rnase  (Worthington Biochemical)


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    Name:
    Ribonuclease A
    Description:
    A chromatographically purified lyophilized powder
    Catalog Number:
    LS003431
    Price:
    75
    Source:
    Bovine Pancreas
    Size:
    200 mg
    Cas Number:
    9001.99.4
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    Structured Review

    Worthington Biochemical rnase
    et-1 , tlr2 and tlr3 expression levels in fibroblasts stimulated with SSc-ICs or NHS-ICs pretreated with <t>DNase/RNase.</t> SSc-ICs treated with DNase I (20 KU/ml) or RNase (8 μg/ml) and then added to fibroblast cultures. a ATA-ICs, ACA-ICs and anti-Th/To-ICs on et-1 ; b ATA-ICs, ACA-ICs, ARA-ICs and anti-Th/To-ICs on tlr2 ; c ATA-ICs and anti-Th/To-ICs on ifn-α ; d ATA-ICs, ACA-ICs, ARA-ICs and anti-Th/To-ICs on tlr3 . * p
    A chromatographically purified lyophilized powder
    https://www.bioz.com/result/rnase/product/Worthington Biochemical
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    rnase - by Bioz Stars, 2021-07
    94/100 stars

    Images

    1) Product Images from "Immune complexes containing scleroderma-specific autoantibodies induce a profibrotic and proinflammatory phenotype in skin fibroblasts"

    Article Title: Immune complexes containing scleroderma-specific autoantibodies induce a profibrotic and proinflammatory phenotype in skin fibroblasts

    Journal: Arthritis Research & Therapy

    doi: 10.1186/s13075-018-1689-6

    et-1 , tlr2 and tlr3 expression levels in fibroblasts stimulated with SSc-ICs or NHS-ICs pretreated with DNase/RNase. SSc-ICs treated with DNase I (20 KU/ml) or RNase (8 μg/ml) and then added to fibroblast cultures. a ATA-ICs, ACA-ICs and anti-Th/To-ICs on et-1 ; b ATA-ICs, ACA-ICs, ARA-ICs and anti-Th/To-ICs on tlr2 ; c ATA-ICs and anti-Th/To-ICs on ifn-α ; d ATA-ICs, ACA-ICs, ARA-ICs and anti-Th/To-ICs on tlr3 . * p
    Figure Legend Snippet: et-1 , tlr2 and tlr3 expression levels in fibroblasts stimulated with SSc-ICs or NHS-ICs pretreated with DNase/RNase. SSc-ICs treated with DNase I (20 KU/ml) or RNase (8 μg/ml) and then added to fibroblast cultures. a ATA-ICs, ACA-ICs and anti-Th/To-ICs on et-1 ; b ATA-ICs, ACA-ICs, ARA-ICs and anti-Th/To-ICs on tlr2 ; c ATA-ICs and anti-Th/To-ICs on ifn-α ; d ATA-ICs, ACA-ICs, ARA-ICs and anti-Th/To-ICs on tlr3 . * p

    Techniques Used: Expressing, Acetylene Reduction Assay

    2) Product Images from "Immune complexes containing scleroderma-specific autoantibodies induce a profibrotic and proinflammatory phenotype in skin fibroblasts"

    Article Title: Immune complexes containing scleroderma-specific autoantibodies induce a profibrotic and proinflammatory phenotype in skin fibroblasts

    Journal: Arthritis Research & Therapy

    doi: 10.1186/s13075-018-1689-6

    et-1 , tlr2 and tlr3 expression levels in fibroblasts stimulated with SSc-ICs or NHS-ICs pretreated with DNase/RNase. SSc-ICs treated with DNase I (20 KU/ml) or RNase (8 μg/ml) and then added to fibroblast cultures. a ATA-ICs, ACA-ICs and anti-Th/To-ICs on et-1 ; b ATA-ICs, ACA-ICs, ARA-ICs and anti-Th/To-ICs on tlr2 ; c ATA-ICs and anti-Th/To-ICs on ifn-α ; d ATA-ICs, ACA-ICs, ARA-ICs and anti-Th/To-ICs on tlr3 . * p
    Figure Legend Snippet: et-1 , tlr2 and tlr3 expression levels in fibroblasts stimulated with SSc-ICs or NHS-ICs pretreated with DNase/RNase. SSc-ICs treated with DNase I (20 KU/ml) or RNase (8 μg/ml) and then added to fibroblast cultures. a ATA-ICs, ACA-ICs and anti-Th/To-ICs on et-1 ; b ATA-ICs, ACA-ICs, ARA-ICs and anti-Th/To-ICs on tlr2 ; c ATA-ICs and anti-Th/To-ICs on ifn-α ; d ATA-ICs, ACA-ICs, ARA-ICs and anti-Th/To-ICs on tlr3 . * p

