Structured Review

GE Healthcare t4 rna ligase
Effect of various modifications on the 3′ terminal nucleotide of a small RNA on <t>T4</t> RNA ligase- and yeast PAP-catalyzed reactions. ( a ) T4 RNA ligase-mediated ligation of various miR173 forms to an RNA linker. ( b ) Activity of yeast PAP on various forms of miR173 in the presence of 2 pmol [α- 32 P]-ATP. The ladders or smears represent products of PAP-catalyzed reaction. ( c ) Activity of yeast PAP on various forms of miR173 in the presence of 10 pmol [α- 32 P]-ATP.
T4 Rna Ligase, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 85/100, based on 27 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Average 85 stars, based on 27 article reviews
Price from $9.99 to $1999.99
t4 rna ligase - by Bioz Stars, 2020-09
85/100 stars

Images

1) Product Images from "HEN1 recognizes 21-24 nt small RNA duplexes and deposits a methyl group onto the 2? OH of the 3? terminal nucleotide"

Article Title: HEN1 recognizes 21-24 nt small RNA duplexes and deposits a methyl group onto the 2? OH of the 3? terminal nucleotide

Journal: Nucleic Acids Research

doi: 10.1093/nar/gkj474

Effect of various modifications on the 3′ terminal nucleotide of a small RNA on T4 RNA ligase- and yeast PAP-catalyzed reactions. ( a ) T4 RNA ligase-mediated ligation of various miR173 forms to an RNA linker. ( b ) Activity of yeast PAP on various forms of miR173 in the presence of 2 pmol [α- 32 P]-ATP. The ladders or smears represent products of PAP-catalyzed reaction. ( c ) Activity of yeast PAP on various forms of miR173 in the presence of 10 pmol [α- 32 P]-ATP.
Figure Legend Snippet: Effect of various modifications on the 3′ terminal nucleotide of a small RNA on T4 RNA ligase- and yeast PAP-catalyzed reactions. ( a ) T4 RNA ligase-mediated ligation of various miR173 forms to an RNA linker. ( b ) Activity of yeast PAP on various forms of miR173 in the presence of 2 pmol [α- 32 P]-ATP. The ladders or smears represent products of PAP-catalyzed reaction. ( c ) Activity of yeast PAP on various forms of miR173 in the presence of 10 pmol [α- 32 P]-ATP.

Techniques Used: Ligation, Activity Assay

2) Product Images from "RISC is a 5? phosphomonoester-producing RNA endonuclease"

Article Title: RISC is a 5? phosphomonoester-producing RNA endonuclease

Journal: Genes & Development

doi: 10.1101/gad.1187904

Target RNA is cleaved endonucleolytically producing 5′-phosphate and 3′-hydroxyl termini. ( A ) Preparation of site-specifically labeled substrates and cleavage assay. 5′- 32 P-labeled and 3′ aminolinker (L) protected 12-nt oligoribonucleotide was ligated to nonphosphorylated 9-nt oligoribonucleotide using T4 RNA ligase. An aliquot of the ligation product was further 5′- 32 P-labeled using T4 polynucleotide kinase. The purified substrates were incubated with affinity-purified RISC programmed with single-stranded guide siRNA. ( B ) PhosphorImaging of cleavage reactions incubated for 2 h at 30°C, and resolved on a 20% denaturing polyacrylamide gel. 5′- 32 P-labeled 9- and 12-nt oligoribonucleotides were loaded as marker in lanes 1 and 2 , respectively. The cleavage reactions with single- and double-labeled 21-nt substrate are loaded in lanes 4 and 5 , respectively. Lane 3 contains the 12-nt cleavage product isolated from a prior cleavage reaction. ( C ) Two-dimensional thin layer chromatography analysis of the ribonuclease T2-digested RISC-cleavage product. The oval depicts the unlabeled pAp as detected by UV shadowing. The radioactive signal comigrates with the 5′ 32 pAp released by ribonuclease T2 digestion from the gel-purified 12-nt cleavage product.
Figure Legend Snippet: Target RNA is cleaved endonucleolytically producing 5′-phosphate and 3′-hydroxyl termini. ( A ) Preparation of site-specifically labeled substrates and cleavage assay. 5′- 32 P-labeled and 3′ aminolinker (L) protected 12-nt oligoribonucleotide was ligated to nonphosphorylated 9-nt oligoribonucleotide using T4 RNA ligase. An aliquot of the ligation product was further 5′- 32 P-labeled using T4 polynucleotide kinase. The purified substrates were incubated with affinity-purified RISC programmed with single-stranded guide siRNA. ( B ) PhosphorImaging of cleavage reactions incubated for 2 h at 30°C, and resolved on a 20% denaturing polyacrylamide gel. 5′- 32 P-labeled 9- and 12-nt oligoribonucleotides were loaded as marker in lanes 1 and 2 , respectively. The cleavage reactions with single- and double-labeled 21-nt substrate are loaded in lanes 4 and 5 , respectively. Lane 3 contains the 12-nt cleavage product isolated from a prior cleavage reaction. ( C ) Two-dimensional thin layer chromatography analysis of the ribonuclease T2-digested RISC-cleavage product. The oval depicts the unlabeled pAp as detected by UV shadowing. The radioactive signal comigrates with the 5′ 32 pAp released by ribonuclease T2 digestion from the gel-purified 12-nt cleavage product.

