bal 31 nuclease assay  (New England Biolabs)


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    Structured Review

    New England Biolabs bal 31 nuclease assay
    The character of hybridization signals revealed using Southern hybridization. The DIG-labeled probe specific to (TTAGG) n ( a ) and (TCAGG)n ( b ) sequence was hybridized with RsaI/HinfI-digested genomic DNAs of selected coleopteran species. ( c ) <t>Bal</t> 31 exonuclease digestion of genomic DNA of Anoplotrupes stercorosus and Clerus muttilarius. Time-course digestion intervals are indicated above the lanes.
    Bal 31 Nuclease Assay, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Average 93 stars, based on 1 article reviews
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    bal 31 nuclease assay - by Bioz Stars, 2022-07
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    Images

    1) Product Images from "Telomeric DNA sequences in beetle taxa vary with species richness"

    Article Title: Telomeric DNA sequences in beetle taxa vary with species richness

    Journal: Scientific Reports

    doi: 10.1038/s41598-021-92705-y

    The character of hybridization signals revealed using Southern hybridization. The DIG-labeled probe specific to (TTAGG) n ( a ) and (TCAGG)n ( b ) sequence was hybridized with RsaI/HinfI-digested genomic DNAs of selected coleopteran species. ( c ) Bal 31 exonuclease digestion of genomic DNA of Anoplotrupes stercorosus and Clerus muttilarius. Time-course digestion intervals are indicated above the lanes.
    Figure Legend Snippet: The character of hybridization signals revealed using Southern hybridization. The DIG-labeled probe specific to (TTAGG) n ( a ) and (TCAGG)n ( b ) sequence was hybridized with RsaI/HinfI-digested genomic DNAs of selected coleopteran species. ( c ) Bal 31 exonuclease digestion of genomic DNA of Anoplotrupes stercorosus and Clerus muttilarius. Time-course digestion intervals are indicated above the lanes.

    Techniques Used: Hybridization, Labeling, Sequencing

    2) Product Images from "Production of DNA minicircles less than 250 base pairs through a novel concentrated DNA circularization assay enabling minicircle design with NF-κB inhibition activity"

    Article Title: Production of DNA minicircles less than 250 base pairs through a novel concentrated DNA circularization assay enabling minicircle design with NF-κB inhibition activity

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkw1034

    CNDCA-based production of supercoiled minicircle. Image of stained native polyacrylamide gel showing 95 bp minicircle topoisomers of decreased linking number (ΔLk values of −1 and −2) (lanes 4, 6) as indicated on the right and obtained by religation of the nicked minicircle in the presence of EtBr. The slowest migrating band corresponds to the relaxed minicircle that migrates expectedly at the same rate as the nicked minicircle (lanes 1 and 2). Digestion of minicircle samples by the nuclease Bal31 is indicated on the right. The position of the molecular mass markers in bp is indicated on the left.
    Figure Legend Snippet: CNDCA-based production of supercoiled minicircle. Image of stained native polyacrylamide gel showing 95 bp minicircle topoisomers of decreased linking number (ΔLk values of −1 and −2) (lanes 4, 6) as indicated on the right and obtained by religation of the nicked minicircle in the presence of EtBr. The slowest migrating band corresponds to the relaxed minicircle that migrates expectedly at the same rate as the nicked minicircle (lanes 1 and 2). Digestion of minicircle samples by the nuclease Bal31 is indicated on the right. The position of the molecular mass markers in bp is indicated on the left.

    Techniques Used: Staining

    3) Product Images from "Design and characterization of decoy oligonucleotides containing locked nucleic acids"

    Article Title: Design and characterization of decoy oligonucleotides containing locked nucleic acids

    Journal: Nucleic Acids Research

    doi:

    Susceptibility to BAL-31 nuclease degradation of LNA-modified ODNs. LNA-modified [NF-κB(a), (b), (c), (c+b) and (b+c)] and control phosphodiester [NF-κB(d)] decoy molecules were incubated for different lengths of time, as indicated, with 0.5 U/ml BAL-31 and then submitted to electrophoretic separation on 2.5% (w/v) agarose gels. Detection and quantitation of the ethidium bromide stained bands were performed in a Molecular Analyst. Volume densities of the bands are expressed as percent ODN decoy remaining with respect to the relative time zero value and are shown as line graphs.
    Figure Legend Snippet: Susceptibility to BAL-31 nuclease degradation of LNA-modified ODNs. LNA-modified [NF-κB(a), (b), (c), (c+b) and (b+c)] and control phosphodiester [NF-κB(d)] decoy molecules were incubated for different lengths of time, as indicated, with 0.5 U/ml BAL-31 and then submitted to electrophoretic separation on 2.5% (w/v) agarose gels. Detection and quantitation of the ethidium bromide stained bands were performed in a Molecular Analyst. Volume densities of the bands are expressed as percent ODN decoy remaining with respect to the relative time zero value and are shown as line graphs.

    Techniques Used: Modification, Incubation, Quantitation Assay, Staining

    4) Product Images from "Bending modes of DNA directly addressed by cryo-electron microscopy of DNA minicircles"

    Article Title: Bending modes of DNA directly addressed by cryo-electron microscopy of DNA minicircles

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkp137

    The formation and purification of 94-bp-long covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that after the ligation reaction of minicircles with nicks, new species appear on the gel (compare lanes 3 with 2). Only these new species resist digestion by Bal31 (lane 4) indicating that they are covalently closed DNA molecules. The electrophoretic migration of these new species is as expected for monomeric, dimeric and trimeric rings that acquired writhe due to ethidium bromide intercalation to covalently closed DNA molecules.
    Figure Legend Snippet: The formation and purification of 94-bp-long covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that after the ligation reaction of minicircles with nicks, new species appear on the gel (compare lanes 3 with 2). Only these new species resist digestion by Bal31 (lane 4) indicating that they are covalently closed DNA molecules. The electrophoretic migration of these new species is as expected for monomeric, dimeric and trimeric rings that acquired writhe due to ethidium bromide intercalation to covalently closed DNA molecules.