    Techniques Used: Expressing, Acetylene Reduction Assay

    Related Articles

    Incubation:

    Article Title: Immune complexes containing scleroderma-specific autoantibodies induce a profibrotic and proinflammatory phenotype in skin fibroblasts
    Article Snippet: The protein secretion of MMP-2 and Pro-CollagenIα1, the mRNA levels of coIα1 and mmp-1 , and the activation rate of NFκB, p38MAPK, p54SAPK-JNK and p46SAPK-JNK were also evaluated in response to stimulation with ICs from SLE and PAPS sera. .. SSc-ICs were incubated for 1 h at 37 °C with recombinant DNase I or RNase (20 KU/ml and 8 μg/ml, respectively; Worthington Biochemical Corporation, Lakewood, NJ, USA) and then added to cells for 24 h. RT-PCR for tlr2 , tlr3 , ifn - α and et-1 was then performed. .. Cells were preincubated for 1 h at 37 °C with inhibitors of NFκB (MG-132, 20 μmol; Sigma-Aldrich) and p38MAPK (SB202190, 20 μmol; Cell Signaling Technology).

    Article Title: Immune complexes containing scleroderma-specific autoantibodies induce a profibrotic and proinflammatory phenotype in skin fibroblasts
    Article Snippet: SLE and APS immune complexes The protein secretion of MMP-2 and Pro-CollagenIα1, the mRNA levels of coIα1 and mmp-1 , and the activation rate of NFκB, p38MAPK, p54SAPK-JNK and p46SAPK-JNK were also evaluated in response to stimulation with ICs from SLE and PAPS sera. .. DNase and RNase treatment SSc-ICs were incubated for 1 h at 37 °C with recombinant DNase I or RNase (20 KU/ml and 8 μg/ml, respectively; Worthington Biochemical Corporation, Lakewood, NJ, USA) and then added to cells for 24 h. RT-PCR for tlr2 , tlr3 , ifn -α and et-1 was then performed. .. NFκB and p38MAPK inhibitors Cells were preincubated for 1 h at 37 °C with inhibitors of NFκB (MG-132, 20 μmol; Sigma-Aldrich) and p38MAPK (SB202190, 20 μmol; Cell Signaling Technology).

    Article Title: Mammalian DEAD Box Protein Ddx51 Acts in 3? End Maturation of 28S rRNA by Promoting the Release of U8 snoRNA ▿Mammalian DEAD Box Protein Ddx51 Acts in 3? End Maturation of 28S rRNA by Promoting the Release of U8 snoRNA ▿ §
    Article Snippet: To isolate proteins, we added 10 μg of bovine serum albumin to each 1-ml fraction, followed by trichloroacetic acid to a 10% final concentration. .. The pellet was washed with 1 ml of ethanol and 1 ml of acetone, air dried, resuspended in 20 mM Tris-HCl (pH 6.8)-1 mM EDTA-50 U/ml RNase A (Worthington), incubated for 15 min at 37°C, and mixed with SDS-PAGE loading buffer for separation on a gel and Western blotting. .. We identified Ddx51, a previously uncharacterized protein belonging to the DEAD box helicase family , in a two-hybrid screen for proteins interacting with the mammalian ribosome assembly factor Nog1.

    Recombinant:

    Article Title: Immune complexes containing scleroderma-specific autoantibodies induce a profibrotic and proinflammatory phenotype in skin fibroblasts
    Article Snippet: The protein secretion of MMP-2 and Pro-CollagenIα1, the mRNA levels of coIα1 and mmp-1 , and the activation rate of NFκB, p38MAPK, p54SAPK-JNK and p46SAPK-JNK were also evaluated in response to stimulation with ICs from SLE and PAPS sera. .. SSc-ICs were incubated for 1 h at 37 °C with recombinant DNase I or RNase (20 KU/ml and 8 μg/ml, respectively; Worthington Biochemical Corporation, Lakewood, NJ, USA) and then added to cells for 24 h. RT-PCR for tlr2 , tlr3 , ifn - α and et-1 was then performed. .. Cells were preincubated for 1 h at 37 °C with inhibitors of NFκB (MG-132, 20 μmol; Sigma-Aldrich) and p38MAPK (SB202190, 20 μmol; Cell Signaling Technology).