Techniques Used: Labeling, Cleavage Assay, Ligation, Purification, Incubation, Affinity Purification, Marker, Isolation, Thin Layer Chromatography

3) Product Images from "Combinatorial minimization and secondary structure determination of a nucleotide synthase ribozyme"

Article Title: Combinatorial minimization and secondary structure determination of a nucleotide synthase ribozyme

Journal: RNA

doi: 10.1261/rna.5500603

Selection scheme designed to retain sequence variation all the way to the 3′ end of RNA transcripts. ( A ) Pool RNA tethered to pRpp was incubated with 4S Ura. Active sequences react to form tethered 4-thiouridine. ( B ) Active sequences were purified away from unreactive sequences, using an APM gel. ( C ) Using T4 RNA ligase, purified ribozymes were then ligated to an adenylated DNA oligo containing an Ear I restriction site and 3′ RT-PCR primer binding region. ( D ) RT-PCR was performed to amplify ribozyme DNA sequence and add a T7 promoter (T7) to the 5′ end of the sequences. ( E ) Digestion with Ear I-generated DNA sequence lacking a fixed 3′ primer-binding site, which was then transcribed to generate RNA for another round of selection.
Figure Legend Snippet: Selection scheme designed to retain sequence variation all the way to the 3′ end of RNA transcripts. ( A ) Pool RNA tethered to pRpp was incubated with 4S Ura. Active sequences react to form tethered 4-thiouridine. ( B ) Active sequences were purified away from unreactive sequences, using an APM gel. ( C ) Using T4 RNA ligase, purified ribozymes were then ligated to an adenylated DNA oligo containing an Ear I restriction site and 3′ RT-PCR primer binding region. ( D ) RT-PCR was performed to amplify ribozyme DNA sequence and add a T7 promoter (T7) to the 5′ end of the sequences. ( E ) Digestion with Ear I-generated DNA sequence lacking a fixed 3′ primer-binding site, which was then transcribed to generate RNA for another round of selection.

Techniques Used: Selection, Sequencing, Incubation, Purification, Reverse Transcription Polymerase Chain Reaction, Binding Assay, Generated

4) Product Images from "Physical and Functional Interactions of the Arf Tumor Suppressor Protein with Nucleophosmin/B23"

Article Title: Physical and Functional Interactions of the Arf Tumor Suppressor Protein with Nucleophosmin/B23