    Techniques Used: Purification, Agarose Gel Electrophoresis, Ligation, Migration

    Bal31 nuclease detects a destabilized duplex DNA structure in covalently closed supercoiled DNA molecules but does not reveal the presence of kinks in 94-bp covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that while negatively supercoiled DNA (lane 6) and nonligated nicked rings are completely digested by Bal31 nuclease (see lanes 2–4), the covalently closed monomeric, dimerc, trimeric and tetrameric circles (indicated with arrows) remain resistant to action of Bal31 nuclease (compare lanes 4 and 3).
    Figure Legend Snippet: Bal31 nuclease detects a destabilized duplex DNA structure in covalently closed supercoiled DNA molecules but does not reveal the presence of kinks in 94-bp covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that while negatively supercoiled DNA (lane 6) and nonligated nicked rings are completely digested by Bal31 nuclease (see lanes 2–4), the covalently closed monomeric, dimerc, trimeric and tetrameric circles (indicated with arrows) remain resistant to action of Bal31 nuclease (compare lanes 4 and 3).

    Techniques Used: Agarose Gel Electrophoresis

    5) Product Images from "Bending modes of DNA directly addressed by cryo-electron microscopy of DNA minicircles"

    Article Title: Bending modes of DNA directly addressed by cryo-electron microscopy of DNA minicircles

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkp137

    The formation and purification of 94-bp-long covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that after the ligation reaction of minicircles with nicks, new species appear on the gel (compare lanes 3 with 2). Only these new species resist digestion by Bal31 (lane 4) indicating that they are covalently closed DNA molecules. The electrophoretic migration of these new species is as expected for monomeric, dimeric and trimeric rings that acquired writhe due to ethidium bromide intercalation to covalently closed DNA molecules.
    Figure Legend Snippet: The formation and purification of 94-bp-long covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that after the ligation reaction of minicircles with nicks, new species appear on the gel (compare lanes 3 with 2). Only these new species resist digestion by Bal31 (lane 4) indicating that they are covalently closed DNA molecules. The electrophoretic migration of these new species is as expected for monomeric, dimeric and trimeric rings that acquired writhe due to ethidium bromide intercalation to covalently closed DNA molecules.

    Techniques Used: Purification, Agarose Gel Electrophoresis, Ligation, Migration

    Bal31 nuclease detects a destabilized duplex DNA structure in covalently closed supercoiled DNA molecules but does not reveal the presence of kinks in 94-bp covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that while negatively supercoiled DNA (lane 6) and nonligated nicked rings are completely digested by Bal31 nuclease (see lanes 2–4), the covalently closed monomeric, dimerc, trimeric and tetrameric circles (indicated with arrows) remain resistant to action of Bal31 nuclease (compare lanes 4 and 3).
    Figure Legend Snippet: Bal31 nuclease detects a destabilized duplex DNA structure in covalently closed supercoiled DNA molecules but does not reveal the presence of kinks in 94-bp covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that while negatively supercoiled DNA (lane 6) and nonligated nicked rings are completely digested by Bal31 nuclease (see lanes 2–4), the covalently closed monomeric, dimerc, trimeric and tetrameric circles (indicated with arrows) remain resistant to action of Bal31 nuclease (compare lanes 4 and 3).

    Techniques Used: Agarose Gel Electrophoresis

    6) Product Images from "Cooperative kinking at distant sites in mechanically stressed DNA"

    Article Title: Cooperative kinking at distant sites in mechanically stressed DNA

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkr666

    DNA minicircles with 100, 106 and 108 bp, their purity and their susceptibility to Bal31 digestion. In a 10% denaturing PAGE gel (7 M urea), the denatured linear DNA fragments used as a marker, migrate much quicker than circular, covalently closed DNA minicircles in which the two strands cannot separate in space. Notice the presence of only one topoisomer in each category of minicircles and the sensitivity of minicircles with 100 bp to Bal31 nuclease.
    Figure Legend Snippet: DNA minicircles with 100, 106 and 108 bp, their purity and their susceptibility to Bal31 digestion. In a 10% denaturing PAGE gel (7 M urea), the denatured linear DNA fragments used as a marker, migrate much quicker than circular, covalently closed DNA minicircles in which the two strands cannot separate in space. Notice the presence of only one topoisomer in each category of minicircles and the sensitivity of minicircles with 100 bp to Bal31 nuclease.

    Techniques Used: Polyacrylamide Gel Electrophoresis, Marker

    7) Product Images from "Cooperative kinking at distant sites in mechanically stressed DNA"

    Article Title: Cooperative kinking at distant sites in mechanically stressed DNA

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkr666

    DNA minicircles with 100, 106 and 108 bp, their purity and their susceptibility to Bal31 digestion. In a 10% denaturing PAGE gel (7 M urea), the denatured linear DNA fragments used as a marker, migrate much quicker than circular, covalently closed DNA minicircles in which the two strands cannot separate in space. Notice the presence of only one topoisomer in each category of minicircles and the sensitivity of minicircles with 100 bp to Bal31 nuclease.
    Figure Legend Snippet: DNA minicircles with 100, 106 and 108 bp, their purity and their susceptibility to Bal31 digestion. In a 10% denaturing PAGE gel (7 M urea), the denatured linear DNA fragments used as a marker, migrate much quicker than circular, covalently closed DNA minicircles in which the two strands cannot separate in space. Notice the presence of only one topoisomer in each category of minicircles and the sensitivity of minicircles with 100 bp to Bal31 nuclease.