    Article Title: Immune complexes containing scleroderma-specific autoantibodies induce a profibrotic and proinflammatory phenotype in skin fibroblasts
    Article Snippet: SLE and APS immune complexes The protein secretion of MMP-2 and Pro-CollagenIα1, the mRNA levels of coIα1 and mmp-1 , and the activation rate of NFκB, p38MAPK, p54SAPK-JNK and p46SAPK-JNK were also evaluated in response to stimulation with ICs from SLE and PAPS sera. .. DNase and RNase treatment SSc-ICs were incubated for 1 h at 37 °C with recombinant DNase I or RNase (20 KU/ml and 8 μg/ml, respectively; Worthington Biochemical Corporation, Lakewood, NJ, USA) and then added to cells for 24 h. RT-PCR for tlr2 , tlr3 , ifn -α and et-1 was then performed. .. NFκB and p38MAPK inhibitors Cells were preincubated for 1 h at 37 °C with inhibitors of NFκB (MG-132, 20 μmol; Sigma-Aldrich) and p38MAPK (SB202190, 20 μmol; Cell Signaling Technology).

    Reverse Transcription Polymerase Chain Reaction:

    Article Title: Immune complexes containing scleroderma-specific autoantibodies induce a profibrotic and proinflammatory phenotype in skin fibroblasts
    Article Snippet: The protein secretion of MMP-2 and Pro-CollagenIα1, the mRNA levels of coIα1 and mmp-1 , and the activation rate of NFκB, p38MAPK, p54SAPK-JNK and p46SAPK-JNK were also evaluated in response to stimulation with ICs from SLE and PAPS sera. .. SSc-ICs were incubated for 1 h at 37 °C with recombinant DNase I or RNase (20 KU/ml and 8 μg/ml, respectively; Worthington Biochemical Corporation, Lakewood, NJ, USA) and then added to cells for 24 h. RT-PCR for tlr2 , tlr3 , ifn - α and et-1 was then performed. .. Cells were preincubated for 1 h at 37 °C with inhibitors of NFκB (MG-132, 20 μmol; Sigma-Aldrich) and p38MAPK (SB202190, 20 μmol; Cell Signaling Technology).

    Article Title: Immune complexes containing scleroderma-specific autoantibodies induce a profibrotic and proinflammatory phenotype in skin fibroblasts
    Article Snippet: SLE and APS immune complexes The protein secretion of MMP-2 and Pro-CollagenIα1, the mRNA levels of coIα1 and mmp-1 , and the activation rate of NFκB, p38MAPK, p54SAPK-JNK and p46SAPK-JNK were also evaluated in response to stimulation with ICs from SLE and PAPS sera. .. DNase and RNase treatment SSc-ICs were incubated for 1 h at 37 °C with recombinant DNase I or RNase (20 KU/ml and 8 μg/ml, respectively; Worthington Biochemical Corporation, Lakewood, NJ, USA) and then added to cells for 24 h. RT-PCR for tlr2 , tlr3 , ifn -α and et-1 was then performed. .. NFκB and p38MAPK inhibitors Cells were preincubated for 1 h at 37 °C with inhibitors of NFκB (MG-132, 20 μmol; Sigma-Aldrich) and p38MAPK (SB202190, 20 μmol; Cell Signaling Technology).

    Synthesized:

    Article Title: PYRIDOXAMINE ANALOGS SCAVENGE LIPID-DERIVED ?-KETOALDEHYDES AND PROTECT AGAINST H2O2-MEDIATED CYTOTOXICITY †
    Article Snippet: .. Salicylamine was purchased from Fischer Scientific USA (Pittsburgh, PA) and additional salicylamine hydrochloride was synthesized by the method of Reany et al ( ) RNase A was obtained from Worthington Biochemical (Lakewood, NJ). .. Dazoxiben was a generous gift from Pfizer Limited (Sandwich, U.K.).

