Refer to nanodrop protocol for instructions on use. Accuscript high fidelity 1st strand cdna synthesis kit agilent. Revertaid first strand cdna synthesis kit is a complete system for efficient synthesis of first strand cdna from rna templates. Coumestrol induces mitochondrial biogenesis by activating. The revertaid h minus first strand cdna synthesis kit contains. A control reaction without reverse transcriptase is recommended to examine the dna contamination in the samples. 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. Firststrand cdna synthesis depending on the intended use of the firststrand cdna, synthesis may be performed using different volumes of bulk firststrand cdna mix, and different types and amounts of primer and rna. Revertaid h minus reverse transcriptase maintains activity over a. Thermo scientific revertaid reverse transcriptase 200 u. 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.
The maxima first strand cdna synthesis kit for rtqpcr is capable of reproducible cdna synthesis from a wide range of total rna amounts 1 pg to 5 g at elevated temperatures 42 to 65c. Lightcycler faststart dna masterplus sybr green i, was then used with primers for g6pdh to quantify geneexpression levels. It differs from the mmulv rt by its structure and catalytic properties. Applications first strand cdna synthesis for pcr or rtpcr gene expression data validation by using rtpcr benefits active at temperatures up to 55 c. 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.
Revertaid reverse transcriptase 10,000 u50,000 u ep0441ep0442 revertaid h minus reverse transcriptase 10,000 u50,000 u ep0451ep0452 cdna synthesis kits revertaid first strand cdna synthesis kit 20 rxns100 rxns k1621k1622 revertaid h minus first strand cdna synthesis kit 20 rxns100 rxns k1631k1632. Add the following components in the indicated order. The amv first strand cdna synthesis kit combines amv enzyme mix and amv reaction mix. Thermo scientific revertaid h minus first strand cdna synthesis kit perform efficient synthesis of first strand cdna from rna templates with this complete synthesis system. Transcriptor first strand cdna synthesis kit roche life science. Synthesis kit is a complete system for efficient synthesis of first strand cdna from mrna or total rna templates. If the first strand cdna synthesis 33 dnarna hybrid itself is the starting material for second strand synthe sis this volume 35, omit step 3 and proceed immediately to step 4. The synthesis reaction can be completed in 15 to 30 minutes. Negative control reactions were carried out with 1x mmulv reaction mix. The first strand cdna synthesis kit is used for the synthesis of the first strand cdna as the starting reaction for rtpcr. Thermo scientific maxima h minus doublestranded cdna synthesis kit is a complete system for efficient synthesis of doublestranded cdna from total rna or mrna.
If random 9mers are used, the first strand synthesis reaction must be incubated at. Make a cdna synthesis reaction mix by adding the iscript reverse transcription supermix to the dnasetreated rna template according to the guidelines in table 4. First strand cdna synthesis kit direct pcr reagents and. Maxima first strand cdna synthesis kit for rtqpcr, with dsdnase. If the synthesis of second strand cdna is contemplated by the classi cal method this volume 34, rna must be hydrolyzed step 3. The transcriptor high fidelity cdna synthesis kit is designed to reverse transcribe rna with an increased fidelity cat. The accupower rt premix is a master mix for cdna synthesis that consists of an easy to resuspend, lyophilized mix of mmlv reverse transcriptase, rnase inhibitor, dntps, reaction buffer, tracking dye, and patented stabilizer. In numbered lanes except lane 3, reverse tran scription was. First strand cdna synthesis kitsreverse transcriptionrtpcr. The kit is highly efficient at producing fulllength cdna from long rna templates at temperatures between 4255. Protoscript ii first strand cdna synthesis kit features two optimized mixes, protoscript ii enzyme mix and protoscript ii reaction mix. Thermo scientific revertaid reverse transcriptase 200 ul. Truscript first strand cdna synthesis kit product insert. It can be used with either sequencespecific primers, polydt15 primers, or random primers, pdn6.
The enzyme possesses an rnadependent and dnadependent polymerase activity and a rnase h activity specific to rna in rnadna hybrids which is significantly lower than that of avian. Oligodt primer, random primer or a sequencespecific primer. The kit can provide high efficient polya modification and reverse transcription and efficiently synthesize the firststrand cdna from 20 pg2. This is recommended when the cdna synthesis reaction uses less than 10 ng of rna input. Mmlv reverse transcriptase 1ststrand cdna synthesis kit lucigen. Store the remaining first strand cdna sample at 20 c until use. It can be used with either sequencespecific primers, polydt15 primers, or. Thermo scientific revertaid rt reverse transcription kit. 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.