    Techniques Used: Polyacrylamide Gel Electrophoresis, Marker

    8) Product Images from "Bending modes of DNA directly addressed by cryo-electron microscopy of DNA minicircles"

    Article Title: Bending modes of DNA directly addressed by cryo-electron microscopy of DNA minicircles

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkp137

    The formation and purification of 94-bp-long covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that after the ligation reaction of minicircles with nicks, new species appear on the gel (compare lanes 3 with 2). Only these new species resist digestion by Bal31 (lane 4) indicating that they are covalently closed DNA molecules. The electrophoretic migration of these new species is as expected for monomeric, dimeric and trimeric rings that acquired writhe due to ethidium bromide intercalation to covalently closed DNA molecules.
    Figure Legend Snippet: The formation and purification of 94-bp-long covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that after the ligation reaction of minicircles with nicks, new species appear on the gel (compare lanes 3 with 2). Only these new species resist digestion by Bal31 (lane 4) indicating that they are covalently closed DNA molecules. The electrophoretic migration of these new species is as expected for monomeric, dimeric and trimeric rings that acquired writhe due to ethidium bromide intercalation to covalently closed DNA molecules.

    Techniques Used: Purification, Agarose Gel Electrophoresis, Ligation, Migration

    Bal31 nuclease detects a destabilized duplex DNA structure in covalently closed supercoiled DNA molecules but does not reveal the presence of kinks in 94-bp covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that while negatively supercoiled DNA (lane 6) and nonligated nicked rings are completely digested by Bal31 nuclease (see lanes 2–4), the covalently closed monomeric, dimerc, trimeric and tetrameric circles (indicated with arrows) remain resistant to action of Bal31 nuclease (compare lanes 4 and 3).
    Figure Legend Snippet: Bal31 nuclease detects a destabilized duplex DNA structure in covalently closed supercoiled DNA molecules but does not reveal the presence of kinks in 94-bp covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that while negatively supercoiled DNA (lane 6) and nonligated nicked rings are completely digested by Bal31 nuclease (see lanes 2–4), the covalently closed monomeric, dimerc, trimeric and tetrameric circles (indicated with arrows) remain resistant to action of Bal31 nuclease (compare lanes 4 and 3).

    Techniques Used: Agarose Gel Electrophoresis

    9) Product Images from "Bending modes of DNA directly addressed by cryo-electron microscopy of DNA minicircles"

    Article Title: Bending modes of DNA directly addressed by cryo-electron microscopy of DNA minicircles

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkp137

    The formation and purification of 94-bp-long covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that after the ligation reaction of minicircles with nicks, new species appear on the gel (compare lanes 3 with 2). Only these new species resist digestion by Bal31 (lane 4) indicating that they are covalently closed DNA molecules. The electrophoretic migration of these new species is as expected for monomeric, dimeric and trimeric rings that acquired writhe due to ethidium bromide intercalation to covalently closed DNA molecules.
    Figure Legend Snippet: The formation and purification of 94-bp-long covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that after the ligation reaction of minicircles with nicks, new species appear on the gel (compare lanes 3 with 2). Only these new species resist digestion by Bal31 (lane 4) indicating that they are covalently closed DNA molecules. The electrophoretic migration of these new species is as expected for monomeric, dimeric and trimeric rings that acquired writhe due to ethidium bromide intercalation to covalently closed DNA molecules.

    Techniques Used: Purification, Agarose Gel Electrophoresis, Ligation, Migration

    Bal31 nuclease detects a destabilized duplex DNA structure in covalently closed supercoiled DNA molecules but does not reveal the presence of kinks in 94-bp covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that while negatively supercoiled DNA (lane 6) and nonligated nicked rings are completely digested by Bal31 nuclease (see lanes 2–4), the covalently closed monomeric, dimerc, trimeric and tetrameric circles (indicated with arrows) remain resistant to action of Bal31 nuclease (compare lanes 4 and 3).
    Figure Legend Snippet: Bal31 nuclease detects a destabilized duplex DNA structure in covalently closed supercoiled DNA molecules but does not reveal the presence of kinks in 94-bp covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that while negatively supercoiled DNA (lane 6) and nonligated nicked rings are completely digested by Bal31 nuclease (see lanes 2–4), the covalently closed monomeric, dimerc, trimeric and tetrameric circles (indicated with arrows) remain resistant to action of Bal31 nuclease (compare lanes 4 and 3).

    Techniques Used: Agarose Gel Electrophoresis

    10) Product Images from "Bending modes of DNA directly addressed by cryo-electron microscopy of DNA minicircles"

    Article Title: Bending modes of DNA directly addressed by cryo-electron microscopy of DNA minicircles

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkp137

    The formation and purification of 94-bp-long covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that after the ligation reaction of minicircles with nicks, new species appear on the gel (compare lanes 3 with 2). Only these new species resist digestion by Bal31 (lane 4) indicating that they are covalently closed DNA molecules. The electrophoretic migration of these new species is as expected for monomeric, dimeric and trimeric rings that acquired writhe due to ethidium bromide intercalation to covalently closed DNA molecules.
    Figure Legend Snippet: The formation and purification of 94-bp-long covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that after the ligation reaction of minicircles with nicks, new species appear on the gel (compare lanes 3 with 2). Only these new species resist digestion by Bal31 (lane 4) indicating that they are covalently closed DNA molecules. The electrophoretic migration of these new species is as expected for monomeric, dimeric and trimeric rings that acquired writhe due to ethidium bromide intercalation to covalently closed DNA molecules.