    Labeling:

    Article Title: Defining the divergent enzymatic properties of RNA polymerases I and II
    Article Snippet: ECs are formed in buffer A and labeled as described for the single-nucleotide addition assay. .. At t = 0, labeled ECs are mixed in equal amount with 10 μM RNase A (Worthington Biochemical, Lakewood, NJ) and 1 M KCl. ..

    Purification:

    Article Title: Protein Phase Behavior in Aqueous Solutions: Crystallization, Liquid-Liquid Phase Separation, Gels, and Aggregates
    Article Snippet: Third, the correlation between the positions of the aggregation and solubility lines and the second osmotic virial coefficient is discussed, and finally, failure of the theoretical phase diagram ( ) to capture the temperature dependence observed experimentally is emphasized to show the limit of current theoretical approaches. .. Ovalbumin was obtained from fresh single-comb white Leghorn eggs following the purification protocol used by Judge et al. ( , ). β -Lactoglobulin from bovine milk (L0130) and lysozyme from chicken egg white (L6876) were purchased from Sigma (St. Louis, MO), STI (21730) was obtained from USB (Cleveland, OH), and ribonuclease A (LS003433) was purchased from Worthington (Lakewood, NJ). .. Sodium chloride (S-271) and sodium phosphate (P-285) were obtained from Fisher Scientific (Hampton, NH).

    SDS Page:

    Article Title: Mammalian DEAD Box Protein Ddx51 Acts in 3? End Maturation of 28S rRNA by Promoting the Release of U8 snoRNA ▿Mammalian DEAD Box Protein Ddx51 Acts in 3? End Maturation of 28S rRNA by Promoting the Release of U8 snoRNA ▿ §
    Article Snippet: To isolate proteins, we added 10 μg of bovine serum albumin to each 1-ml fraction, followed by trichloroacetic acid to a 10% final concentration. .. The pellet was washed with 1 ml of ethanol and 1 ml of acetone, air dried, resuspended in 20 mM Tris-HCl (pH 6.8)-1 mM EDTA-50 U/ml RNase A (Worthington), incubated for 15 min at 37°C, and mixed with SDS-PAGE loading buffer for separation on a gel and Western blotting. .. We identified Ddx51, a previously uncharacterized protein belonging to the DEAD box helicase family , in a two-hybrid screen for proteins interacting with the mammalian ribosome assembly factor Nog1.

    Western Blot:

    Article Title: Mammalian DEAD Box Protein Ddx51 Acts in 3? End Maturation of 28S rRNA by Promoting the Release of U8 snoRNA ▿Mammalian DEAD Box Protein Ddx51 Acts in 3? End Maturation of 28S rRNA by Promoting the Release of U8 snoRNA ▿ §
    Article Snippet: To isolate proteins, we added 10 μg of bovine serum albumin to each 1-ml fraction, followed by trichloroacetic acid to a 10% final concentration. .. The pellet was washed with 1 ml of ethanol and 1 ml of acetone, air dried, resuspended in 20 mM Tris-HCl (pH 6.8)-1 mM EDTA-50 U/ml RNase A (Worthington), incubated for 15 min at 37°C, and mixed with SDS-PAGE loading buffer for separation on a gel and Western blotting. .. We identified Ddx51, a previously uncharacterized protein belonging to the DEAD box helicase family , in a two-hybrid screen for proteins interacting with the mammalian ribosome assembly factor Nog1.