Sensifast cdna synthesis kit, 50 reactions, first strand synthesis, fulllength reverse transcription. Product information thermo scientific possesses rna. Add the following reagents into a sterile, nuclease free tube on ice in the indicated order. Accuscript high fidelity 1st strand cdna synthesis kit.
The enzyme maintains activity at 4250c and is suitable for synthesis of cdna up to kb. Revertaid first strand cdna synthesis kit fermentas and the last, said to be the best but not explained to me why it is so smarter race cdna amplification kit clontech. 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. Random 9mers, also provided with this kit, are efficient primers for the detection of multiple short rtpcr targets. The kit is suitable for synthesis of cdna up to kb. The allinone firststrand cdna synthesis kit includes a reverse transcriptase and a specialized set of reagents designed to yield firststrand cdna that is optimal for gene cloning, cdna library creation and quantitative pcr amplification. Avoid repeated freezethaw cycles for control neo pa rna, rnase. Revertaid h minus reverse transcriptase does not degrade rna in rnadna hybrids during synthesis of the first strand cdna, and therefore high yields of fulllength cdna from long templates are obtained. The amv enzyme mix is an optimized blend of amv reverse transcriptase and a murine rnase inhibitor. Thermo scientific revertaid first strand cdna synthesis kit. Firststrand cdna synthesis kit ge healthcare, 27926101. Thermo scientific revertaid rt kit is a complete system for efficient synthesis of first strand cdna from mrna or total rna templates. Thermo scientific molecular biology workflow solutions. The rna pcr kit was purchased from takara biotechnology co.
Readytouse master mix for cdna synthesis from total rna. Mint2 cdna synthesis kit utilizes the specific features of mint reverse transcriptase mint rt. Not recommended or intended for diagnosis of disease in humans or animals. After completion of cdna synthesis, use 12 l of the first strand reaction out of the 20 l for pcr amplification. Researchers can choose from among a wide variety of commercial cdna synthesis kits. Mix thoroughly by pipetting up and down several times. For robust twostep rtpcr and rt qpcr of rna up to 14 kb. Rna isolationpurification and firststrand cdna synthesis. The maxima first strand cdna synthesis kit contains oligodt 18 and random hexamer primers to prime synthesis of first strand cdna. Applications first strand cdna synthesis for rt pcr and real time rtpcr, see protocol on back page. 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. 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.
The first strand cdna product generated is up to 10 kb. Thermo scientific maxima first strand cdna synthesis kit for. Thermo scientific revertaid first strand cdna synthesis kit is a complete system for efficient synthesis of first strand cdna from rna templates. For robust twostep rtpcr and rtqpcr of rna up to 14 kb. Reverse transcriptases rts use an rna template and a primer complementary to the rna to direct the synthesis of the first strand cdna, which can be used directly as a template for the polymerase chain reaction pcr. The first strand cdna can then be used as a template for pcr. Instruction manual protoscript ii first strand cdna.
Rnas were isolated using the trizol reagent invitrogen, and cdnas were synthesized using a revertaid first strand cdna synthesis kit fermentas. The kit includes revertaid h minus reverse transcriptase, which has a point mutation that completely eliminates rnase h activity. Contains protoscript ii reverse transcriptase, an enzyme with increased thermostability and reduced rnase h activity. Revertaid h minus reverse transcriptase, ribolock rnase inhibitor, 5x reaction buffer, dntp mix, oligodt18 primer, random hexamer primer, control rna, control primers and nucleasefree water. Thermo scientific reverse transcription thermo fisher. The energicscript cdna synthesis kit is intended for cdna synthesis for twostep quantitative reverse transcriptionpcr qrtpcr applications, where amplicons are usually around 200 bp in length. Usb firststrand cdna synthesis kit for realtime pcr. Some are designed strictly for first strand cdna synthesis. 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. Quickly merge multiple pdf files or part of them into a single one. This kit is can be used in conjunction with shineprobe qpcr kits or with any other qpcr kit suitable for the application. 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. Firststrand cdna synthesis for quantitative rtpcr origene.