    Techniques Used: Purification, Agarose Gel Electrophoresis, Ligation, Migration

    Bal31 nuclease detects a destabilized duplex DNA structure in covalently closed supercoiled DNA molecules but does not reveal the presence of kinks in 94-bp covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that while negatively supercoiled DNA (lane 6) and nonligated nicked rings are completely digested by Bal31 nuclease (see lanes 2–4), the covalently closed monomeric, dimerc, trimeric and tetrameric circles (indicated with arrows) remain resistant to action of Bal31 nuclease (compare lanes 4 and 3).
    Figure Legend Snippet: Bal31 nuclease detects a destabilized duplex DNA structure in covalently closed supercoiled DNA molecules but does not reveal the presence of kinks in 94-bp covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that while negatively supercoiled DNA (lane 6) and nonligated nicked rings are completely digested by Bal31 nuclease (see lanes 2–4), the covalently closed monomeric, dimerc, trimeric and tetrameric circles (indicated with arrows) remain resistant to action of Bal31 nuclease (compare lanes 4 and 3).

    Techniques Used: Agarose Gel Electrophoresis

    11) Product Images from "Chromatin features of plant telomeric sequences at terminal vs. internal positions"

    Article Title: Chromatin features of plant telomeric sequences at terminal vs. internal positions

    Journal: Frontiers in Plant Science

    doi: 10.3389/fpls.2014.00593

    Internal localization of telomeric repeats in the B. antipoda genome. (A) High molecular weight DNA digested by Bal31 nuclease and a mix of frequently cutting restriction enzymes was analyzed by pulse field gel electrophoresis. Signals after hybridization with the radioactively labeled telomeric oligonucleotide showed resistance of telomeric sequences to Bal31 cleavage. The apparent decrease in signal intensity after 30 min of Bal31 digestion reflects variation in sample loading rather than a change due to the Bal31 treatment (compare to the signal after 45 min of Bal31). (B) DNA released into solution in the course of Bal31 digestions was purified and subjected to conventional agarose electrophoresis. The numbers above lines indicate the time of Bal31 digestion in minutes. M, DNA size markers.
    Figure Legend Snippet: Internal localization of telomeric repeats in the B. antipoda genome. (A) High molecular weight DNA digested by Bal31 nuclease and a mix of frequently cutting restriction enzymes was analyzed by pulse field gel electrophoresis. Signals after hybridization with the radioactively labeled telomeric oligonucleotide showed resistance of telomeric sequences to Bal31 cleavage. The apparent decrease in signal intensity after 30 min of Bal31 digestion reflects variation in sample loading rather than a change due to the Bal31 treatment (compare to the signal after 45 min of Bal31). (B) DNA released into solution in the course of Bal31 digestions was purified and subjected to conventional agarose electrophoresis. The numbers above lines indicate the time of Bal31 digestion in minutes. M, DNA size markers.

    Techniques Used: Molecular Weight, Nucleic Acid Electrophoresis, Hybridization, Labeling, Purification, Electrophoresis

    Relative cytosine methylation in telomeres of Nicotiana species. (A) After Bal31 treatment of N. tabacum high molecular weight DNA, TRF analysis was used to control the efficiency of digestion. A loss of the telomere-specific hybridization signal was observed in the course of Bal31 treatment. After 15 min of Bal31 digestion loss of the signal is evident but telomere erosion is not clear, whereas after 45- and 90-min treatments, telomeres were efficiently degraded. Time of Bal31 digestion is given above the lanes. M – DNA size marker. (B) Dot-blot analysis of Bal31-digested DNA from N. tabacum and N. tomentosiformis after treatment with sodium bisulfite. Samples were loaded onto a membrane and hybridized with radioactively labeled probes to detect the total signal of telomeres (loading probe pltel-C complementary to the telomeric G-strand) and the portion of methylated telomeres (DEGENER probe). Time of the Bal31 digestion in minutes is given above the membranes. +, positive hybridization control (tobacco DNA without the bisulfite treatment); −, negative control (DNA from the pUC19 plasmid). (C) Relative density of methylated cytosines along telomeres, calculated as the DEGENER/loading hybridization signals ratio. The ratio in Bal31 non-treated samples was arbitrarily taken as 1. Six independent experiments were performed.
    Figure Legend Snippet: Relative cytosine methylation in telomeres of Nicotiana species. (A) After Bal31 treatment of N. tabacum high molecular weight DNA, TRF analysis was used to control the efficiency of digestion. A loss of the telomere-specific hybridization signal was observed in the course of Bal31 treatment. After 15 min of Bal31 digestion loss of the signal is evident but telomere erosion is not clear, whereas after 45- and 90-min treatments, telomeres were efficiently degraded. Time of Bal31 digestion is given above the lanes. M – DNA size marker. (B) Dot-blot analysis of Bal31-digested DNA from N. tabacum and N. tomentosiformis after treatment with sodium bisulfite. Samples were loaded onto a membrane and hybridized with radioactively labeled probes to detect the total signal of telomeres (loading probe pltel-C complementary to the telomeric G-strand) and the portion of methylated telomeres (DEGENER probe). Time of the Bal31 digestion in minutes is given above the membranes. +, positive hybridization control (tobacco DNA without the bisulfite treatment); −, negative control (DNA from the pUC19 plasmid). (C) Relative density of methylated cytosines along telomeres, calculated as the DEGENER/loading hybridization signals ratio. The ratio in Bal31 non-treated samples was arbitrarily taken as 1. Six independent experiments were performed.