Journal: Molecular and Cellular Biology

doi: 10.1128/MCB.24.3.985-996.2004

Arf forms complexes with NPM. (A, upper two panels) Lysates prepared from NIH 3T3 and MT-Arf cells treated with zinc sulfate for 24 h (+) or not (−) were precipitated with antibodies to NPM (lanes 5 to 8), p19 Arf (lanes 9 to 12), or control IgG (lanes 13 and 14). Denatured immune complexes electrophoretically separated on gels were transferred to a membrane and blotted with the same antibodies. Cell lysates (10%) separated on denaturing gels were directly blotted with antibodies to NPM and p19 Arf in order to estimate levels of the expressed proteins (lanes 1 to 4; see text). (A, lower panel) RNAs eluted from immune complexes were labeled with [ 32 P]pCp and T4 RNA ligase, separated on a denaturing gel, and detected by autoradiography. (B) Primary MEFs established from day 15 embryos were propagated on a 3T3 protocol for seven passages. Cell lysates expressing both p19 Arf and NPM (lane 1) were precipitated with control antibodies (lane 2) or with antibodies to NPM or Arf (lanes 3 and 4). Proteins separated on denaturing gels were immunoblotted with the same antibodies, indicated at the right of the panel. (C) Arf/Mdm2/p53 triple knockout (TKO) MEFs infected with retroviruses encoding p19 Arf or the p19 Arf Δ2-14 mutant were lysed and precipitated with antibodies to p19 Arf (lanes 4 to 6) or nonimmune rabbit serum (NRS) (lanes 7 to 9) and subjected to immunoblotting with antibodies to Arf (top panel) or NPM (bottom panel). A portion of the cell lysate was used to demonstrate levels of protein expression (lanes 1 to 3) as in panel A.
Figure Legend Snippet: Arf forms complexes with NPM. (A, upper two panels) Lysates prepared from NIH 3T3 and MT-Arf cells treated with zinc sulfate for 24 h (+) or not (−) were precipitated with antibodies to NPM (lanes 5 to 8), p19 Arf (lanes 9 to 12), or control IgG (lanes 13 and 14). Denatured immune complexes electrophoretically separated on gels were transferred to a membrane and blotted with the same antibodies. Cell lysates (10%) separated on denaturing gels were directly blotted with antibodies to NPM and p19 Arf in order to estimate levels of the expressed proteins (lanes 1 to 4; see text). (A, lower panel) RNAs eluted from immune complexes were labeled with [ 32 P]pCp and T4 RNA ligase, separated on a denaturing gel, and detected by autoradiography. (B) Primary MEFs established from day 15 embryos were propagated on a 3T3 protocol for seven passages. Cell lysates expressing both p19 Arf and NPM (lane 1) were precipitated with control antibodies (lane 2) or with antibodies to NPM or Arf (lanes 3 and 4). Proteins separated on denaturing gels were immunoblotted with the same antibodies, indicated at the right of the panel. (C) Arf/Mdm2/p53 triple knockout (TKO) MEFs infected with retroviruses encoding p19 Arf or the p19 Arf Δ2-14 mutant were lysed and precipitated with antibodies to p19 Arf (lanes 4 to 6) or nonimmune rabbit serum (NRS) (lanes 7 to 9) and subjected to immunoblotting with antibodies to Arf (top panel) or NPM (bottom panel). A portion of the cell lysate was used to demonstrate levels of protein expression (lanes 1 to 3) as in panel A.

Techniques Used: Labeling, Autoradiography, Expressing, Triple Knockout, Infection, Mutagenesis

5) Product Images from "RNA-Dependent DNA Binding Activity of the Pur Factor, Potentially Involved in DNA Replication and Gene Transcription"

Article Title: RNA-Dependent DNA Binding Activity of the Pur Factor, Potentially Involved in DNA Replication and Gene Transcription

Journal: Gene Expression

doi:

RNAs interacting with Pur are present in rabbit retic-ulocyte lysates. (A) Sensitivity to RNase A of the PUR binding activity of the in vitro translated Purα protein. The ability of an in vitro-translated Purα protein to bind 32 P-labeled PUR oligonucleotides was examined by mobility shift assay. Binding reactions were done with the reticulocyte lysate immediately after the translation reaction (lane 1), or after an additional 30-min incubation at 37°C in the absence (lane 2) or presence (lane 3) of RNase A. (B) Presence in the rabbit reticulocyte lysate of RNAs similar to the Pur-associated RNAs of fraction 14 of RSV-infected QEF nuclear extracts. Total RNAs were prepared from fraction 14 of nuclear extracts from RSV-infected QEF (lane 1), or the rabbit reticulocyte lysate (lane 2), and 3′ end labeled using the T4 RNA polymerase, and run in parallel in a denaturing 20% polyacrylamide gel, along with 22, 27, and 32 mer radioactive oligonucleotides as molecular weight markers. Identical 28- and 29-bases-long RNAs, similar to those previously found associated with the PUR complexes, are present in both preparations, and are indicated by arrows.
Figure Legend Snippet: RNAs interacting with Pur are present in rabbit retic-ulocyte lysates. (A) Sensitivity to RNase A of the PUR binding activity of the in vitro translated Purα protein. The ability of an in vitro-translated Purα protein to bind 32 P-labeled PUR oligonucleotides was examined by mobility shift assay. Binding reactions were done with the reticulocyte lysate immediately after the translation reaction (lane 1), or after an additional 30-min incubation at 37°C in the absence (lane 2) or presence (lane 3) of RNase A. (B) Presence in the rabbit reticulocyte lysate of RNAs similar to the Pur-associated RNAs of fraction 14 of RSV-infected QEF nuclear extracts. Total RNAs were prepared from fraction 14 of nuclear extracts from RSV-infected QEF (lane 1), or the rabbit reticulocyte lysate (lane 2), and 3′ end labeled using the T4 RNA polymerase, and run in parallel in a denaturing 20% polyacrylamide gel, along with 22, 27, and 32 mer radioactive oligonucleotides as molecular weight markers. Identical 28- and 29-bases-long RNAs, similar to those previously found associated with the PUR complexes, are present in both preparations, and are indicated by arrows.

Techniques Used: Binding Assay, Activity Assay, In Vitro, Labeling, Mobility Shift, Incubation, Infection, Molecular Weight

Related Articles

Mutagenesis:

Article Title: Efficient gene replacement and direct hyphal transformation in Sclerotinia sclerotiorum
Article Snippet: .. For Southern blot analysis, total genomic DNA was extracted from ssku80 mutant strain 20 and the wild type, digested with either Bam HI or Eco RV and transferred to a Hybond‐XL (Amersham Biosciences, UK) nylon membrane. .. Hybridization was performed at 42 °C under low‐stringency conditions in the presence of ULTRAhyb solution (Ambion, Austin, TX).

Electrophoresis:

Article Title: Regulation of RNA polymerase III transcription during transformation of human IMR90 fibroblasts with defined genetic elements
Article Snippet: .. 8 µg of total RNA was separated by electrophoresis and blotted onto Hybond-XL nylon membrane (GE Healthcare, RPN2020S). .. The membrane was incubated in 1x Church buffer (0.25 M NaH2 PO4 ; 1 mM EDTA; 7% SDS; 20 mg/ml salmon sperm DNA; 0.5% BSA) for 2 hrs at 28°C and the denatured radioactive probe was added for incubation overnight.

Agarose Gel Electrophoresis:

Article Title: Modeling cancer genomic data in yeast reveals selection against ATM function during tumorigenesis
Article Snippet: .. Genomic DNA was either Pst I-or Xho I-digested (as specified in Figure legends), run on 1.3% agarose gel and transferred on an Amersham Hybond-XL membrane (GE Healthcare) and detected by Southern blot with a 32 P-labeled telomere-specific probe (5′-TGTGGTGTGTGGGTGTGGTGT-3′) as described [ ]. .. Rad50 overexpression and purification from yeast cells The yeast galactose inducible expression plasmid, pR50.1 (2μ, GAL-PGK-RAD50, leu2-d), was a gift from Patrick Sung [ ].

Article Title: rahu is a mutant allele of Dnmt3c, encoding a DNA methyltransferase homolog required for meiosis and transposon repression in the mouse male germline
Article Snippet: .. ~250 ng of digested DNA was electrophoresed on a 0.9% agarose gel and transferred as described [ ] to an Amersham Hybond-XL membrane (GE Healthcare). .. The L1 5′UTR probe has been described elsewhere [ ] and corresponds to nucleotides 515–1628 of the L1 sequence (GenBank accession number M13002).