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  • 91
    Worthington Biochemical rnase t1
    LC-MS analysis of RNaseT1 digests of yeast tRNA Phe−1 . ( a ) Base peak chromatogram of the <t>RNase</t> T1 digest of the yeast tRNA Phe−1 preparation (200 fmol). Chromatography was performed as described in the ‘Materials and Methods’ section. Sixteen major oligoribonucleotide peaks, indicated by arrows with peak numbers, are assigned to the fragments of yeast tRNA Phe−1 , tRNA Phe−2 , tRNA Lys−2 , or tRNA Tyr (see Table 1 ). ( b and c ) A typical MS spectrum of the RNase T1 digest of tRNA Phe−1 ; (b) [AUUUAm 2 G > p] 2− and (c) [ACmUGmAAyWAΨUm 5 CUG > p] 3− .
    Rnase T1, supplied by Worthington Biochemical, used in various techniques. Bioz Stars score: 91/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Average 91 stars, based on 1 article reviews
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    94
    Worthington Biochemical rnase a
    p50 binds both to the 5′ end of osk ) was incubated and UV cross-linked to proteins in the oocyte extract in the absence of competitor RNA (lane 1 ). Competitions were carried out with a 100-fold excess of either repressor fragment (rep, lane 2 ), Eco RI– Bgl II fragment containing the 5′ activator in sense orientation (5′/act-s, lane 3), or the same Eco RI– Bgl II fragment in antisense orientation (5′/act-as, lane 4 ). Immunoprecipitation of proteins UV cross-linked to radiolabeled repressor fragment (lane 5 ) by anti-Bruno antiserum (lane 6 ) or preimmune serum (lane 7 ). Simultaneous binding of p50 and Bruno was tested by treating with <t>RNase</t> A only after the immunoprecipitation. (Lane 8 ) Anti-Bruno antiserum; (lane 9 ) preimmune serum. We noted that in some cases (as shown here) the intensity of the upper band of the p50 doublet was reduced after coprecipitation. The locations of the 5′ and 3′ competitors in the osk transcript are indicated below. The radiolabeled probe is indicated by an asterisk. The reverse experiment, with activator element RNA as a radioactive probe, and cold activator and repressor RNA elements as unlabeled competitors, yielded the same result with respect to p50 (data not shown).
    Rnase A, supplied by Worthington Biochemical, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rnase a/product/Worthington Biochemical
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
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    93
    Worthington Biochemical dnase 1
    Identification of NET-associated proteins. (A) Silver stained SDS-PAGE and (B) immunoblots with samples from NET protein purification procedure. Human neutrophils were stimulated to form NETs. Supernatants from unstimulated (lane 1) and stimulated (lane 2) neutrophils; first wash (lane 3); second wash (lane 4); medium containing <t>DNase-1</t> incubated with unstimulated neutrophils (lane 5); DNase-1-free medium incubated with washed NETs (lane 6); medium containing DNase-1 incubated with washed NETs (lane 7); medium containing DNase-1 incubated with washed NETs including protease inhibitor cocktail (lane 8).
    Dnase 1, supplied by Worthington Biochemical, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/dnase 1/product/Worthington Biochemical
    Average 93 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    dnase 1 - by Bioz Stars, 2021-07
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    N/A
    Worthington RNase Code R filtered through a 0 22 micron pore size membrane and lyophilized in sterile vials containing 100 mg vial
      Buy from Supplier

    Image Search Results


    LC-MS analysis of RNaseT1 digests of yeast tRNA Phe−1 . ( a ) Base peak chromatogram of the RNase T1 digest of the yeast tRNA Phe−1 preparation (200 fmol). Chromatography was performed as described in the ‘Materials and Methods’ section. Sixteen major oligoribonucleotide peaks, indicated by arrows with peak numbers, are assigned to the fragments of yeast tRNA Phe−1 , tRNA Phe−2 , tRNA Lys−2 , or tRNA Tyr (see Table 1 ). ( b and c ) A typical MS spectrum of the RNase T1 digest of tRNA Phe−1 ; (b) [AUUUAm 2 G > p] 2− and (c) [ACmUGmAAyWAΨUm 5 CUG > p] 3− .

    Journal: Nucleic Acids Research

    Article Title: An analytical platform for mass spectrometry-based identification and chemical analysis of RNA in ribonucleoprotein complexes

    doi: 10.1093/nar/gkp732

    Figure Lengend Snippet: LC-MS analysis of RNaseT1 digests of yeast tRNA Phe−1 . ( a ) Base peak chromatogram of the RNase T1 digest of the yeast tRNA Phe−1 preparation (200 fmol). Chromatography was performed as described in the ‘Materials and Methods’ section. Sixteen major oligoribonucleotide peaks, indicated by arrows with peak numbers, are assigned to the fragments of yeast tRNA Phe−1 , tRNA Phe−2 , tRNA Lys−2 , or tRNA Tyr (see Table 1 ). ( b and c ) A typical MS spectrum of the RNase T1 digest of tRNA Phe−1 ; (b) [AUUUAm 2 G > p] 2− and (c) [ACmUGmAAyWAΨUm 5 CUG > p] 3− .

    Article Snippet: RNase T1 was purchased from Worthington (Lakewood, NJ) and further purified by reversed-phase liquid chromatography (RPLC) before use.