    Techniques Used: Methylation, Molecular Weight, Hybridization, Marker, Dot Blot, Labeling, Negative Control, Plasmid Preparation

    Methylation of cytosines of B. antipoda telomeric repeats. (A) Analysis of global methylation of cytosines in telomeric repeats of B. antipoda . High molecular weight DNA was treated with Bal31, converted with bisulfite and analyzed by Southern hybridization against loading and DEGENER probes. Signals with the DEGENER probe indicating methylated cytosines in telomeric repeats were obtained in all samples. The time of Bal31 digestion is given above the membranes. +, positive hybridization control (tobacco DNA without the bisulfite treatment); −, negative control (DNA from pUC19 plasmid). (B) An example of bisulfite sequencing data for Ba493 and Ba576 regions in adult leaves. Non-symmetrically located cytosines are depicted as green triangles; full triangle, methylated; empty triangle, non-methylated cytosine. Cytosines located in perfect telomeric repeats are underlined. The first line of the scheme where all cytosines in all sequence contexts are presented as full figures represents position of respective cytosine. (C) Graphical representation of telomeric cytosine methylation within the ITR-containing sequences Ba493 and Ba576. Three tissues were analyzed to reveal possible tissue-specific methylation pattern. “all,” all cytosines located in non-symmetrical sequence context; “telomeric,” cytosines located in perfect telomeric repeats. Data were processed by One-Way independent ANOVA test with standard weighted-means, * is used for P
    Figure Legend Snippet: Methylation of cytosines of B. antipoda telomeric repeats. (A) Analysis of global methylation of cytosines in telomeric repeats of B. antipoda . High molecular weight DNA was treated with Bal31, converted with bisulfite and analyzed by Southern hybridization against loading and DEGENER probes. Signals with the DEGENER probe indicating methylated cytosines in telomeric repeats were obtained in all samples. The time of Bal31 digestion is given above the membranes. +, positive hybridization control (tobacco DNA without the bisulfite treatment); −, negative control (DNA from pUC19 plasmid). (B) An example of bisulfite sequencing data for Ba493 and Ba576 regions in adult leaves. Non-symmetrically located cytosines are depicted as green triangles; full triangle, methylated; empty triangle, non-methylated cytosine. Cytosines located in perfect telomeric repeats are underlined. The first line of the scheme where all cytosines in all sequence contexts are presented as full figures represents position of respective cytosine. (C) Graphical representation of telomeric cytosine methylation within the ITR-containing sequences Ba493 and Ba576. Three tissues were analyzed to reveal possible tissue-specific methylation pattern. “all,” all cytosines located in non-symmetrical sequence context; “telomeric,” cytosines located in perfect telomeric repeats. Data were processed by One-Way independent ANOVA test with standard weighted-means, * is used for P

    Techniques Used: Methylation, Molecular Weight, Hybridization, Negative Control, Plasmid Preparation, Methylation Sequencing, Sequencing

    12) Product Images from "Analysis of Carica papaya Telomeres and Telomere-Associated Proteins: Insights into the Evolution of Telomere Maintenance in Brassicales"

    Article Title: Analysis of Carica papaya Telomeres and Telomere-Associated Proteins: Insights into the Evolution of Telomere Maintenance in Brassicales

    Journal:

    doi: 10.1007/s12042-008-9018-x

    Telomere length analysis in members of the Brassicales Order. a TRF analysis of papaya telomeres. Lane 1, Tru1 I digestion of genomic DNA without prior Bal 31 treatment (0 min), lanes 2–6, Tru1 I digestion of genomic DNA after various incubation
    Figure Legend Snippet: Telomere length analysis in members of the Brassicales Order. a TRF analysis of papaya telomeres. Lane 1, Tru1 I digestion of genomic DNA without prior Bal 31 treatment (0 min), lanes 2–6, Tru1 I digestion of genomic DNA after various incubation

    Techniques Used: Incubation

    13) Product Images from "Bending modes of DNA directly addressed by cryo-electron microscopy of DNA minicircles"

    Article Title: Bending modes of DNA directly addressed by cryo-electron microscopy of DNA minicircles

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkp137

    The formation and purification of 94-bp-long covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that after the ligation reaction of minicircles with nicks, new species appear on the gel (compare lanes 3 with 2). Only these new species resist digestion by Bal31 (lane 4) indicating that they are covalently closed DNA molecules. The electrophoretic migration of these new species is as expected for monomeric, dimeric and trimeric rings that acquired writhe due to ethidium bromide intercalation to covalently closed DNA molecules.
    Figure Legend Snippet: The formation and purification of 94-bp-long covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that after the ligation reaction of minicircles with nicks, new species appear on the gel (compare lanes 3 with 2). Only these new species resist digestion by Bal31 (lane 4) indicating that they are covalently closed DNA molecules. The electrophoretic migration of these new species is as expected for monomeric, dimeric and trimeric rings that acquired writhe due to ethidium bromide intercalation to covalently closed DNA molecules.

    Techniques Used: Purification, Agarose Gel Electrophoresis, Ligation, Migration

    Bal31 nuclease detects a destabilized duplex DNA structure in covalently closed supercoiled DNA molecules but does not reveal the presence of kinks in 94-bp covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that while negatively supercoiled DNA (lane 6) and nonligated nicked rings are completely digested by Bal31 nuclease (see lanes 2–4), the covalently closed monomeric, dimerc, trimeric and tetrameric circles (indicated with arrows) remain resistant to action of Bal31 nuclease (compare lanes 4 and 3).
    Figure Legend Snippet: Bal31 nuclease detects a destabilized duplex DNA structure in covalently closed supercoiled DNA molecules but does not reveal the presence of kinks in 94-bp covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that while negatively supercoiled DNA (lane 6) and nonligated nicked rings are completely digested by Bal31 nuclease (see lanes 2–4), the covalently closed monomeric, dimerc, trimeric and tetrameric circles (indicated with arrows) remain resistant to action of Bal31 nuclease (compare lanes 4 and 3).

