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HARTMANN ANALYTIC α 32 p atp utp
Cid14/Cid16 and Rrp6 degrade Argonaute-bound sRNAs. ( a ) Western blotting analysis of co-immunoprecipitation assay showing that Argonaute interacts with Cid14 in vivo . ( b ) Autoradiograph of denaturing polyacrylamide gel showing Cid14 and Cid16 activity on Argonaute-bound sRNA. 22 nucleotide long sRNA was loaded onto empty Argonaute purified from dcr1Δtri1Δ cells 9 . Argonaute was incubated with Cid14/Cid16 and 32 P <t>α-ATP/α-UTP</t> and sRNA was analysed on denaturing polyacrylamide gel. ( c ) Quantification of Argonaute-bound sRNAs that have non-templated nucleotides at the 3′ end in indicated strains. ( d , e ) Autoradiograph of denaturing polyacrylamide gel showing degradation of Argonaute-bound sRNA by Cid14, Cid16 and Rrp6. 5′ 32 P-labelled 22 nucleotide long sRNA was loaded onto empty Argonaute purified from dcr1Δtri1 Δ cells. Argonaute was incubated with Cid14, Cid16 and Triman or Rrp6. ( e ) Autoradiograph of denaturing polyacrylamide gel showing degradation of Argonaute-bound sRNA by Cid16 and Rrp6. 5′ 32 P-labelled 22 nucleotide long sRNA was loaded onto empty Argonaute purified from dcr1Δtri1Δ cells. Argonaute was incubated with Cid16 and Triman or Rrp6. ( f ) Autoradiograph of denaturing polyacrylamide gel showing degradation of Argonaute-bound sRNA by Cid14 and Rrp6. 5′ 32 P-labelled 22 nucleotide long sRNA was loaded onto empty Argonaute purified from dcr1Δtri1Δ cells. Argonaute was incubated with Rrp6, Cid14 and Cid14DADA. 30 nucleotide long DNA was used as a loading control. ( g ) Autoradiograph of denaturing polyacrylamide gel showing degradation of Argonaute-bound sRNA by Cid16 and Rrp6. 5′ 32 P-labelled 22 nucleotide long sRNA was loaded onto empty Argonaute purified from dcr1Δtri1Δ cells. Argonaute was incubated with Rrp6, Cid16 and Cid16DADA. 30 nucleotide long DNA was used as a loading control. ( h , i ) Autoradiograph of denaturing polyacrylamide gel showing time course of Argonaute-bound sRNA degradation by Cid14/Cid16 and Rrp6. 32 P-labelled 22 nucleotide long sRNA was loaded onto empty Argonaute purified from dcr1Δtri1Δ cells. Argonaute was incubated with Rrp6, Cid14 ( h ) or Cid16 ( i ). Time when reaction was stopped is indicated above the image.
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1) Product Images from "Tailing and degradation of Argonaute-bound small RNAs protect the genome from uncontrolled RNAi"

Article Title: Tailing and degradation of Argonaute-bound small RNAs protect the genome from uncontrolled RNAi

Journal: Nature Communications

doi: 10.1038/ncomms15332

Cid14/Cid16 and Rrp6 degrade Argonaute-bound sRNAs. ( a ) Western blotting analysis of co-immunoprecipitation assay showing that Argonaute interacts with Cid14 in vivo . ( b ) Autoradiograph of denaturing polyacrylamide gel showing Cid14 and Cid16 activity on Argonaute-bound sRNA. 22 nucleotide long sRNA was loaded onto empty Argonaute purified from dcr1Δtri1Δ cells 9 . Argonaute was incubated with Cid14/Cid16 and 32 P α-ATP/α-UTP and sRNA was analysed on denaturing polyacrylamide gel. ( c ) Quantification of Argonaute-bound sRNAs that have non-templated nucleotides at the 3′ end in indicated strains. ( d , e ) Autoradiograph of denaturing polyacrylamide gel showing degradation of Argonaute-bound sRNA by Cid14, Cid16 and Rrp6. 5′ 32 P-labelled 22 nucleotide long sRNA was loaded onto empty Argonaute purified from dcr1Δtri1 Δ cells. Argonaute was incubated with Cid14, Cid16 and Triman or Rrp6. ( e ) Autoradiograph of denaturing polyacrylamide gel showing degradation of Argonaute-bound sRNA by Cid16 and Rrp6. 5′ 32 P-labelled 22 nucleotide long sRNA was loaded onto empty Argonaute purified from dcr1Δtri1Δ cells. Argonaute was incubated with Cid16 and Triman or Rrp6. ( f ) Autoradiograph of denaturing polyacrylamide gel showing degradation of Argonaute-bound sRNA by Cid14 and Rrp6. 5′ 32 P-labelled 22 nucleotide long sRNA was loaded onto empty Argonaute purified from dcr1Δtri1Δ cells. Argonaute was incubated with Rrp6, Cid14 and Cid14DADA. 30 nucleotide long DNA was used as a loading control. ( g ) Autoradiograph of denaturing polyacrylamide gel showing degradation of Argonaute-bound sRNA by Cid16 and Rrp6. 5′ 32 P-labelled 22 nucleotide long sRNA was loaded onto empty Argonaute purified from dcr1Δtri1Δ cells. Argonaute was incubated with Rrp6, Cid16 and Cid16DADA. 30 nucleotide long DNA was used as a loading control. ( h , i ) Autoradiograph of denaturing polyacrylamide gel showing time course of Argonaute-bound sRNA degradation by Cid14/Cid16 and Rrp6. 32 P-labelled 22 nucleotide long sRNA was loaded onto empty Argonaute purified from dcr1Δtri1Δ cells. Argonaute was incubated with Rrp6, Cid14 ( h ) or Cid16 ( i ). Time when reaction was stopped is indicated above the image.
Figure Legend Snippet: Cid14/Cid16 and Rrp6 degrade Argonaute-bound sRNAs. ( a ) Western blotting analysis of co-immunoprecipitation assay showing that Argonaute interacts with Cid14 in vivo . ( b ) Autoradiograph of denaturing polyacrylamide gel showing Cid14 and Cid16 activity on Argonaute-bound sRNA. 22 nucleotide long sRNA was loaded onto empty Argonaute purified from dcr1Δtri1Δ cells 9 . Argonaute was incubated with Cid14/Cid16 and 32 P α-ATP/α-UTP and sRNA was analysed on denaturing polyacrylamide gel. ( c ) Quantification of Argonaute-bound sRNAs that have non-templated nucleotides at the 3′ end in indicated strains. ( d , e ) Autoradiograph of denaturing polyacrylamide gel showing degradation of Argonaute-bound sRNA by Cid14, Cid16 and Rrp6. 5′ 32 P-labelled 22 nucleotide long sRNA was loaded onto empty Argonaute purified from dcr1Δtri1 Δ cells. Argonaute was incubated with Cid14, Cid16 and Triman or Rrp6. ( e ) Autoradiograph of denaturing polyacrylamide gel showing degradation of Argonaute-bound sRNA by Cid16 and Rrp6. 5′ 32 P-labelled 22 nucleotide long sRNA was loaded onto empty Argonaute purified from dcr1Δtri1Δ cells. Argonaute was incubated with Cid16 and Triman or Rrp6. ( f ) Autoradiograph of denaturing polyacrylamide gel showing degradation of Argonaute-bound sRNA by Cid14 and Rrp6. 5′ 32 P-labelled 22 nucleotide long sRNA was loaded onto empty Argonaute purified from dcr1Δtri1Δ cells. Argonaute was incubated with Rrp6, Cid14 and Cid14DADA. 30 nucleotide long DNA was used as a loading control. ( g ) Autoradiograph of denaturing polyacrylamide gel showing degradation of Argonaute-bound sRNA by Cid16 and Rrp6. 5′ 32 P-labelled 22 nucleotide long sRNA was loaded onto empty Argonaute purified from dcr1Δtri1Δ cells. Argonaute was incubated with Rrp6, Cid16 and Cid16DADA. 30 nucleotide long DNA was used as a loading control. ( h , i ) Autoradiograph of denaturing polyacrylamide gel showing time course of Argonaute-bound sRNA degradation by Cid14/Cid16 and Rrp6. 32 P-labelled 22 nucleotide long sRNA was loaded onto empty Argonaute purified from dcr1Δtri1Δ cells. Argonaute was incubated with Rrp6, Cid14 ( h ) or Cid16 ( i ). Time when reaction was stopped is indicated above the image.

