Description:
ShortCut®RNaseIII,usedwithitsmanganese-containingreactionbuffer,convertslongdouble-strandedRNAintoaheterogeneousmixofshort(18–25bp)interferingRNAs(siRNA)suitableforRNAinterferenceinmammaliancells(1–3).1.5units(1 µl)ofShortCutRNaseIIIissufficienttoconvert1µgofdsRNAintosiRNAsuitableforRNAinterferenceinmammaliancells.
ShortCutRNaseIIIdigestionofdsRNA:(A)VaryingamountsofShortCutRNaseIIIwereincubatedwith2µgofa500bpdsRNAfor20minutesat37°Cina50µlreaction.Digestswereanalyzedby20%TBEpolyacrylamideelectrophoresis.Markerlanecontainsamixtureof21bpsiRNAMarkerand100bpDNALadder(NEB#N3231).(B)dsRNAfragments(1kband175bp)weredigestedwithShortCutRNaseIII.Digestswereanalyzedby20%TBEpolyacrylamidegelelectrophoresis.
GFPSilencinginCOS-7Cells:COS-7cellsco-transfectedina24wellplatewithaplasmidexpressingGFPintheabsence(control)orthepresenceof30ng(4nM)ofGFPsiRNApreparedusingtheShortCutRNAiKit.Cellswerephotographed48hourspost-transfection.
Highlights
- Isolatedfromarecombinantsource
- Genesilencing
- Targetvalidation
- Suppliedwith10XReactionBuffer
ProductSource
AnE.colistraincontainingageneticfusionoftheE.coliRNaseIIIgene(rnc)andthegenecodingformaltosebindingprotein(MBP).ReagentsSupplied
Thefollowingreagentsaresuppliedwiththisproduct:
| Storeat(°C) | Concentration |
10XEDTA | | 500mM |
MnCl2 | | 10X |
GlycogenRNase-Free | | 10mg/ml |
ShortcutReactionBuffer | | 10X |