    Techniques Used: Agarose Gel Electrophoresis

    14) Product Images from "Bending modes of DNA directly addressed by cryo-electron microscopy of DNA minicircles"

    Article Title: Bending modes of DNA directly addressed by cryo-electron microscopy of DNA minicircles

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkp137

    The formation and purification of 94-bp-long covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that after the ligation reaction of minicircles with nicks, new species appear on the gel (compare lanes 3 with 2). Only these new species resist digestion by Bal31 (lane 4) indicating that they are covalently closed DNA molecules. The electrophoretic migration of these new species is as expected for monomeric, dimeric and trimeric rings that acquired writhe due to ethidium bromide intercalation to covalently closed DNA molecules.
    Figure Legend Snippet: The formation and purification of 94-bp-long covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that after the ligation reaction of minicircles with nicks, new species appear on the gel (compare lanes 3 with 2). Only these new species resist digestion by Bal31 (lane 4) indicating that they are covalently closed DNA molecules. The electrophoretic migration of these new species is as expected for monomeric, dimeric and trimeric rings that acquired writhe due to ethidium bromide intercalation to covalently closed DNA molecules.

    Techniques Used: Purification, Agarose Gel Electrophoresis, Ligation, Migration

    Bal31 nuclease detects a destabilized duplex DNA structure in covalently closed supercoiled DNA molecules but does not reveal the presence of kinks in 94-bp covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that while negatively supercoiled DNA (lane 6) and nonligated nicked rings are completely digested by Bal31 nuclease (see lanes 2–4), the covalently closed monomeric, dimerc, trimeric and tetrameric circles (indicated with arrows) remain resistant to action of Bal31 nuclease (compare lanes 4 and 3).
    Figure Legend Snippet: Bal31 nuclease detects a destabilized duplex DNA structure in covalently closed supercoiled DNA molecules but does not reveal the presence of kinks in 94-bp covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that while negatively supercoiled DNA (lane 6) and nonligated nicked rings are completely digested by Bal31 nuclease (see lanes 2–4), the covalently closed monomeric, dimerc, trimeric and tetrameric circles (indicated with arrows) remain resistant to action of Bal31 nuclease (compare lanes 4 and 3).

    Techniques Used: Agarose Gel Electrophoresis

    15) Product Images from "Application of 5-Methylcytosine DNA Glycosylase to the Quantitative Analysis of DNA Methylation"

    Article Title: Application of 5-Methylcytosine DNA Glycosylase to the Quantitative Analysis of DNA Methylation

    Journal: International Journal of Molecular Sciences

    doi: 10.3390/ijms22031072

    Quantitative analysis of DNA methylation by DEMETER (DME) treatment and PCR amplification. ( A ) Schematic diagrams of differentially methylated regions. Yeast genomic DNA was in vitro methylated with Hpa II methyltransferase for CCGG methylation. Methylated regions with a single or three CCGG sites were subjected to DME treatment and PCR amplification with F1a/R1 and F1b/R1 primer pairs, respectively. Unmethylated control region was amplified with F2/R2 primer pairs. ( B ) DME-induced SSBs decreases PCR amplification in proportion to DNA methylation levels. Hpa II-methylated yeast genomic DNA was treated with DME and subjected to multiplex PCR amplification. Primer pairs are denoted to the right of the panel. ( C ) DME and BAL-31 treatment induces DSBs at methylated regions. DSB formation by DME and BAL-31 treatment prevents PCR amplification regardless of the number of methylated sites.
    Figure Legend Snippet: Quantitative analysis of DNA methylation by DEMETER (DME) treatment and PCR amplification. ( A ) Schematic diagrams of differentially methylated regions. Yeast genomic DNA was in vitro methylated with Hpa II methyltransferase for CCGG methylation. Methylated regions with a single or three CCGG sites were subjected to DME treatment and PCR amplification with F1a/R1 and F1b/R1 primer pairs, respectively. Unmethylated control region was amplified with F2/R2 primer pairs. ( B ) DME-induced SSBs decreases PCR amplification in proportion to DNA methylation levels. Hpa II-methylated yeast genomic DNA was treated with DME and subjected to multiplex PCR amplification. Primer pairs are denoted to the right of the panel. ( C ) DME and BAL-31 treatment induces DSBs at methylated regions. DSB formation by DME and BAL-31 treatment prevents PCR amplification regardless of the number of methylated sites.

    Techniques Used: DNA Methylation Assay, Polymerase Chain Reaction, Amplification, Methylation, In Vitro, Multiplex Assay

    16) Product Images from "Bending modes of DNA directly addressed by cryo-electron microscopy of DNA minicircles"

    Article Title: Bending modes of DNA directly addressed by cryo-electron microscopy of DNA minicircles

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkp137

    The formation and purification of 94-bp-long covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that after the ligation reaction of minicircles with nicks, new species appear on the gel (compare lanes 3 with 2). Only these new species resist digestion by Bal31 (lane 4) indicating that they are covalently closed DNA molecules. The electrophoretic migration of these new species is as expected for monomeric, dimeric and trimeric rings that acquired writhe due to ethidium bromide intercalation to covalently closed DNA molecules.
    Figure Legend Snippet: The formation and purification of 94-bp-long covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that after the ligation reaction of minicircles with nicks, new species appear on the gel (compare lanes 3 with 2). Only these new species resist digestion by Bal31 (lane 4) indicating that they are covalently closed DNA molecules. The electrophoretic migration of these new species is as expected for monomeric, dimeric and trimeric rings that acquired writhe due to ethidium bromide intercalation to covalently closed DNA molecules.

    Techniques Used: Purification, Agarose Gel Electrophoresis, Ligation, Migration

    Bal31 nuclease detects a destabilized duplex DNA structure in covalently closed supercoiled DNA molecules but does not reveal the presence of kinks in 94-bp covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that while negatively supercoiled DNA (lane 6) and nonligated nicked rings are completely digested by Bal31 nuclease (see lanes 2–4), the covalently closed monomeric, dimerc, trimeric and tetrameric circles (indicated with arrows) remain resistant to action of Bal31 nuclease (compare lanes 4 and 3).
    Figure Legend Snippet: Bal31 nuclease detects a destabilized duplex DNA structure in covalently closed supercoiled DNA molecules but does not reveal the presence of kinks in 94-bp covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that while negatively supercoiled DNA (lane 6) and nonligated nicked rings are completely digested by Bal31 nuclease (see lanes 2–4), the covalently closed monomeric, dimerc, trimeric and tetrameric circles (indicated with arrows) remain resistant to action of Bal31 nuclease (compare lanes 4 and 3).

    Techniques Used: Agarose Gel Electrophoresis

    17) Product Images from "Bending modes of DNA directly addressed by cryo-electron microscopy of DNA minicircles"

    Article Title: Bending modes of DNA directly addressed by cryo-electron microscopy of DNA minicircles

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkp137

    The formation and purification of 94-bp-long covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that after the ligation reaction of minicircles with nicks, new species appear on the gel (compare lanes 3 with 2). Only these new species resist digestion by Bal31 (lane 4) indicating that they are covalently closed DNA molecules. The electrophoretic migration of these new species is as expected for monomeric, dimeric and trimeric rings that acquired writhe due to ethidium bromide intercalation to covalently closed DNA molecules.
    Figure Legend Snippet: The formation and purification of 94-bp-long covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that after the ligation reaction of minicircles with nicks, new species appear on the gel (compare lanes 3 with 2). Only these new species resist digestion by Bal31 (lane 4) indicating that they are covalently closed DNA molecules. The electrophoretic migration of these new species is as expected for monomeric, dimeric and trimeric rings that acquired writhe due to ethidium bromide intercalation to covalently closed DNA molecules.

    Techniques Used: Purification, Agarose Gel Electrophoresis, Ligation, Migration

    Bal31 nuclease detects a destabilized duplex DNA structure in covalently closed supercoiled DNA molecules but does not reveal the presence of kinks in 94-bp covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that while negatively supercoiled DNA (lane 6) and nonligated nicked rings are completely digested by Bal31 nuclease (see lanes 2–4), the covalently closed monomeric, dimerc, trimeric and tetrameric circles (indicated with arrows) remain resistant to action of Bal31 nuclease (compare lanes 4 and 3).
    Figure Legend Snippet: Bal31 nuclease detects a destabilized duplex DNA structure in covalently closed supercoiled DNA molecules but does not reveal the presence of kinks in 94-bp covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that while negatively supercoiled DNA (lane 6) and nonligated nicked rings are completely digested by Bal31 nuclease (see lanes 2–4), the covalently closed monomeric, dimerc, trimeric and tetrameric circles (indicated with arrows) remain resistant to action of Bal31 nuclease (compare lanes 4 and 3).

    Techniques Used: Agarose Gel Electrophoresis

    18) Product Images from "Cooperative kinking at distant sites in mechanically stressed DNA"

    Article Title: Cooperative kinking at distant sites in mechanically stressed DNA

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkr666

    DNA minicircles with 100, 106 and 108 bp, their purity and their susceptibility to Bal31 digestion. In a 10% denaturing PAGE gel (7 M urea), the denatured linear DNA fragments used as a marker, migrate much quicker than circular, covalently closed DNA minicircles in which the two strands cannot separate in space. Notice the presence of only one topoisomer in each category of minicircles and the sensitivity of minicircles with 100 bp to Bal31 nuclease.
    Figure Legend Snippet: DNA minicircles with 100, 106 and 108 bp, their purity and their susceptibility to Bal31 digestion. In a 10% denaturing PAGE gel (7 M urea), the denatured linear DNA fragments used as a marker, migrate much quicker than circular, covalently closed DNA minicircles in which the two strands cannot separate in space. Notice the presence of only one topoisomer in each category of minicircles and the sensitivity of minicircles with 100 bp to Bal31 nuclease.

    Techniques Used: Polyacrylamide Gel Electrophoresis, Marker

    19) Product Images from "Bending modes of DNA directly addressed by cryo-electron microscopy of DNA minicircles"

    Article Title: Bending modes of DNA directly addressed by cryo-electron microscopy of DNA minicircles

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkp137

    The formation and purification of 94-bp-long covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that after the ligation reaction of minicircles with nicks, new species appear on the gel (compare lanes 3 with 2). Only these new species resist digestion by Bal31 (lane 4) indicating that they are covalently closed DNA molecules. The electrophoretic migration of these new species is as expected for monomeric, dimeric and trimeric rings that acquired writhe due to ethidium bromide intercalation to covalently closed DNA molecules.
    Figure Legend Snippet: The formation and purification of 94-bp-long covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that after the ligation reaction of minicircles with nicks, new species appear on the gel (compare lanes 3 with 2). Only these new species resist digestion by Bal31 (lane 4) indicating that they are covalently closed DNA molecules. The electrophoretic migration of these new species is as expected for monomeric, dimeric and trimeric rings that acquired writhe due to ethidium bromide intercalation to covalently closed DNA molecules.

    Techniques Used: Purification, Agarose Gel Electrophoresis, Ligation, Migration

    Bal31 nuclease detects a destabilized duplex DNA structure in covalently closed supercoiled DNA molecules but does not reveal the presence of kinks in 94-bp covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that while negatively supercoiled DNA (lane 6) and nonligated nicked rings are completely digested by Bal31 nuclease (see lanes 2–4), the covalently closed monomeric, dimerc, trimeric and tetrameric circles (indicated with arrows) remain resistant to action of Bal31 nuclease (compare lanes 4 and 3).
    Figure Legend Snippet: Bal31 nuclease detects a destabilized duplex DNA structure in covalently closed supercoiled DNA molecules but does not reveal the presence of kinks in 94-bp covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that while negatively supercoiled DNA (lane 6) and nonligated nicked rings are completely digested by Bal31 nuclease (see lanes 2–4), the covalently closed monomeric, dimerc, trimeric and tetrameric circles (indicated with arrows) remain resistant to action of Bal31 nuclease (compare lanes 4 and 3).