Techniques Used: Western Blot, Co-Immunoprecipitation Assay, In Vivo, Autoradiography, Activity Assay, Purification, Incubation

Related Articles

Concentration Assay:

Article Title: Tailing and degradation of Argonaute-bound small RNAs protect the genome from uncontrolled RNAi
Article Snippet: sRNAs tailing assay Overall, 500 fmol to 1 pmol of double-strand or single strand 22 nucleotides long RNAs were incubated with 80 ng of Cid14 and Cid16 in buffer containing 1 mM Hepes pH 7.5, 0.5 mM MgCl2 , 0.5 mM MnCl2 , 25 mM KCl, 0.2 mM DTT, 40U Ribolock (Thermo Scientific) and 150 nM α-32 P-ATP/UTP (Hartmann Analytic) for 2 h at 32 °C. .. Double-strand RNA was obtained by incubating RNAs 71 and 72 ( ) at same molar concentration in annealing buffer (10 mM Tris pH 7.5, 50 mM KCl, 1 mM EDTA) at 95 °C for 5 min followed by incubation at RT for 1 h. RNA was extracted and detected as described above.

Incubation:

Article Title: Tailing and degradation of Argonaute-bound small RNAs protect the genome from uncontrolled RNAi
Article Snippet: .. sRNAs tailing assay Overall, 500 fmol to 1 pmol of double-strand or single strand 22 nucleotides long RNAs were incubated with 80 ng of Cid14 and Cid16 in buffer containing 1 mM Hepes pH 7.5, 0.5 mM MgCl2 , 0.5 mM MnCl2 , 25 mM KCl, 0.2 mM DTT, 40U Ribolock (Thermo Scientific) and 150 nM α-32 P-ATP/UTP (Hartmann Analytic) for 2 h at 32 °C. .. Double-strand RNA was obtained by incubating RNAs 71 and 72 ( ) at same molar concentration in annealing buffer (10 mM Tris pH 7.5, 50 mM KCl, 1 mM EDTA) at 95 °C for 5 min followed by incubation at RT for 1 h. RNA was extracted and detected as described above.

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    HARTMANN ANALYTIC α 32 p atp utp
    Cid14/Cid16 and Rrp6 degrade Argonaute-bound sRNAs. ( a ) Western blotting analysis of co-immunoprecipitation assay showing that Argonaute interacts with Cid14 in vivo . ( b ) Autoradiograph of denaturing polyacrylamide gel showing Cid14 and Cid16 activity on Argonaute-bound sRNA. 22 nucleotide long sRNA was loaded onto empty Argonaute purified from dcr1Δtri1Δ cells 9 . Argonaute was incubated with Cid14/Cid16 and 32 P <t>α-ATP/α-UTP</t> and sRNA was analysed on denaturing polyacrylamide gel. ( c ) Quantification of Argonaute-bound sRNAs that have non-templated nucleotides at the 3′ end in indicated strains. ( d , e ) Autoradiograph of denaturing polyacrylamide gel showing degradation of Argonaute-bound sRNA by Cid14, Cid16 and Rrp6. 5′ 32 P-labelled 22 nucleotide long sRNA was loaded onto empty Argonaute purified from dcr1Δtri1 Δ cells. Argonaute was incubated with Cid14, Cid16 and Triman or Rrp6. ( e ) Autoradiograph of denaturing polyacrylamide gel showing degradation of Argonaute-bound sRNA by Cid16 and Rrp6. 5′ 32 P-labelled 22 nucleotide long sRNA was loaded onto empty Argonaute purified from dcr1Δtri1Δ cells. Argonaute was incubated with Cid16 and Triman or Rrp6. ( f ) Autoradiograph of denaturing polyacrylamide gel showing degradation of Argonaute-bound sRNA by Cid14 and Rrp6. 5′ 32 P-labelled 22 nucleotide long sRNA was loaded onto empty Argonaute purified from dcr1Δtri1Δ cells. Argonaute was incubated with Rrp6, Cid14 and Cid14DADA. 30 nucleotide long DNA was used as a loading control. ( g ) Autoradiograph of denaturing polyacrylamide gel showing degradation of Argonaute-bound sRNA by Cid16 and Rrp6. 5′ 32 P-labelled 22 nucleotide long sRNA was loaded onto empty Argonaute purified from dcr1Δtri1Δ cells. Argonaute was incubated with Rrp6, Cid16 and Cid16DADA. 30 nucleotide long DNA was used as a loading control. ( h , i ) Autoradiograph of denaturing polyacrylamide gel showing time course of Argonaute-bound sRNA degradation by Cid14/Cid16 and Rrp6. 32 P-labelled 22 nucleotide long sRNA was loaded onto empty Argonaute purified from dcr1Δtri1Δ cells. Argonaute was incubated with Rrp6, Cid14 ( h ) or Cid16 ( i ). Time when reaction was stopped is indicated above the image.
    α 32 P Atp Utp, supplied by HARTMANN ANALYTIC, used in various techniques. Bioz Stars score: 91/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/α 32 p atp utp/product/HARTMANN ANALYTIC
    Average 91 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    α 32 p atp utp - by Bioz Stars, 2020-04
    91/100 stars
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    Cid14/Cid16 and Rrp6 degrade Argonaute-bound sRNAs. ( a ) Western blotting analysis of co-immunoprecipitation assay showing that Argonaute interacts with Cid14 in vivo . ( b ) Autoradiograph of denaturing polyacrylamide gel showing Cid14 and Cid16 activity on Argonaute-bound sRNA. 22 nucleotide long sRNA was loaded onto empty Argonaute purified from dcr1Δtri1Δ cells 9 . Argonaute was incubated with Cid14/Cid16 and 32 P α-ATP/α-UTP and sRNA was analysed on denaturing polyacrylamide gel. ( c ) Quantification of Argonaute-bound sRNAs that have non-templated nucleotides at the 3′ end in indicated strains. ( d , e ) Autoradiograph of denaturing polyacrylamide gel showing degradation of Argonaute-bound sRNA by Cid14, Cid16 and Rrp6. 5′ 32 P-labelled 22 nucleotide long sRNA was loaded onto empty Argonaute purified from dcr1Δtri1 Δ cells. Argonaute was incubated with Cid14, Cid16 and Triman or Rrp6. ( e ) Autoradiograph of denaturing polyacrylamide gel showing degradation of Argonaute-bound sRNA by Cid16 and Rrp6. 5′ 32 P-labelled 22 nucleotide long sRNA was loaded onto empty Argonaute purified from dcr1Δtri1Δ cells. Argonaute was incubated with Cid16 and Triman or Rrp6. ( f ) Autoradiograph of denaturing polyacrylamide gel showing degradation of Argonaute-bound sRNA by Cid14 and Rrp6. 5′ 32 P-labelled 22 nucleotide long sRNA was loaded onto empty Argonaute purified from dcr1Δtri1Δ cells. Argonaute was incubated with Rrp6, Cid14 and Cid14DADA. 30 nucleotide long DNA was used as a loading control. ( g ) Autoradiograph of denaturing polyacrylamide gel showing degradation of Argonaute-bound sRNA by Cid16 and Rrp6. 5′ 32 P-labelled 22 nucleotide long sRNA was loaded onto empty Argonaute purified from dcr1Δtri1Δ cells. Argonaute was incubated with Rrp6, Cid16 and Cid16DADA. 30 nucleotide long DNA was used as a loading control. ( h , i ) Autoradiograph of denaturing polyacrylamide gel showing time course of Argonaute-bound sRNA degradation by Cid14/Cid16 and Rrp6. 32 P-labelled 22 nucleotide long sRNA was loaded onto empty Argonaute purified from dcr1Δtri1Δ cells. Argonaute was incubated with Rrp6, Cid14 ( h ) or Cid16 ( i ). Time when reaction was stopped is indicated above the image.

    Journal: Nature Communications

    Article Title: Tailing and degradation of Argonaute-bound small RNAs protect the genome from uncontrolled RNAi

    doi: 10.1038/ncomms15332

    Figure Lengend Snippet: Cid14/Cid16 and Rrp6 degrade Argonaute-bound sRNAs. ( a ) Western blotting analysis of co-immunoprecipitation assay showing that Argonaute interacts with Cid14 in vivo . ( b ) Autoradiograph of denaturing polyacrylamide gel showing Cid14 and Cid16 activity on Argonaute-bound sRNA. 22 nucleotide long sRNA was loaded onto empty Argonaute purified from dcr1Δtri1Δ cells 9 . Argonaute was incubated with Cid14/Cid16 and 32 P α-ATP/α-UTP and sRNA was analysed on denaturing polyacrylamide gel. ( c ) Quantification of Argonaute-bound sRNAs that have non-templated nucleotides at the 3′ end in indicated strains. ( d , e ) Autoradiograph of denaturing polyacrylamide gel showing degradation of Argonaute-bound sRNA by Cid14, Cid16 and Rrp6. 5′ 32 P-labelled 22 nucleotide long sRNA was loaded onto empty Argonaute purified from dcr1Δtri1 Δ cells. Argonaute was incubated with Cid14, Cid16 and Triman or Rrp6. ( e ) Autoradiograph of denaturing polyacrylamide gel showing degradation of Argonaute-bound sRNA by Cid16 and Rrp6. 5′ 32 P-labelled 22 nucleotide long sRNA was loaded onto empty Argonaute purified from dcr1Δtri1Δ cells. Argonaute was incubated with Cid16 and Triman or Rrp6. ( f ) Autoradiograph of denaturing polyacrylamide gel showing degradation of Argonaute-bound sRNA by Cid14 and Rrp6. 5′ 32 P-labelled 22 nucleotide long sRNA was loaded onto empty Argonaute purified from dcr1Δtri1Δ cells. Argonaute was incubated with Rrp6, Cid14 and Cid14DADA. 30 nucleotide long DNA was used as a loading control. ( g ) Autoradiograph of denaturing polyacrylamide gel showing degradation of Argonaute-bound sRNA by Cid16 and Rrp6. 5′ 32 P-labelled 22 nucleotide long sRNA was loaded onto empty Argonaute purified from dcr1Δtri1Δ cells. Argonaute was incubated with Rrp6, Cid16 and Cid16DADA. 30 nucleotide long DNA was used as a loading control. ( h , i ) Autoradiograph of denaturing polyacrylamide gel showing time course of Argonaute-bound sRNA degradation by Cid14/Cid16 and Rrp6. 32 P-labelled 22 nucleotide long sRNA was loaded onto empty Argonaute purified from dcr1Δtri1Δ cells. Argonaute was incubated with Rrp6, Cid14 ( h ) or Cid16 ( i ). Time when reaction was stopped is indicated above the image.

    Article Snippet: sRNAs tailing assay Overall, 500 fmol to 1 pmol of double-strand or single strand 22 nucleotides long RNAs were incubated with 80 ng of Cid14 and Cid16 in buffer containing 1 mM Hepes pH 7.5, 0.5 mM MgCl2 , 0.5 mM MnCl2 , 25 mM KCl, 0.2 mM DTT, 40U Ribolock (Thermo Scientific) and 150 nM α-32 P-ATP/UTP (Hartmann Analytic) for 2 h at 32 °C.

    Techniques: Western Blot, Co-Immunoprecipitation Assay, In Vivo, Autoradiography, Activity Assay, Purification, Incubation