Southern Blot:

Article Title: Efficient gene replacement and direct hyphal transformation in Sclerotinia sclerotiorum
Article Snippet: .. For Southern blot analysis, total genomic DNA was extracted from ssku80 mutant strain 20 and the wild type, digested with either Bam HI or Eco RV and transferred to a Hybond‐XL (Amersham Biosciences, UK) nylon membrane. .. Hybridization was performed at 42 °C under low‐stringency conditions in the presence of ULTRAhyb solution (Ambion, Austin, TX).

Article Title: Preferential Repair of the Transcribed DNA Strand in Plants
Article Snippet: .. Samples (at least 1 μg of DNA/lane) were electrophoresed on a 0.5% alkaline gel (9.5 cm) in 1 mM EDTA and 30 mM NaOH at 22 V for around 16 h. Southern blotting, hybridization and washes were performed as described (Spivak and Hanawalt Philip, ) with the exception that Hybond-N+ or Hybond-XL nylon membranes (Amersham Pharmacia, UK) were used. ..

Article Title: Modeling cancer genomic data in yeast reveals selection against ATM function during tumorigenesis
Article Snippet: .. Genomic DNA was either Pst I-or Xho I-digested (as specified in Figure legends), run on 1.3% agarose gel and transferred on an Amersham Hybond-XL membrane (GE Healthcare) and detected by Southern blot with a 32 P-labeled telomere-specific probe (5′-TGTGGTGTGTGGGTGTGGTGT-3′) as described [ ]. .. Rad50 overexpression and purification from yeast cells The yeast galactose inducible expression plasmid, pR50.1 (2μ, GAL-PGK-RAD50, leu2-d), was a gift from Patrick Sung [ ].

Hybridization:

Article Title: Preferential Repair of the Transcribed DNA Strand in Plants
Article Snippet: .. Samples (at least 1 μg of DNA/lane) were electrophoresed on a 0.5% alkaline gel (9.5 cm) in 1 mM EDTA and 30 mM NaOH at 22 V for around 16 h. Southern blotting, hybridization and washes were performed as described (Spivak and Hanawalt Philip, ) with the exception that Hybond-N+ or Hybond-XL nylon membranes (Amersham Pharmacia, UK) were used. ..

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    GE Healthcare t4 rna ligase
    RNA-DNA ligation with <t>T4</t> RNA ligase. ( A ) The 21- to 24-nt smRNAs from infected HC-Pro + or HC-Pro - plants (lanes 5-12) or the 24-nt smRNAs from both infected and uninfected HC-Pro + and HC-Pro - plants (lanes 13-28) were gel-purified and ligated to a 16-nt
    T4 Rna Ligase, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 92/100, based on 27 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/t4 rna ligase/product/GE Healthcare
    Average 92 stars, based on 27 article reviews
    Price from $9.99 to $1999.99
    t4 rna ligase - by Bioz Stars, 2020-09
    92/100 stars
      Buy from Supplier

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    RNA-DNA ligation with T4 RNA ligase. ( A ) The 21- to 24-nt smRNAs from infected HC-Pro + or HC-Pro - plants (lanes 5-12) or the 24-nt smRNAs from both infected and uninfected HC-Pro + and HC-Pro - plants (lanes 13-28) were gel-purified and ligated to a 16-nt

    Journal:

    Article Title: Extensive 3? modification of plant small RNAs is modulated by helper component-proteinase expression

    doi: 10.1073/pnas.0506597102

    Figure Lengend Snippet: RNA-DNA ligation with T4 RNA ligase. ( A ) The 21- to 24-nt smRNAs from infected HC-Pro + or HC-Pro - plants (lanes 5-12) or the 24-nt smRNAs from both infected and uninfected HC-Pro + and HC-Pro - plants (lanes 13-28) were gel-purified and ligated to a 16-nt

    Article Snippet: Radiolabeled 21- to 24-nt smRNAs from virus-infected HC-Pro+ or HC-Pro- plants, together with the four gel-purified samples of 24-nt smRNAs (the same as used in ), were tested for their ability to serve as substrates for T4 RNA ligase.

    Techniques: DNA Ligation, Infection, Purification