mutation that completely eliminates rnase h activity. Efficient cdna synthesis by allinone firststrand cdna synthesis kit. The kit uses revertaid reverse transcriptase rt, a. Add the following components in the indicated order.
Omniscript reverse transcription handbook en print bookmark share omniscript reverse transcriptase for firststrand cdna synthesis twotube rtpcr pdf 381kb english format file size. For use with application firststrand cdna synthesis, rtpcr. Vectorprimed cdna synthesis the preparation of cdna libraries with combined vectorprimers is detailed in this volume 41. Mix the following components and incubate at 42c for 1 hour. The transcriptor first strand cdna synthesis kit is designed to reverse transcribe rna mrna, total rna, viral rna, and in vitrotranscribed rna from a variety of sources for the following applications. Protoscript first strand cdna synthesis kit features two optimized mixes, mmulv enzyme mix and mmulv reaction mix. Description legene express 1st strand cdna synthesis system is optimized to synthesize high yield cdna from total or poly a. The first strand cdna kit combines genomic dna elimination of rna samples and subsequent first strand cdna synthesis in one easy workflow to ensure accuracy of realtime rtpcr results. It can be used with either sequencespecific primers, polydt15 primers, or. Efficient synthesis of fulllength first strand cdna up to kb optimum activity at 42c active up to 50c incorporates modified nucleotides e. For reliable first strand cdna synthesis from all rna sources. It can be used with either sequencespecific primers, polydt15 primers, or random primers, pdn6.
The reverse transcriptase rt enzyme synthesizes the new cdna strand at a site determined by the type of primer used. The transcriptor first strand cdna synthesis kit is designed to reverse transcribe rna mrna, total rna, viral rna, and in vitro transcribed rna from a. Rna isolationpurification and firststrand cdna synthesis. Transcriptor first strand cdna synthesis kit roche. The primescript first strand cdna synthesis kit contains all of the reagents necessary for synthesis of firststrand cdna from total or. The first strand cdna can then be used as a template for pcr. Rna isolationpurification and firststrand cdna synthesis entered by kevin janes janes lab protocols 122816 2 wet pellets can be warmed for a few seconds at 37oc to increase evaporation smaller or broken pellets will need to have the remaining liquid evaporated off at 42. Reverse transcription rt is a widely used application in molecular biology research for rna and expression analysis. For firststrand cdna synthesis twotube rtpcr pdf 381kb english. Ribolock rnase inhibitor, supplied with the kit, effectively protects rna templates from degradation.
Legene express 1st strand cdna synthesis system, size. Marligens firststrand cdna synthesis system for quantitative rtpcr has been designed for the highest efficiency conversion of rna to cdna and is. First strand cdna synthesis with high sensitivity and efficiency. Thermo scientific maxima first strand cdna synthesis kit for rtqpcr is a convenient system optimized for cdna synthesis in 2step quantitative rtpcr rtqpcr applications. The cdna reaction product can be used directly in the next step without being purified. Find product specific information including cas, msds, protocols and references. The goscript reverse transcription system is a convenient kit that includes a reverse transcriptase and an optimized set of reagents for efficient synthesis of firststrand cdna optimized in preparation for pcr amplification. Fermentas first strand cdna synthesis kit is a complete system for the efficient synthesis of first strand cdna from rna templates. This is the first time that i will be peforming qrtpcr. Using the first strand cdna synthesis kit, rna is reverse transcribed into single stranded cdna. Revertaid h minus first strand cdna synthesis kit is a complete system for efficient synthesis of first strand cdna from rna templates. Protoscript ii first strand cdna synthesis kit features two optimized mixes, protoscript ii enzyme mix and protoscript ii reaction mix. First strand cdna synthesis kit 11801025 the use of this others has been cited in the following citations controlling obesity and metabolic diseases by hydrodynamic delivery of a fusion gene of.
Therefore, degradation of rna does not occur during first strand. First strand cdna synthesis kit for rtpcr amv sigma. Easy protocol mix the following components and incubate at 42c for 1 hour. Applied biosystems high capacity cdna reverse transcription kit biorad iscript cdna synthesis kit invitrogen superscript iii first strand synthesis supermix quanta qscript cdna synthesis kit quanta qscript cdna supermix qiagen quantitect reverse transcription kit stratagene affinityscript qpcr cdna synthesis kit. The cdna synthesis procedure is illustrated in fig. Total rna isolated from human placenta was used as template rna in reverse transcription reactions using the allinonetm firststrand cdna synthesis kit together with the oligodt primer. First strand synthesis reaction denaturation of rna and primer at 6570c for 5 minutes can remove secondary structures that may impede long cdna synthesis. The kit includes revertaid h minus reverse transcriptase, which has a point mutation that completely eliminates rnase h activity. Thermo scientific revertaid rt kit is a complete system for efficient synthesis of first strand cdna from mrna or total rna templates. Thermo scientific revertaid rt reverse transcription kit. The first strand cdna synthesis kit is used for the synthesis of the first strand cdna as the starting reaction for rtpcr. Mint2 cdna synthesis kit utilizes the specific features of mint reverse transcriptase mint rt.
Here we present a method for synthesis of first strand cdna which makes use of the same components found to be advantageous with oligodt as the primer. First strand cdna synthesis kit is designed to generate fulllength first strand cdna from mrna templates. Mar 07, 2011 cdna synthesis exploits the viral enzyme reverse transcriptase rt, which uses mrna as a template to drive dna synthesis, a step called reverse transcription. Thermo scientific revertaid premium first strand cdna.
Firststrand cdna synthesis kit ge healthcare, 27926101. The enzyme mix combines protoscript ii reverse transcriptase and. First strand cdna synthesis protocols e6560 thaw kit components on ice and mix by inverting several times. First strand cdna synthesis kit 11801025 the use of this others has been cited in the following citations controlling obesity and metabolic diseases by hydrodynamic delivery of a fusion gene of exendin4 and. The kit uses maxima reverse transcriptase rt, an advanced enzyme derived by in vitro evolution of mmulv rt. The applicability of the transcriptor first strand cdna synthesis kit to partially. Revertaid first strand cdna synthesis kit is a complete system for efficient synthesis of first strand cdna from rna templates. The reverse transcription efficiency of the allinone first strand synthesis kit is assessed by examining the amplification results of different genes or gene regions using the oligodt synthesized cdna prepared from the allinone firststrand cdna kit. Some are designed strictly for first strand cdna synthesis.
Protoscript ii first strand cdna synthesis kit neb. The enzyme maintains activity at 4250 c and is suitable for. Firststrand cdna synthesis kit is designed to generate fulllength firststrand cdna from mrna templates. Following firststrand cdna synthesis, the sample can be used directly for in vitro.
Marligens firststrand cdna synthesis system for quantitative rtpcr has been designed for the highest efficiency conversion of rna to cdna and is fully optimized for quantitative realtime pcr applications. Betaine reduces pausing and stops by the reverse transcriptase, enabling improved reverse transcription through difficult sequences, such as regions of high gc content. The azuraquant cdna synthesis kit provides a rapid and sensitive method for generating high quality, qpcrready firststrand cdna for use in downstream realtime pcr. The first strand of cdna can be directly used as a template in pcr see rtpcr protocol, realtime pcr or in second strand cdna synthesis see protocol. Thermo scientific revertaid rt reverse transcription kit 500. The first is the difficulty in reaching the cap site in the first strand synthesis with the reverse transcriptase rt, the first enzyme involved in the preparation of a cdna library. Instruction manual protoscript ii first strand cdna.
If random primer mix is used, an incubation step at 25c for 5 minutes is recommended before the 42c incubation. Since there is no need to perform a separate dnase digestion and repurify your rna samples, your time and costs are saved. The maxima first strand cdna synthesis kit with dsdnase is optimized for use with. Quantification of the g6pdh gene with the transcriptor first strand cdna synthesis kit and the lightcycler 2. The kit includes revertaid h minus reverse transcriptase, which. First strand cdna synthesis kitsreverse transcriptionrt. The kit uses revertaid reverse transcriptase rt, a recombinant mmulv rt which maintains activity at 4250c and is suitable for synthesis of cdna up to kb. Mmulv enzyme mix combines mmulv reverse transcriptase and murine. The azuraquant cdna synthesis kit delivers both highly efficient firststrand synthesis and higher cdna yields, leading to enhanced reproducibility and data accuracy. Amv reverse transcriptase provides first strand cdna synthesis reactions with a broader optimal reaction temperature from 37c to 50c. Mmulv enzyme mix combines mmulv reverse transcriptase and murine rnase inhibitor, while mmulv reaction mix contains dntps and an optimized buffer. Oligodt primer, random primer or a sequencespecific primer. Thermo scientific revertaid h minus first strand cdna.
Accuscript high fidelity 1st strand cdna synthesis kit agilent. The thermo scientific revertaid rt kit is a complete system for efficient synthesis of first strand cdna from mrna or total rna templates. Normally, 2 l of the rt mixture is used as template for subsequent qpcr in a 25 l total volume. The azuraquant cdna synthesis kit exhibits highly reproducible, unbiased synthesis and representation of 5 and 3 mrna ends. Invitrogen cloned amv firststrand cdna synthesis kit. First strand cdna synthesis kit can be used to synthesize the first strand cdna by reverse transcription rt at higher temperatures than the wild type mmulv to reach higher cdna yields for difficult rna transcriptions. Protocol for cdna synthesis and qrtpcr cdna synthesis. Revertaid first strand cdna synthesis kit fermentas and the last, said to be the best but not explained to me why it is so. Thermo scientific revertaid h minus first strand cdna synthesis kit is a complete system for efficient synthesis of first strand cdna from rna templates. The thermo scientific revertaid first strand cdna synthesis kit is a complete system for efficient synthesis of first strand cdna from mrna or total rna templates. Primers are not included in the preassembled first strand reaction.
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