Thermo scientific revertaid reverse transcriptase rt is a recombinant mmulv rt. Only four to ten genetic differences between the strains circulating in the us and the original virus that was isolated from patients in. Hyperscripttm first strand synthesis kit is ideally organized for synthe sizing reaction of first strand cdna from purified mrna or total rna. If the first strand cdna synthesis 33 dnarna hybrid itself is the starting material for second strand synthe sis this volume 35. The goscript reverse transcription system is a convenient kit that includes a reverse transcriptase and an optimized set of reagents for efficient synthesis of first strand cdna optimized in preparation for pcr amplification. If the cdna is to be used for secondstrand synthesis by the.
For reliable first strand cdna synthesis from all rna sources. Astragalus membranaceus inhibits inflammation via phospho. Thermo scientific revertaid h minus first strand cdna. K1612, first strand cdna synthesis, rev15 thermo fisher scientific. This primer mixture ensures high sensitivity in low copy number transcript detection assays. The cdna synthesis procedure is illustrated in fig. A robust experimental design delivers a universal kit that is suitable for first. The thermo scientific revertaid rt kit is a complete system for efficient synthesis of first strand cdna from mrna or total rna templates. Our pdf merger allows you to quickly combine multiple pdf files into one single pdf document, in just a few clicks. Thermo scientific ep0441 view more versions of this product catalog no. Once all the options are properly set, click run to start the execution. Using the revertaid first strand cdna synthesis kit quantify we need 0.
Efficient cdna synthesis by allinone firststrand cdna synthesis kit. The enzyme is capable of first strand cdna synthesis up to kb. Amv reverse transcriptase provides first strand cdna synthesis reactions with a broader optimal reaction temperature from 37c to 50c. Thermo scientific revertaid h minus reverse transcriptase. Mix rna sample and primer dt 23 vn in two sterile rnasefree microfuge tubes. Perform efficient synthesis of fulllength first strand cdna up to kb with this genetically modified mmulv rt for routine reverse transcription.
This kit is powered by primescript reverse transcriptase primescript rt, which has exceptionally strong strand displacement activity. Quantification of the g6pdh gene with the transcriptor first strand cdna synthesis kit and the lightcycler 2. The master mix kit is used for first strand cdna synthesis from rna. Experience with revertaid rt reverse transcription kit. A fraction of the first strand cdna product was used to amplify sequences specific for three different messenger rnas using 1x longamp taq 2x master mix. Either the transcriptor first strand cdna synthesis kit blue curves or another supplierss reverse transcriptase red curves was used to perform reverse transcription on 100 ng to 10 pg of total rna from k562 cells.
The kit uses revertaid reverse transcriptase rt, which has lower rnase h activity compared to amv reverse transcriptase. Use 1 g of isolated mrna to generate first strand cdna for secondstrand synthesis and subsequent cloning reactions. First strand cdna synthesis kit is designed to generate fulllength first strand cdna from mrna templates. In this experiment, cdnas prepared using each method were amplified in real time with primers specific for the 5 end of the adar gene using three different. The n cov19, it has been found, does not mutate rapidly. First strand cdna synthesis starts from the 3end cds adapter containing an oligodt sequence that anneals to polya stretches of rna. Thermo scientific revertaid h minus first strand cdna synthesis kit is a complete system for efficient synthesis of first strand cdna from rna templates. First strand cdna synthesis kit thermo scientific fermentas first strand cdna synthesis kit is a complete system for the efficient synthesis of first strand cdna from rna templates. Figure 2 shows that cdna prepared with the marligens first strand cdna synthesis system provides significantly enhanced sensitivity compared to cdna prepared with a competitors kit. Each entry pointing to the first page of the merged file. Sensifast cdna synthesis kit, 50 reactions, first strand.
Thermo scientific revertaid rt reverse transcription kit 500. The revertaid first strand cdna synthesis kit was purchased from fermentas international inc. Revertaid h minus first strand cdna synthesis kit is a complete system for efficient synthesis of first strand cdna from rna templates. First and second strand cdna synthesis reactions are performed in the same tube without the need for intermediate organic extraction or ethanol precipitation steps. First strand cdna synthesis after thawing, mix and briefly centrifuge the components of the kit. First strand cdna synthesis protocols e6300 introduction. The mmlv reverse transcriptase 1ststrand cdna synthesis kit is optimized for. Protocol after thawing, mix and briefly centrifuge the components of the kit. Hyperscript first strand synthesis kit 1 bohemia genetics. Highly efficient at producing fulllength cdna from as. Using the first strand cdna synthesis kit, rna is reverse transcribed into single stranded cdna. Includes dntps, oligodt primer and random primer mix. 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.
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