    Techniques: Liquid Chromatography with Mass Spectroscopy, Chromatography, Mass Spectrometry

    CID-MS/MS spectrum of [AUUUAm 2 G] 2− derived from RNase T1 digestion of yeast tRNA Phe−1 . The doubly charged ion with m / z = 966.6 was analyzed by CID. The nucleotide sequence was verified by manual interpretation of the a- and c-type (normal text) and w - and y -type ( italic text ) product ion series as indicated in the figure. The parent ion losing methyl guanine [P−B(mG) 2− ], the parent ion losing adenine [P−B(A) 2− ], the y5 ion losing methyl guanine [y5−B(mG) 2− ], the y5 ion losing adenine [y5−B(A) 2− ], y5 2− and c5 2− were doubly charged products. All other assigned signals were singly charged products, unless indicated otherwise. The asterisks indicate hydrated or dehydrated ions of a-, c-, w- or y-type products.

    Journal: Nucleic Acids Research

    Article Title: An analytical platform for mass spectrometry-based identification and chemical analysis of RNA in ribonucleoprotein complexes

    doi: 10.1093/nar/gkp732

    Figure Lengend Snippet: CID-MS/MS spectrum of [AUUUAm 2 G] 2− derived from RNase T1 digestion of yeast tRNA Phe−1 . The doubly charged ion with m / z = 966.6 was analyzed by CID. The nucleotide sequence was verified by manual interpretation of the a- and c-type (normal text) and w - and y -type ( italic text ) product ion series as indicated in the figure. The parent ion losing methyl guanine [P−B(mG) 2− ], the parent ion losing adenine [P−B(A) 2− ], the y5 ion losing methyl guanine [y5−B(mG) 2− ], the y5 ion losing adenine [y5−B(A) 2− ], y5 2− and c5 2− were doubly charged products. All other assigned signals were singly charged products, unless indicated otherwise. The asterisks indicate hydrated or dehydrated ions of a-, c-, w- or y-type products.

    Article Snippet: RNase T1 was purchased from Worthington (Lakewood, NJ) and further purified by reversed-phase liquid chromatography (RPLC) before use.

    Techniques: Mass Spectrometry, Derivative Assay, Sequencing

    Base peak chromatogram of the RNaseT1 digest of yeast U4 snRNA isolated from the Lsm3-associated RNP complex. The gel piece containing U4 snRNA was in-gel digested with RNaseT1 and subjected to the LC-MS analysis. Major oligoribonucleotide peaks assigned as RNase T1 fragments of yeast U4 snRNA are indicated by arrows with the corresponding sequence. Detailed data for MS/MS-based assignment of each fragment are given in Supplementary Table S3 .

    Journal: Nucleic Acids Research

    Article Title: An analytical platform for mass spectrometry-based identification and chemical analysis of RNA in ribonucleoprotein complexes

    doi: 10.1093/nar/gkp732

    Figure Lengend Snippet: Base peak chromatogram of the RNaseT1 digest of yeast U4 snRNA isolated from the Lsm3-associated RNP complex. The gel piece containing U4 snRNA was in-gel digested with RNaseT1 and subjected to the LC-MS analysis. Major oligoribonucleotide peaks assigned as RNase T1 fragments of yeast U4 snRNA are indicated by arrows with the corresponding sequence. Detailed data for MS/MS-based assignment of each fragment are given in Supplementary Table S3 .

    Article Snippet: RNase T1 was purchased from Worthington (Lakewood, NJ) and further purified by reversed-phase liquid chromatography (RPLC) before use.

    Techniques: Isolation, Liquid Chromatography with Mass Spectroscopy, Sequencing, Mass Spectrometry

    Mt FRS activity. ( A ) Aminoacylation of unmodified M. tuberculosis tRNA Phe . The 3′- 32 P-labeled tRNA transcript was charged with L-Phe in the presence of purified Mt FRS. The extent of aminoacylation was analyzed according to Varshney et al. ( 37 ). Prior to polyacrylamide gel electrophoresis, tRNA and Phe-tRNA were digested with RNase T1 to yield 3′- 32 P-labeled CCACCA and CCACCA-Phe, respectively. (+) and (-) indicate the presence and the absence of the enzyme. (B–D) Synthesis of the Phe-AMP intermediate by Mt FRS in the presence of unmodified M. tuberculosis tRNA Phe ( B ), E. coli tRNA Phe ( C ) and yeast tRNA Phe ( D ). Phe-[ 32 P]AMP synthesis was monitored by thin layer chromatography. Phe-AMP and ATP denote 32 P-labeled compounds. F – full reaction; -E, -tRNA and -Phe – control reactions lacking the enzyme, tRNA or amino acid.

