Xrn1, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 97/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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1) Product Images from "The conserved stem-loop II structure at the 3' untranslated region of Japanese encephalitis virus genome is required for the formation of subgenomic flaviviral RNA"
Article Title: The conserved stem-loop II structure at the 3' untranslated region of Japanese encephalitis virus genome is required for the formation of subgenomic flaviviral RNA
Journal: PLoS ONE
Figure Legend Snippet: Both knocking-down and knocking-out of XRN1 had no effect on the accumulation of sfRNA. XRN1-knockdown (KD) cells were prepared as described in Materials and Methods. Knockdown efficiency was determined by Western blot using anti-XRN1 and anti-β-actin antibodies. The Rel. % value represents the percentage of XRN1 expression in cells transfected with shXRN1 compared to wild-type cells (WT)(100%) as shown at the top (A-C). The XRN1-KD HEK293T (A) or A549 (B) cells were infected with JEV. Total RNA were extracted at the indicated times post infection and Northern blots were done with a DIG-labeled riboprobe detecting nt 10454 to nt 10976 in the 3’UTR (A, D, E) or an IRD 700-labeled JEV(-)10950-10976 probe (B). (C) XRN1-KD HEK293T cells were infected with DENV-2 at an MOI of 5 as a control. Total RNAs were extracted at 72 h post-infection and Northern blots were analyzed using a DIG-labeled riboprobe detecting nt 10270 to nt 10723 in the 3’UTR. Relative amounts of sfRNA were quantified (%) in the XRN1-depleted cells. (D) XRN1-knockout (KO) cells were infected with JEV or DENN-2 at an MOI of 5. RNA isolated from these cells at 48 h post-infection was subjected to Northern blot analysis. (E) RNA degradation analysis of non-replicative 800-nt 3’-terminal monophosphate transcripts derived from genome of JEV or DENV as indicated was measured in vitro by incubating with the indicated amounts of XRN1. Total RNAs extracted from JEV or DENV-2 infected cells (1 μg) were used as the sfRNA size marker (lanes 5 and 10). RNAs were separated by denaturing gel and analyzed by Northern hybridization.
Techniques Used: Western Blot, Expressing, Transfection, Infection, Northern Blot, Labeling, Knock-Out, Isolation, Derivative Assay, In Vitro, Marker, Hybridization
Figure Legend Snippet: Schematic model depicting the mechanism of JEV sfRNA formation. The JEV sfRNA is likely made (i) by transcription initially by RdRp in conjunction with other host factors (HFs), and (ii) the synthesized products could be further trimmed by exoribonuclease XRN1 and/or other unidentified enzymes.
Techniques Used: Synthesized