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Boehringer Mannheim dnaase i
Dnaase I, supplied by Boehringer Mannheim, used in various techniques. Bioz Stars score: 86/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/dnaase i/product/Boehringer Mannheim
Average 86 stars, based on 3 article reviews
Price from $9.99 to $1999.99
dnaase i - by Bioz Stars, 2020-04
86/100 stars

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Related Articles

Nucleic Acid Electrophoresis:

Article Title: Characterization of Novel Parent-Specific Epigenetic Modifications Upstream of the Imprinted Mouse H19 Gene
Article Snippet: .. The cells were immediately pelleted, then resuspended in solution II containing DNase I (1 μg/ml; Boehringer Mannheim catalog no. 104132), and incubated for 1 to 6 min at RT to obtain an optimal nicking frequency of one nick every 200 to 800 nucleotides, as assessed by alkaline gel electrophoresis after DNA purification and before LMPCR. .. Cells were pelleted and resuspended in 2 ml of buffer C containing proteinase K (600 μg/ml), then 2 ml of buffer B was added, and the sample was incubated at 37°C overnight ( ).

Amplification:

Article Title: Repressor CopG prevents access of RNA polymerase to promoter and actively dissociates open complexes
Article Snippet: The amplified fragment spanned from −144 to +95 relative to the Pcr transcription start site ( ). .. DNase I and E. coli RNAP holoenzyme were from Boehringer Mannheim.

Article Title: Temporal Regulation of Genes Encoding the Flagellar Proximal Rod in Caulobacter crescentus
Article Snippet: Products were amplified with Pfu polymerase (Stratagene) according to the manufacturer's directions, cloned directly into pET15b, and verified by dye termination sequencing. .. The mix (volume of 190 μl) was allowed to bind for 20 min at room temperature, and then 0.05 U of DNase I (Boehringer Mannheim) was added and allowed to digest for 3 min. At that time, 100 μl of DNase stop buffer (0.6 M ammonium acetate, 150 mM EDTA, 20 mg of calf thymus DNA per ml) was added to stop the reactions, and the reaction mixtures were then extracted once with phenol-chloroform-isoamyl alcohol (25:24:1; equilibrated with Tris-EDTA, pH 8.0), precipitated with 3 volumes of ethanol, washed with 70% ethanol, dried, and resuspended in 12 μl of formamide loading buffer.

Clone Assay:

Article Title: Temporal Regulation of Genes Encoding the Flagellar Proximal Rod in Caulobacter crescentus
Article Snippet: Products were amplified with Pfu polymerase (Stratagene) according to the manufacturer's directions, cloned directly into pET15b, and verified by dye termination sequencing. .. The mix (volume of 190 μl) was allowed to bind for 20 min at room temperature, and then 0.05 U of DNase I (Boehringer Mannheim) was added and allowed to digest for 3 min. At that time, 100 μl of DNase stop buffer (0.6 M ammonium acetate, 150 mM EDTA, 20 mg of calf thymus DNA per ml) was added to stop the reactions, and the reaction mixtures were then extracted once with phenol-chloroform-isoamyl alcohol (25:24:1; equilibrated with Tris-EDTA, pH 8.0), precipitated with 3 volumes of ethanol, washed with 70% ethanol, dried, and resuspended in 12 μl of formamide loading buffer.

In Vitro:

Article Title: Chromatin Degradation in Differentiating Fiber Cells of the Eye Lens
Article Snippet: .. To control for this, some slices were preincubated in DNase I to cause extensive DNA fragmentation in vitro. .. In DNase I–treated tissue, all of the nuclei were labeled by the TdT assay, including those of the epithelial cells and annular pad (data not shown) and superficial fiber cells (Fig. B ).

Methylation:

Article Title: CpG methylation of an endogenous retroviral enhancer inhibits transcription factor binding and activity
Article Snippet: Paragraph title: Band shift, DNAse I footprinting, methylation interference, and UV crosslinking assays ... The probe and extracts were incubated as in the band-shift assays and digested with DNAse I (20 μg/ml; Boehringer Mannheim) for 30 seconds.

Isolation:

Article Title: Identification and reconstitution of the origin recognition complex from Schizosaccharomyces pombe
Article Snippet: .. The isolated chromatin was incubated with 0.3 ml of 0.1 M NaCl in buffer H containing DNase I (1,000 units/ml) at 4°C for 30 min to extract the ORC proteins and then centrifuged for 15 min at 15,000 × g at 4°C. .. HA or FLAG cross-linked agarose beads (15 μl; Sigma) were added to the supernatant, and the mixture was incubated for 2 hr at 4°C with shaking.

Article Title: Characterization of Novel Parent-Specific Epigenetic Modifications Upstream of the Imprinted Mouse H19 Gene
Article Snippet: The cells were immediately pelleted, then resuspended in solution II containing DNase I (1 μg/ml; Boehringer Mannheim catalog no. 104132), and incubated for 1 to 6 min at RT to obtain an optimal nicking frequency of one nick every 200 to 800 nucleotides, as assessed by alkaline gel electrophoresis after DNA purification and before LMPCR. .. As a control, 40 μg of genomic DNA isolated from untreated ES cells as described above was mixed with 10 μg of DNase I per ml in a 200-μl reaction volume and incubated at RT for 5 min.