    Journal: Nucleic Acids Research

    Article Title: Mycobacterium tuberculosis Phe-tRNA synthetase: structural insights into tRNA recognition and aminoacylation

    doi: 10.1093/nar/gkab272

    Figure Lengend Snippet: Mt FRS activity. ( A ) Aminoacylation of unmodified M. tuberculosis tRNA Phe . The 3′- 32 P-labeled tRNA transcript was charged with L-Phe in the presence of purified Mt FRS. The extent of aminoacylation was analyzed according to Varshney et al. ( 37 ). Prior to polyacrylamide gel electrophoresis, tRNA and Phe-tRNA were digested with RNase T1 to yield 3′- 32 P-labeled CCACCA and CCACCA-Phe, respectively. (+) and (-) indicate the presence and the absence of the enzyme. (B–D) Synthesis of the Phe-AMP intermediate by Mt FRS in the presence of unmodified M. tuberculosis tRNA Phe ( B ), E. coli tRNA Phe ( C ) and yeast tRNA Phe ( D ). Phe-[ 32 P]AMP synthesis was monitored by thin layer chromatography. Phe-AMP and ATP denote 32 P-labeled compounds. F – full reaction; -E, -tRNA and -Phe – control reactions lacking the enzyme, tRNA or amino acid.

    Article Snippet: Prior to electrophoresis, on a 10% acid/7 M urea polyacrylamide gel, 10 pmoles of uncharged and aminoacylated tRNA (Phe-tRNAPhe) were completely digested with 2 U of RNase T1 (Worthington) in 10 mM sodium acetate (pH 4.5).

    Techniques: Activity Assay, Labeling, Purification, Polyacrylamide Gel Electrophoresis, Thin Layer Chromatography

    p50 binds both to the 5′ end of osk ) was incubated and UV cross-linked to proteins in the oocyte extract in the absence of competitor RNA (lane 1 ). Competitions were carried out with a 100-fold excess of either repressor fragment (rep, lane 2 ), Eco RI– Bgl II fragment containing the 5′ activator in sense orientation (5′/act-s, lane 3), or the same Eco RI– Bgl II fragment in antisense orientation (5′/act-as, lane 4 ). Immunoprecipitation of proteins UV cross-linked to radiolabeled repressor fragment (lane 5 ) by anti-Bruno antiserum (lane 6 ) or preimmune serum (lane 7 ). Simultaneous binding of p50 and Bruno was tested by treating with RNase A only after the immunoprecipitation. (Lane 8 ) Anti-Bruno antiserum; (lane 9 ) preimmune serum. We noted that in some cases (as shown here) the intensity of the upper band of the p50 doublet was reduced after coprecipitation. The locations of the 5′ and 3′ competitors in the osk transcript are indicated below. The radiolabeled probe is indicated by an asterisk. The reverse experiment, with activator element RNA as a radioactive probe, and cold activator and repressor RNA elements as unlabeled competitors, yielded the same result with respect to p50 (data not shown).

    Journal: Genes & Development

    Article Title: Localization-dependent translation requires a functional interaction between the 5? and 3? ends of oskar mRNA

    doi:

    Figure Lengend Snippet: p50 binds both to the 5′ end of osk ) was incubated and UV cross-linked to proteins in the oocyte extract in the absence of competitor RNA (lane 1 ). Competitions were carried out with a 100-fold excess of either repressor fragment (rep, lane 2 ), Eco RI– Bgl II fragment containing the 5′ activator in sense orientation (5′/act-s, lane 3), or the same Eco RI– Bgl II fragment in antisense orientation (5′/act-as, lane 4 ). Immunoprecipitation of proteins UV cross-linked to radiolabeled repressor fragment (lane 5 ) by anti-Bruno antiserum (lane 6 ) or preimmune serum (lane 7 ). Simultaneous binding of p50 and Bruno was tested by treating with RNase A only after the immunoprecipitation. (Lane 8 ) Anti-Bruno antiserum; (lane 9 ) preimmune serum. We noted that in some cases (as shown here) the intensity of the upper band of the p50 doublet was reduced after coprecipitation. The locations of the 5′ and 3′ competitors in the osk transcript are indicated below. The radiolabeled probe is indicated by an asterisk. The reverse experiment, with activator element RNA as a radioactive probe, and cold activator and repressor RNA elements as unlabeled competitors, yielded the same result with respect to p50 (data not shown).

