ARNaseH-reversetrancriptasewithimprovedthermalstABIlity,efficiencyandprocessivityfortheproductionofCDNAfromlonganddifficultmRNAtemplates.
- Improvedthermalstability:Activeupto85ºC
- HighyieldsoffirststrandcDNA
- Highprocessivity:ProducescDNAoflongestmRNAsinthemammaliantranscriptome
- Greatsensitivity: WorkswithpgamountsofstartingtotalRNA
- Highreproducibility
- Robustandreliable
SunScript™ReverseTranscriptaseRNaseH-isaengineeredreversetranscriptasebasedonthewellcharacterizedHIV-1RTwithhighThermostabilityandnoRnaseHactivity.IthasRNA-andDNA-dependentpolymeraseactivity.Thisenzymeisactiveupto85°C.TheeliminationofRNaseHactivityenablesthisenzymetoproducehighyieldsoffulllengthcDNAfromlongRNAmolecules,andthehighthermalstabilitypromotesreversetransriptionthroughcomplexsecondarystructures.
- Reversetransciptionattemperaturesashighas85°C
- HighsensitivityoverawideRNAofstartingRNAamounts
- ReversetranscriptionoflongmRNAs
- HighcDNAsynthesisrate
Reversetransciptionattemperaturesashighas85°C
100ngoftotalRNAfromHeLacellswerereversetranscribedattheindicatedtemperaturesfor60minwithSunScriptReverseTranscriptasesandOligo(dT)18 primer,followedbyinactivationat95°Cfor10min.PresenceofGAPDHcDNAwasdetectedbyPCR(productlength305bp).
HighreactiontemperaturesareadvantageoustotranscribeRNAswithcomplicatedsecondarystructures.
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HighsensitivityoverawiderangeofRNAconcentrations
Sensitivitytest:RTreactionswereperformedat70°Cfor60min.Inactivationat95°Cfor10min.
Primerset:HumanGAPDH305bpset
1-1µgHumantotalRNAfromHeLacells
2-100ngHumantotalRNAfromHeLacells
3-10ngHumantotalRNAfromHeLacells
4-1ngHumantotalRNAfromHeLacells
5-100pgHumantotalRNAfromHeLacells
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TranscriptionoflongmRNAs
1-SunScript™RTRNaseH+replicate1
2-SunScript™RTRNaseH+replicate2
3-SunScript™RTRNaseH+replicate3
4-SunScript™RTRNaseH-replicate1
5-SunScript™RTRNaseH-replicate2
6-SunScript™RTRNaseH-replicate3
AssayforlongmRNAreversetranscription:3independentRTreactionswereperformedat65°Cfor60minusingOligo(dT)20 primers.Inactivationat95°Cfor10min.PCRPrimersets:mmNEB(>16kbfrom3’endofmRNA;mRNAtotallength22,5kb);mmDnah8(>8,5kbfrom3’endofmRNA;mRNAtotallength14,5kb)
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HighcDNAsynthesisrate
1µgoftotalRNAfromHeLacellswasreversetranscribedat65°CfortheindicatedtimeswithSunScript™ReverseTranscriptasesandOligo(dT)18primer,followedbyinactivationat95°C10min.
PCRPrimerset:hsHERC4033(>11kbfrom3’endofmRNA;mRNAtotallength15.3kb)
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ORDERINFORMATION
EachSunScript™RTRNaseH-contains:enzyme,5XReactionBufferandDTT.ThecompletemanualisavailableundertheManualtab.LucigenisanauthorizeddistributorofSygnisproductsintheUS.
SunScript™ReverseTranscriptaseRNaseH-isintendedformolecularBIOLOGyuseonlyandinvitrouseonly.Thisproductisnotintendedfordiagnosis,preventionortreatmentofadiseaseinhumanbeingsoranimals.
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我在做5RACE,选用TaKaRa公司的RNA反转录酶（RNaseH-），不知道这个酶好不好，有什么需要注意的。
没有序列识别的功能，他就是简单的催化相邻DNA链的5-P末端和3'-OH末端以磷酸二酯键结合的反应，需ATP作辅酶。 不仅可以催化粘性末端之间或平滑末端之间的DNA的连接，也催化DNA与RNA之间以及少数RNA之间的连接。
反转录酶（Reversetranscripatase）是以RNA为模板指导三磷酸脱氧核苷酸合成互补DNA（cDNA）的酶。哺乳类C型病毒的反转录酶和鼠类B型病毒的反转录酶都是一条多肽链。鸟类RNA病毒的反转录酶则由两上亚基结构。真核生物中也都分离出具有不同结构的反转录酶。
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