Footprinting:

Article Title: CpG methylation of an endogenous retroviral enhancer inhibits transcription factor binding and activity
Article Snippet: Paragraph title: Band shift, DNAse I footprinting, methylation interference, and UV crosslinking assays ... The probe and extracts were incubated as in the band-shift assays and digested with DNAse I (20 μg/ml; Boehringer Mannheim) for 30 seconds.

Article Title: Cooperative Interaction of CI Protein Regulates Lysogeny of Lactobacillus casei by Bacteriophage A2
Article Snippet: Paragraph title: DNase I footprinting. ... End-labeled DNA (81 pM) was incubated for 30 min at 30°C in the absence or presence of CI and/or B. subtilis ςA -RNAP in 20 μl of buffer B plus poly(dI)-poly(dC) as an unspecific competitor (at a final concentration of 50 ng/μl), followed by the addition of freshly diluted DNase I (Boehringer Mannheim) (at a concentration to obtain, on average, one cut per molecule).

Article Title: Temporal Regulation of Genes Encoding the Flagellar Proximal Rod in Caulobacter crescentus
Article Snippet: Paragraph title: CtrA footprinting assays. ... The mix (volume of 190 μl) was allowed to bind for 20 min at room temperature, and then 0.05 U of DNase I (Boehringer Mannheim) was added and allowed to digest for 3 min. At that time, 100 μl of DNase stop buffer (0.6 M ammonium acetate, 150 mM EDTA, 20 mg of calf thymus DNA per ml) was added to stop the reactions, and the reaction mixtures were then extracted once with phenol-chloroform-isoamyl alcohol (25:24:1; equilibrated with Tris-EDTA, pH 8.0), precipitated with 3 volumes of ethanol, washed with 70% ethanol, dried, and resuspended in 12 μl of formamide loading buffer.

Incubation:

Article Title: Identification and reconstitution of the origin recognition complex from Schizosaccharomyces pombe
Article Snippet: .. The isolated chromatin was incubated with 0.3 ml of 0.1 M NaCl in buffer H containing DNase I (1,000 units/ml) at 4°C for 30 min to extract the ORC proteins and then centrifuged for 15 min at 15,000 × g at 4°C. .. HA or FLAG cross-linked agarose beads (15 μl; Sigma) were added to the supernatant, and the mixture was incubated for 2 hr at 4°C with shaking.

Article Title: CpG methylation of an endogenous retroviral enhancer inhibits transcription factor binding and activity
Article Snippet: .. The probe and extracts were incubated as in the band-shift assays and digested with DNAse I (20 μg/ml; Boehringer Mannheim) for 30 seconds. .. The products were run on a 6% sequencing gel, and the position of the footprint was determined by running the A + G Maxam-Gilbert sequencing reaction ( , pp.

Article Title: Cooperative Interaction of CI Protein Regulates Lysogeny of Lactobacillus casei by Bacteriophage A2
Article Snippet: .. End-labeled DNA (81 pM) was incubated for 30 min at 30°C in the absence or presence of CI and/or B. subtilis ςA -RNAP in 20 μl of buffer B plus poly(dI)-poly(dC) as an unspecific competitor (at a final concentration of 50 ng/μl), followed by the addition of freshly diluted DNase I (Boehringer Mannheim) (at a concentration to obtain, on average, one cut per molecule). .. The mixture was incubated at 30°C for 3 min, and the digestion was stopped by the addition of 1 μl of 0.5 M EDTA, pH 8.

Article Title: Characterization of Novel Parent-Specific Epigenetic Modifications Upstream of the Imprinted Mouse H19 Gene
Article Snippet: .. The cells were immediately pelleted, then resuspended in solution II containing DNase I (1 μg/ml; Boehringer Mannheim catalog no. 104132), and incubated for 1 to 6 min at RT to obtain an optimal nicking frequency of one nick every 200 to 800 nucleotides, as assessed by alkaline gel electrophoresis after DNA purification and before LMPCR. .. Cells were pelleted and resuspended in 2 ml of buffer C containing proteinase K (600 μg/ml), then 2 ml of buffer B was added, and the sample was incubated at 37°C overnight ( ).