    Article Snippet: Fifty micrograms of RNase A and 4 units of RNase T1 (Worthington Biochemical) were used.

    Techniques: Incubation, Activated Clotting Time Assay, Immunoprecipitation, Binding Assay

    Identification of NET-associated proteins. (A) Silver stained SDS-PAGE and (B) immunoblots with samples from NET protein purification procedure. Human neutrophils were stimulated to form NETs. Supernatants from unstimulated (lane 1) and stimulated (lane 2) neutrophils; first wash (lane 3); second wash (lane 4); medium containing DNase-1 incubated with unstimulated neutrophils (lane 5); DNase-1-free medium incubated with washed NETs (lane 6); medium containing DNase-1 incubated with washed NETs (lane 7); medium containing DNase-1 incubated with washed NETs including protease inhibitor cocktail (lane 8).

    Journal: PLoS Pathogens

    Article Title: Neutrophil Extracellular Traps Contain Calprotectin, a Cytosolic Protein Complex Involved in Host Defense against Candida albicans

    doi: 10.1371/journal.ppat.1000639

    Figure Lengend Snippet: Identification of NET-associated proteins. (A) Silver stained SDS-PAGE and (B) immunoblots with samples from NET protein purification procedure. Human neutrophils were stimulated to form NETs. Supernatants from unstimulated (lane 1) and stimulated (lane 2) neutrophils; first wash (lane 3); second wash (lane 4); medium containing DNase-1 incubated with unstimulated neutrophils (lane 5); DNase-1-free medium incubated with washed NETs (lane 6); medium containing DNase-1 incubated with washed NETs (lane 7); medium containing DNase-1 incubated with washed NETs including protease inhibitor cocktail (lane 8).

    Article Snippet: Subsequently the NETs were digested for 20 min in 1 ml RPMI with 10 U/ml DNase-1 (Worthington).

    Techniques: Staining, SDS Page, Western Blot, Protein Purification, Incubation, Protease Inhibitor

    Histones are altered during NET formation. NETs from human neutrophils were washed and digested with DNase-1. (A) The NET-fraction (N) and the remaining pellet after DNase-1 digest (P) were analyzed by immunoblotting at the indicated time points. Unstimulated neutrophils served as controls. All core histones have a reduced molecular mass (2–5 kDa less) in NETs compared to the pellet fraction and the unstimulated control. A representative experiment out of three in total is shown. (B) High-resolution SEM analysis of NETs which consist of smooth fibers (white box) and globular domains (diameter 25–50 nm, arrows), scale bar = 100 nm. (C) High-resolution FESEM analysis of smooth stretch of a singular NET-fiber. Signal intensities were profiled vertically and horizontally showing similar diameters to nucleosomes (depicted as cartoon structure models taken from [41] , with approximate horizontal and vertical diameters of 5 nm and 10 nm, respectively). One experiment out of two is shown.

    Journal: PLoS Pathogens

    Article Title: Neutrophil Extracellular Traps Contain Calprotectin, a Cytosolic Protein Complex Involved in Host Defense against Candida albicans

    doi: 10.1371/journal.ppat.1000639

    Figure Lengend Snippet: Histones are altered during NET formation. NETs from human neutrophils were washed and digested with DNase-1. (A) The NET-fraction (N) and the remaining pellet after DNase-1 digest (P) were analyzed by immunoblotting at the indicated time points. Unstimulated neutrophils served as controls. All core histones have a reduced molecular mass (2–5 kDa less) in NETs compared to the pellet fraction and the unstimulated control. A representative experiment out of three in total is shown. (B) High-resolution SEM analysis of NETs which consist of smooth fibers (white box) and globular domains (diameter 25–50 nm, arrows), scale bar = 100 nm. (C) High-resolution FESEM analysis of smooth stretch of a singular NET-fiber. Signal intensities were profiled vertically and horizontally showing similar diameters to nucleosomes (depicted as cartoon structure models taken from [41] , with approximate horizontal and vertical diameters of 5 nm and 10 nm, respectively). One experiment out of two is shown.

    Article Snippet: Subsequently the NETs were digested for 20 min in 1 ml RPMI with 10 U/ml DNase-1 (Worthington).

    Techniques: