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.