Article Title: Temporal Regulation of Genes Encoding the Flagellar Proximal Rod in Caulobacter crescentus
Article Snippet: CtrA-His6 (4.8 mM) and EnvZ′-MalE (0.5 mM) were added to phosphorylation buffer (50 mM Tris [pH 7.5], 50 mM KCl, 20 mM MgCl2 , 1 mM dithiothreitol, 0.5 mM ATP) and incubated at 37°C for 1 h ( ). .. The mix (volume of 190 μl) was allowed to bind for 20 min at room temperature, and then 0.05 U of DNase I (Boehringer Mannheim) was added and allowed to digest for 3 min. At that time, 100 μl of DNase stop buffer (0.6 M ammonium acetate, 150 mM EDTA, 20 mg of calf thymus DNA per ml) was added to stop the reactions, and the reaction mixtures were then extracted once with phenol-chloroform-isoamyl alcohol (25:24:1; equilibrated with Tris-EDTA, pH 8.0), precipitated with 3 volumes of ethanol, washed with 70% ethanol, dried, and resuspended in 12 μl of formamide loading buffer.

Electrophoretic Mobility Shift Assay:

Article Title: CpG methylation of an endogenous retroviral enhancer inhibits transcription factor binding and activity
Article Snippet: .. The probe and extracts were incubated as in the band-shift assays and digested with DNAse I (20 μg/ml; Boehringer Mannheim) for 30 seconds. .. The products were run on a 6% sequencing gel, and the position of the footprint was determined by running the A + G Maxam-Gilbert sequencing reaction ( , pp.

Purification:

Article Title: Temporal Regulation of Genes Encoding the Flagellar Proximal Rod in Caulobacter crescentus
Article Snippet: EnvZ′-MalE fusion protein from pKJH5 ( , ) was purified as described in the pMAL protein fusion manual (New England Biolabs). .. The mix (volume of 190 μl) was allowed to bind for 20 min at room temperature, and then 0.05 U of DNase I (Boehringer Mannheim) was added and allowed to digest for 3 min. At that time, 100 μl of DNase stop buffer (0.6 M ammonium acetate, 150 mM EDTA, 20 mg of calf thymus DNA per ml) was added to stop the reactions, and the reaction mixtures were then extracted once with phenol-chloroform-isoamyl alcohol (25:24:1; equilibrated with Tris-EDTA, pH 8.0), precipitated with 3 volumes of ethanol, washed with 70% ethanol, dried, and resuspended in 12 μl of formamide loading buffer.

Sequencing:

Article Title: CpG methylation of an endogenous retroviral enhancer inhibits transcription factor binding and activity
Article Snippet: The Sp1 oligonucleotide (a gift from Dr. Roberto Weinmann, The Wistar Institute) contains the sequence 5′-TCCCGCCCCTAACTCCGCCCAT-3′ from the human metallothionein-IIA promoter. .. The probe and extracts were incubated as in the band-shift assays and digested with DNAse I (20 μg/ml; Boehringer Mannheim) for 30 seconds.

Article Title: Cooperative Interaction of CI Protein Regulates Lysogeny of Lactobacillus casei by Bacteriophage A2
Article Snippet: End-labeled DNA (81 pM) was incubated for 30 min at 30°C in the absence or presence of CI and/or B. subtilis ςA -RNAP in 20 μl of buffer B plus poly(dI)-poly(dC) as an unspecific competitor (at a final concentration of 50 ng/μl), followed by the addition of freshly diluted DNase I (Boehringer Mannheim) (at a concentration to obtain, on average, one cut per molecule). .. Chemical sequencing reactions ( ) for purines, obtained by an express protocol , were run in parallel to determine the sizes of the DNA fragments generated.

Article Title: Temporal Regulation of Genes Encoding the Flagellar Proximal Rod in Caulobacter crescentus
Article Snippet: Products were amplified with Pfu polymerase (Stratagene) according to the manufacturer's directions, cloned directly into pET15b, and verified by dye termination sequencing. .. The mix (volume of 190 μl) was allowed to bind for 20 min at room temperature, and then 0.05 U of DNase I (Boehringer Mannheim) was added and allowed to digest for 3 min. At that time, 100 μl of DNase stop buffer (0.6 M ammonium acetate, 150 mM EDTA, 20 mg of calf thymus DNA per ml) was added to stop the reactions, and the reaction mixtures were then extracted once with phenol-chloroform-isoamyl alcohol (25:24:1; equilibrated with Tris-EDTA, pH 8.0), precipitated with 3 volumes of ethanol, washed with 70% ethanol, dried, and resuspended in 12 μl of formamide loading buffer.

Concentration Assay:

Article Title: Cooperative Interaction of CI Protein Regulates Lysogeny of Lactobacillus casei by Bacteriophage A2
Article Snippet: .. End-labeled DNA (81 pM) was incubated for 30 min at 30°C in the absence or presence of CI and/or B. subtilis ςA -RNAP in 20 μl of buffer B plus poly(dI)-poly(dC) as an unspecific competitor (at a final concentration of 50 ng/μl), followed by the addition of freshly diluted DNase I (Boehringer Mannheim) (at a concentration to obtain, on average, one cut per molecule). .. The mixture was incubated at 30°C for 3 min, and the digestion was stopped by the addition of 1 μl of 0.5 M EDTA, pH 8.

Article Title: Repressor CopG prevents access of RNA polymerase to promoter and actively dissociates open complexes
Article Snippet: DNase I and E. coli RNAP holoenzyme were from Boehringer Mannheim. .. Promoter binding activity of the commercial preparations of RNAP was determined by EMSA, using DNA concentrations 10, 20 or 40 times the total concentration of RNAP (4 nM).

Generated:

Article Title: Cooperative Interaction of CI Protein Regulates Lysogeny of Lactobacillus casei by Bacteriophage A2
Article Snippet: End-labeled DNA (81 pM) was incubated for 30 min at 30°C in the absence or presence of CI and/or B. subtilis ςA -RNAP in 20 μl of buffer B plus poly(dI)-poly(dC) as an unspecific competitor (at a final concentration of 50 ng/μl), followed by the addition of freshly diluted DNase I (Boehringer Mannheim) (at a concentration to obtain, on average, one cut per molecule). .. Chemical sequencing reactions ( ) for purines, obtained by an express protocol , were run in parallel to determine the sizes of the DNA fragments generated.

Article Title: Repressor CopG prevents access of RNA polymerase to promoter and actively dissociates open complexes
Article Snippet: DNA, proteins and chemicals A 239-bp DNA fragment containing Pcr and the overlapping CopG target was generated by PCR using Pfu DNA polymerase (Stratagene), a pair of synthetic primers (5′-CGCCTTTAGCCTTAGAG-3′ and 5′-CCATCTCTCTTGCCAT-3′), and pMV158 DNA as template. .. DNase I and E. coli RNAP holoenzyme were from Boehringer Mannheim.

other:

Article Title: Two Widely Spaced Initiator Binding Sites Create an HMG1-Dependent Parvovirus Rolling-Hairpin Replication Origin
Article Snippet: Surprisingly, MPE-Fe(II) footprints show that NS1 makes extremely close contact with the DNA throughout much of the region protected from DNase I.

Article Title: Leishmania infantum EndoG Is an Endo/Exo-Nuclease Essential for Parasite Survival
Article Snippet: By contrast, the peak pattern observed after digestion with DNase I is much more dispersed along the x axis and the smallest fragments are not the most abundant ( ).

Article Title: Concerted formation of macromolecular Suppressor–mutator transposition complexes
Article Snippet: The reaction was terminated by addition of 700 μl of DNase I stop solution (645 μl of 100% ethanol/5 μg of tRNA/50 μl of saturated ammonium acetate) and chilled in a dry ice/ethanol bath.

Activity Assay:

Article Title: Chromatin Degradation in Differentiating Fiber Cells of the Eye Lens
Article Snippet: .. This activity may correspond to a DNase I–like, 30-kD protein identified in lens tissue by DNase activity gels and immunoblots ( ; ; ). ..

Article Title: Repressor CopG prevents access of RNA polymerase to promoter and actively dissociates open complexes
Article Snippet: DNase I and E. coli RNAP holoenzyme were from Boehringer Mannheim. .. Promoter binding activity of the commercial preparations of RNAP was determined by EMSA, using DNA concentrations 10, 20 or 40 times the total concentration of RNAP (4 nM).

Labeling:

Article Title: Cooperative Interaction of CI Protein Regulates Lysogeny of Lactobacillus casei by Bacteriophage A2
Article Snippet: The Eco RI- Sph I or Hin dIII- Kpn I DNA fragments obtained from pUO183 were end labeled at the Eco RI or Hin dIII sites with Klenow DNA polymerase and [α-32 P]dATP (3,000 Ci/mmol; Amersham). .. End-labeled DNA (81 pM) was incubated for 30 min at 30°C in the absence or presence of CI and/or B. subtilis ςA -RNAP in 20 μl of buffer B plus poly(dI)-poly(dC) as an unspecific competitor (at a final concentration of 50 ng/μl), followed by the addition of freshly diluted DNase I (Boehringer Mannheim) (at a concentration to obtain, on average, one cut per molecule).

DNA Purification:

Article Title: Characterization of Novel Parent-Specific Epigenetic Modifications Upstream of the Imprinted Mouse H19 Gene
Article Snippet: .. The cells were immediately pelleted, then resuspended in solution II containing DNase I (1 μg/ml; Boehringer Mannheim catalog no. 104132), and incubated for 1 to 6 min at RT to obtain an optimal nicking frequency of one nick every 200 to 800 nucleotides, as assessed by alkaline gel electrophoresis after DNA purification and before LMPCR. .. Cells were pelleted and resuspended in 2 ml of buffer C containing proteinase K (600 μg/ml), then 2 ml of buffer B was added, and the sample was incubated at 37°C overnight ( ).

End Labeling:

Article Title: CpG methylation of an endogenous retroviral enhancer inhibits transcription factor binding and activity
Article Snippet: DNAse I footprinting was carried out by end-labeling the −307/−67 restriction fragment from pIAP.l by filling recessed 3′ termini as described ( , pp. .. The probe and extracts were incubated as in the band-shift assays and digested with DNAse I (20 μg/ml; Boehringer Mannheim) for 30 seconds.

Polymerase Chain Reaction:

Article Title: Repressor CopG prevents access of RNA polymerase to promoter and actively dissociates open complexes
Article Snippet: The amplified fragment spanned from −144 to +95 relative to the Pcr transcription start site ( ). .. DNase I and E. coli RNAP holoenzyme were from Boehringer Mannheim.

Article Title: Temporal Regulation of Genes Encoding the Flagellar Proximal Rod in Caulobacter crescentus
Article Snippet: PCR primers used to amplify ctrA for cloning into pET15b were 5′-ATGCAGCATATGCGCGTACTGTTGATCGAG and 3′-ATGCAGCGCGAGTCAGGCGGCGTTAACCTGCT. .. The mix (volume of 190 μl) was allowed to bind for 20 min at room temperature, and then 0.05 U of DNase I (Boehringer Mannheim) was added and allowed to digest for 3 min. At that time, 100 μl of DNase stop buffer (0.6 M ammonium acetate, 150 mM EDTA, 20 mg of calf thymus DNA per ml) was added to stop the reactions, and the reaction mixtures were then extracted once with phenol-chloroform-isoamyl alcohol (25:24:1; equilibrated with Tris-EDTA, pH 8.0), precipitated with 3 volumes of ethanol, washed with 70% ethanol, dried, and resuspended in 12 μl of formamide loading buffer.

Western Blot:

Article Title: Chromatin Degradation in Differentiating Fiber Cells of the Eye Lens
Article Snippet: .. This activity may correspond to a DNase I–like, 30-kD protein identified in lens tissue by DNase activity gels and immunoblots ( ; ; ). ..

Binding Assay:

Article Title: Activation Domain-Specific and General Transcription Stimulation by Native Histone Acetyltransferase Complexes
Article Snippet: .. Binding reaction mixtures with GAL4 derivatives (Fig. D) were treated with DNase I for 0.5 min, except GAL4-Gln, which was treated for 1.5 min. DNase I was terminated with stop mix ( ). .. After ethanol precipitation, DNA was dissolved in 10 mM Tris-HCl (pH 7.5)–1 mM EDTA–50 mM NaCl and electrophoresed through 1.5% 1× Tris-borate-EDTA–agarose gel.

Article Title: Repressor CopG prevents access of RNA polymerase to promoter and actively dissociates open complexes
Article Snippet: DNase I and E. coli RNAP holoenzyme were from Boehringer Mannheim. .. Promoter binding activity of the commercial preparations of RNAP was determined by EMSA, using DNA concentrations 10, 20 or 40 times the total concentration of RNAP (4 nM).

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    Boehringer Mannheim dnase i
    Exonuclease activity of rLiEndoG. 500′-labeled with FAM were digested with rLiEndoG or DNase I. The amount of enzyme used for digestion is indicated. Digestion products were processed by both agarose gel (1%) and capillary electrophoresis.  A ) Agarose gel of the DNA fragments generated after 1 h of digestion with rLiEndoG or DNase I.  B ) Capillary electrophoresis of the DNA probe digested for 1 h with 0.1 µg of rLiEndoG.  C ) Capillary electrophoresis results obtained for the DNA probe digested for 1 hour with 0.01 units of DNase I. Digested DNA was heat-denatured prior to capillary electrophoresis. Fluorescence intensities (arbitrary units) of the single-stranded DNA fragments generated after digestion and denaturation are shown on the y axis. Sizes of the ssDNA fragments (in nucleotides) are shown on the x axis. Fragment sizes were analyzed with the Peak Scanner (Applied Biosystems) software. Accurate sizes can only be predicted for fragments longer than 50 nucleotides.
    Dnase I, supplied by Boehringer Mannheim, used in various techniques. Bioz Stars score: 93/100, based on 98 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/dnase i/product/Boehringer Mannheim
    Average 93 stars, based on 98 article reviews
    Price from $9.99 to $1999.99
    dnase i - by Bioz Stars, 2020-04
    93/100 stars
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    Exonuclease activity of rLiEndoG. 500′-labeled with FAM were digested with rLiEndoG or DNase I. The amount of enzyme used for digestion is indicated. Digestion products were processed by both agarose gel (1%) and capillary electrophoresis.  A ) Agarose gel of the DNA fragments generated after 1 h of digestion with rLiEndoG or DNase I.  B ) Capillary electrophoresis of the DNA probe digested for 1 h with 0.1 µg of rLiEndoG.  C ) Capillary electrophoresis results obtained for the DNA probe digested for 1 hour with 0.01 units of DNase I. Digested DNA was heat-denatured prior to capillary electrophoresis. Fluorescence intensities (arbitrary units) of the single-stranded DNA fragments generated after digestion and denaturation are shown on the y axis. Sizes of the ssDNA fragments (in nucleotides) are shown on the x axis. Fragment sizes were analyzed with the Peak Scanner (Applied Biosystems) software. Accurate sizes can only be predicted for fragments longer than 50 nucleotides.

    Journal: PLoS ONE

    Article Title: Leishmania infantum EndoG Is an Endo/Exo-Nuclease Essential for Parasite Survival

    doi: 10.1371/journal.pone.0089526

    Figure Lengend Snippet: Exonuclease activity of rLiEndoG. 500′-labeled with FAM were digested with rLiEndoG or DNase I. The amount of enzyme used for digestion is indicated. Digestion products were processed by both agarose gel (1%) and capillary electrophoresis. A ) Agarose gel of the DNA fragments generated after 1 h of digestion with rLiEndoG or DNase I. B ) Capillary electrophoresis of the DNA probe digested for 1 h with 0.1 µg of rLiEndoG. C ) Capillary electrophoresis results obtained for the DNA probe digested for 1 hour with 0.01 units of DNase I. Digested DNA was heat-denatured prior to capillary electrophoresis. Fluorescence intensities (arbitrary units) of the single-stranded DNA fragments generated after digestion and denaturation are shown on the y axis. Sizes of the ssDNA fragments (in nucleotides) are shown on the x axis. Fragment sizes were analyzed with the Peak Scanner (Applied Biosystems) software. Accurate sizes can only be predicted for fragments longer than 50 nucleotides.

    Article Snippet: By contrast, the peak pattern observed after digestion with DNase I is much more dispersed along the x axis and the smallest fragments are not the most abundant ( ).

    Techniques: Activity Assay, Labeling, Agarose Gel Electrophoresis, Electrophoresis, Generated, Fluorescence, Software

    In situ electrophoresis of an E15 lens slice. ( A ) Low-magnification image of an ethidium-stained lens slice after electrophoresis. The slice is embedded in a block of 0.3% agarose. The dark space between the two bright arms ( arrowheads ) represents the OFZ. The arms are formed by the ethidium-stained fiber cell nuclei and are continuous with the band of annular pad and epithelial nuclei seen dimly in the background. The uneven bright strip extending across the image is a reflection from the edge of the agarose block. The arrow depicts the direction of the electrical field. ( B ) Intermediate-magnification view of a lens slice that was pretreated with DNase I before electrophoresis to cause fragmentation of fiber cell DNA. Note the diffuse clouds of ethidium-stained material emanating from all of the fiber cell nuclei. ( C ) Untreated lens slice showing two streams of ethidium-stained material emanating from nuclei immediately adjacent to the OFZ. ( D ) High-magnification image of cells at the border of the OFZ. Brightly stained individual nuclei are visible. A stream of ethidium-stained material can be seen issuing from cells immediately adjacent to the OFZ. Bars: ( A ) 500 μm; ( B ) 250 μm; ( C ) 100 μm; ( D ) 25 μm.

    Journal: The Journal of Cell Biology

    Article Title: Chromatin Degradation in Differentiating Fiber Cells of the Eye Lens

    doi:

    Figure Lengend Snippet: In situ electrophoresis of an E15 lens slice. ( A ) Low-magnification image of an ethidium-stained lens slice after electrophoresis. The slice is embedded in a block of 0.3% agarose. The dark space between the two bright arms ( arrowheads ) represents the OFZ. The arms are formed by the ethidium-stained fiber cell nuclei and are continuous with the band of annular pad and epithelial nuclei seen dimly in the background. The uneven bright strip extending across the image is a reflection from the edge of the agarose block. The arrow depicts the direction of the electrical field. ( B ) Intermediate-magnification view of a lens slice that was pretreated with DNase I before electrophoresis to cause fragmentation of fiber cell DNA. Note the diffuse clouds of ethidium-stained material emanating from all of the fiber cell nuclei. ( C ) Untreated lens slice showing two streams of ethidium-stained material emanating from nuclei immediately adjacent to the OFZ. ( D ) High-magnification image of cells at the border of the OFZ. Brightly stained individual nuclei are visible. A stream of ethidium-stained material can be seen issuing from cells immediately adjacent to the OFZ. Bars: ( A ) 500 μm; ( B ) 250 μm; ( C ) 100 μm; ( D ) 25 μm.

    Article Snippet: To control for this, some slices were preincubated in DNase I to cause extensive DNA fragmentation in vitro.

    Techniques: In Situ, Electrophoresis, Staining, Blocking Assay, Stripping Membranes

    Merged confocal and DIC images of lens slices after TdT labeling with fluorescein-dUTP. The DIC images are shown in green and positively labeled nuclei (containing fragmented DNA) are shown in red. ( A ) At the border of the OFZ, the nuclei lose their regular shape ( arrows ) and collapse into condensed structures that are strongly labeled by the TdT assay ( arrowheads ). Positively labeled debris, resulting presumably from the disintegration of labeled nuclei, extends deep into the OFZ. ( B ) Cortical fiber cells from a lens slice that was pretreated for 30 min with 50 U/ml DNase I. Note that after DNase I treatment, all nuclei are labeled by the TdT assay and that the labeling is strongest immediately beneath the nuclear membrane. ( C ) Equatorial region of a lens slice that had been incubated with CIAP before TdT labeling. None of the nuclei are labeled, indicating that the superficial fibers do not contain fragmented DNA with 3′PO 4 termini (see text for details). ( D ) Equatorial region of a lens slice that was treated sequentially with micrococcal nuclease and CIAP before TdT labeling. All the nuclei are labeled, demonstrating the efficacy of the CIAP technique for detecting fragmented DNA with 3′-PO 4 termini. Bars: ( A ) 50 μm; ( B ) 10 μm; ( C ) 50 μm; ( D ) 50 μm.

    Journal: The Journal of Cell Biology

    Article Title: Chromatin Degradation in Differentiating Fiber Cells of the Eye Lens

    doi:

    Figure Lengend Snippet: Merged confocal and DIC images of lens slices after TdT labeling with fluorescein-dUTP. The DIC images are shown in green and positively labeled nuclei (containing fragmented DNA) are shown in red. ( A ) At the border of the OFZ, the nuclei lose their regular shape ( arrows ) and collapse into condensed structures that are strongly labeled by the TdT assay ( arrowheads ). Positively labeled debris, resulting presumably from the disintegration of labeled nuclei, extends deep into the OFZ. ( B ) Cortical fiber cells from a lens slice that was pretreated for 30 min with 50 U/ml DNase I. Note that after DNase I treatment, all nuclei are labeled by the TdT assay and that the labeling is strongest immediately beneath the nuclear membrane. ( C ) Equatorial region of a lens slice that had been incubated with CIAP before TdT labeling. None of the nuclei are labeled, indicating that the superficial fibers do not contain fragmented DNA with 3′PO 4 termini (see text for details). ( D ) Equatorial region of a lens slice that was treated sequentially with micrococcal nuclease and CIAP before TdT labeling. All the nuclei are labeled, demonstrating the efficacy of the CIAP technique for detecting fragmented DNA with 3′-PO 4 termini. Bars: ( A ) 50 μm; ( B ) 10 μm; ( C ) 50 μm; ( D ) 50 μm.

    Article Snippet: To control for this, some slices were preincubated in DNase I to cause extensive DNA fragmentation in vitro.

    Techniques: Labeling, Incubation

    Summary of the analysis of the far upstream sequences. (A) Maternal chromosome, PG ES cells; (B) paternal chromosome, AG ES cells. Arrows pointing up or down indicate increased or decreased DNase I sensitivity of the in vivo-treated versus the in vitro-treated samples; shorter arrows indicate lower DNase I sensitivity in AG cells compared to PG cells; filled or open squares indicate increases or decreases, respectively, in DMS reactivity; open or closed circles indicate hypomethylated or hypermethylated CpGs, respectively; grey circles stand for partially methylated sites; brackets indicate the sequences analyzed.

    Journal: Molecular and Cellular Biology

    Article Title: Characterization of Novel Parent-Specific Epigenetic Modifications Upstream of the Imprinted Mouse H19 Gene

    doi:

    Figure Lengend Snippet: Summary of the analysis of the far upstream sequences. (A) Maternal chromosome, PG ES cells; (B) paternal chromosome, AG ES cells. Arrows pointing up or down indicate increased or decreased DNase I sensitivity of the in vivo-treated versus the in vitro-treated samples; shorter arrows indicate lower DNase I sensitivity in AG cells compared to PG cells; filled or open squares indicate increases or decreases, respectively, in DMS reactivity; open or closed circles indicate hypomethylated or hypermethylated CpGs, respectively; grey circles stand for partially methylated sites; brackets indicate the sequences analyzed.

    Article Snippet: The cells were immediately pelleted, then resuspended in solution II containing DNase I (1 μg/ml; Boehringer Mannheim catalog no. 104132), and incubated for 1 to 6 min at RT to obtain an optimal nicking frequency of one nick every 200 to 800 nucleotides, as assessed by alkaline gel electrophoresis after DNA purification and before LMPCR.

    Techniques: In Vivo, In Vitro, Methylation

    In vivo footprinting of the proximal H19 promoter in PG ES cells, WT ES cells, and AG ES cells. Methylation (Meth.), genomic DNase I footprinting (DNA-se I), and in vivo DMS footprinting (DMS) analyses were conducted. The N primer set was used for LMPCR analysis of the upper strand. Pg, PG ES cells; Wt, WT ES cells; Ag, AG ES cells. Open and filled circles indicate unmethylated and fully methylated CpGs, respectively, representing PG ES cells on the left and AG ES cells on the right; grey bars indicate partial DNase I footprints in PG ES cells; black bars indicate complete protection; black and open squares indicate increases and decreases, respectively, in DMS reactivity; A, AG ES cells; P, PG ES cells; brackets on the right side show consensus binding sites for transcription factors.

    Journal: Molecular and Cellular Biology

    Article Title: Characterization of Novel Parent-Specific Epigenetic Modifications Upstream of the Imprinted Mouse H19 Gene

    doi:

    Figure Lengend Snippet: In vivo footprinting of the proximal H19 promoter in PG ES cells, WT ES cells, and AG ES cells. Methylation (Meth.), genomic DNase I footprinting (DNA-se I), and in vivo DMS footprinting (DMS) analyses were conducted. The N primer set was used for LMPCR analysis of the upper strand. Pg, PG ES cells; Wt, WT ES cells; Ag, AG ES cells. Open and filled circles indicate unmethylated and fully methylated CpGs, respectively, representing PG ES cells on the left and AG ES cells on the right; grey bars indicate partial DNase I footprints in PG ES cells; black bars indicate complete protection; black and open squares indicate increases and decreases, respectively, in DMS reactivity; A, AG ES cells; P, PG ES cells; brackets on the right side show consensus binding sites for transcription factors.

    Article Snippet: The cells were immediately pelleted, then resuspended in solution II containing DNase I (1 μg/ml; Boehringer Mannheim catalog no. 104132), and incubated for 1 to 6 min at RT to obtain an optimal nicking frequency of one nick every 200 to 800 nucleotides, as assessed by alkaline gel electrophoresis after DNA purification and before LMPCR.

    Techniques: In Vivo, Footprinting, Methylation, Binding Assay

    Summary of the promoter footprinting data. Primary data were taken from experiments with the K, L, M, N, and O primer sets. (A) Maternal chromosome, PG ES cells; (B) paternal chromosome, AG ES cells. Circles around the DNA sequences indicate consensus sequences for transcription factors. Striped bars indicate partial DNase I footprints; black bars indicate complete protection; arrows pointing up or down indicate increased DNase I sensitivity of the in vivo-treated versus the in vitro-treated samples; filled or open squares indicate increases or decreases, respectively, in DMS reactivity; open or closed circles stand for hypomethylated or hypermethylated CpGs, respectively; grey circles stand for partially methylated sites.

    Journal: Molecular and Cellular Biology

    Article Title: Characterization of Novel Parent-Specific Epigenetic Modifications Upstream of the Imprinted Mouse H19 Gene

    doi:

    Figure Lengend Snippet: Summary of the promoter footprinting data. Primary data were taken from experiments with the K, L, M, N, and O primer sets. (A) Maternal chromosome, PG ES cells; (B) paternal chromosome, AG ES cells. Circles around the DNA sequences indicate consensus sequences for transcription factors. Striped bars indicate partial DNase I footprints; black bars indicate complete protection; arrows pointing up or down indicate increased DNase I sensitivity of the in vivo-treated versus the in vitro-treated samples; filled or open squares indicate increases or decreases, respectively, in DMS reactivity; open or closed circles stand for hypomethylated or hypermethylated CpGs, respectively; grey circles stand for partially methylated sites.

    Article Snippet: The cells were immediately pelleted, then resuspended in solution II containing DNase I (1 μg/ml; Boehringer Mannheim catalog no. 104132), and incubated for 1 to 6 min at RT to obtain an optimal nicking frequency of one nick every 200 to 800 nucleotides, as assessed by alkaline gel electrophoresis after DNA purification and before LMPCR.

    Techniques: Footprinting, In Vivo, In Vitro, Methylation

    A schematic representation of the origin is aligned down the left-hand side of autoradiographs from denaturing polyacrylamide gels displaying hydroxyl radical (MPE) and DNase I cleavage profiles obtained with naked 32 P-, 3′-end-labeled DNA or following its incubation with NS1 and/or HMG1 as indicated. Each lane displays products from the upper strand, containing the nick site. The positions of the ACCA tetranucleotide motifs that make up the NS1 binding sites are indicated down the right-hand side, together with a region of overcutting and undercutting, obtained with DNase I in the presence of both NS1 and HMG1, that reflects the formation of a double helical DNA loop.

    Journal: Journal of Virology

    Article Title: Two Widely Spaced Initiator Binding Sites Create an HMG1-Dependent Parvovirus Rolling-Hairpin Replication Origin

    doi:

    Figure Lengend Snippet: A schematic representation of the origin is aligned down the left-hand side of autoradiographs from denaturing polyacrylamide gels displaying hydroxyl radical (MPE) and DNase I cleavage profiles obtained with naked 32 P-, 3′-end-labeled DNA or following its incubation with NS1 and/or HMG1 as indicated. Each lane displays products from the upper strand, containing the nick site. The positions of the ACCA tetranucleotide motifs that make up the NS1 binding sites are indicated down the right-hand side, together with a region of overcutting and undercutting, obtained with DNase I in the presence of both NS1 and HMG1, that reflects the formation of a double helical DNA loop.

    Article Snippet: Surprisingly, MPE-Fe(II) footprints show that NS1 makes extremely close contact with the DNA throughout much of the region protected from DNase I.

    Techniques: Labeling, Incubation, Binding Assay