    Techniques Used: Agarose Gel Electrophoresis

    20) Product Images from "Cooperative kinking at distant sites in mechanically stressed DNA"

    Article Title: Cooperative kinking at distant sites in mechanically stressed DNA

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkr666

    DNA minicircles with 100, 106 and 108 bp, their purity and their susceptibility to Bal31 digestion. In a 10% denaturing PAGE gel (7 M urea), the denatured linear DNA fragments used as a marker, migrate much quicker than circular, covalently closed DNA minicircles in which the two strands cannot separate in space. Notice the presence of only one topoisomer in each category of minicircles and the sensitivity of minicircles with 100 bp to Bal31 nuclease.
    Figure Legend Snippet: DNA minicircles with 100, 106 and 108 bp, their purity and their susceptibility to Bal31 digestion. In a 10% denaturing PAGE gel (7 M urea), the denatured linear DNA fragments used as a marker, migrate much quicker than circular, covalently closed DNA minicircles in which the two strands cannot separate in space. Notice the presence of only one topoisomer in each category of minicircles and the sensitivity of minicircles with 100 bp to Bal31 nuclease.

    Techniques Used: Polyacrylamide Gel Electrophoresis, Marker

    21) Product Images from "Novel Virulent and Broad-Host-Range Erwinia amylovora Bacteriophages Reveal a High Degree of Mosaicism and a Relationship to Enterobacteriaceae Phages ▿ Phages ▿ †"

    Article Title: Novel Virulent and Broad-Host-Range Erwinia amylovora Bacteriophages Reveal a High Degree of Mosaicism and a Relationship to Enterobacteriaceae Phages ▿ Phages ▿ †

    Journal: Applied and Environmental Microbiology

    doi: 10.1128/AEM.03022-10

    Analysis of phage genome physical structures. Shown are fragment patterns of phage DNA, following time-limited BAL31 treatment and subsequent digestion with different endonucleases. Phage L1 was digested with MfeI, M7 with FspBI, S6 with three different
    Figure Legend Snippet: Analysis of phage genome physical structures. Shown are fragment patterns of phage DNA, following time-limited BAL31 treatment and subsequent digestion with different endonucleases. Phage L1 was digested with MfeI, M7 with FspBI, S6 with three different

    Techniques Used:

    22) Product Images from "Cooperative kinking at distant sites in mechanically stressed DNA"

    Article Title: Cooperative kinking at distant sites in mechanically stressed DNA

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkr666

    DNA minicircles with 100, 106 and 108 bp, their purity and their susceptibility to Bal31 digestion. In a 10% denaturing PAGE gel (7 M urea), the denatured linear DNA fragments used as a marker, migrate much quicker than circular, covalently closed DNA minicircles in which the two strands cannot separate in space. Notice the presence of only one topoisomer in each category of minicircles and the sensitivity of minicircles with 100 bp to Bal31 nuclease.
    Figure Legend Snippet: DNA minicircles with 100, 106 and 108 bp, their purity and their susceptibility to Bal31 digestion. In a 10% denaturing PAGE gel (7 M urea), the denatured linear DNA fragments used as a marker, migrate much quicker than circular, covalently closed DNA minicircles in which the two strands cannot separate in space. Notice the presence of only one topoisomer in each category of minicircles and the sensitivity of minicircles with 100 bp to Bal31 nuclease.

    Techniques Used: Polyacrylamide Gel Electrophoresis, Marker

    23) Product Images from "Bending modes of DNA directly addressed by cryo-electron microscopy of DNA minicircles"

    Article Title: Bending modes of DNA directly addressed by cryo-electron microscopy of DNA minicircles

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkp137

    The formation and purification of 94-bp-long covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that after the ligation reaction of minicircles with nicks, new species appear on the gel (compare lanes 3 with 2). Only these new species resist digestion by Bal31 (lane 4) indicating that they are covalently closed DNA molecules. The electrophoretic migration of these new species is as expected for monomeric, dimeric and trimeric rings that acquired writhe due to ethidium bromide intercalation to covalently closed DNA molecules.
    Figure Legend Snippet: The formation and purification of 94-bp-long covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that after the ligation reaction of minicircles with nicks, new species appear on the gel (compare lanes 3 with 2). Only these new species resist digestion by Bal31 (lane 4) indicating that they are covalently closed DNA molecules. The electrophoretic migration of these new species is as expected for monomeric, dimeric and trimeric rings that acquired writhe due to ethidium bromide intercalation to covalently closed DNA molecules.

    Techniques Used: Purification, Agarose Gel Electrophoresis, Ligation, Migration

    Bal31 nuclease detects a destabilized duplex DNA structure in covalently closed supercoiled DNA molecules but does not reveal the presence of kinks in 94-bp covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that while negatively supercoiled DNA (lane 6) and nonligated nicked rings are completely digested by Bal31 nuclease (see lanes 2–4), the covalently closed monomeric, dimerc, trimeric and tetrameric circles (indicated with arrows) remain resistant to action of Bal31 nuclease (compare lanes 4 and 3).
    Figure Legend Snippet: Bal31 nuclease detects a destabilized duplex DNA structure in covalently closed supercoiled DNA molecules but does not reveal the presence of kinks in 94-bp covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that while negatively supercoiled DNA (lane 6) and nonligated nicked rings are completely digested by Bal31 nuclease (see lanes 2–4), the covalently closed monomeric, dimerc, trimeric and tetrameric circles (indicated with arrows) remain resistant to action of Bal31 nuclease (compare lanes 4 and 3).

    Techniques Used: Agarose Gel Electrophoresis

    24) Product Images from "T5 Exonuclease Hydrolysis of Hepatitis B Virus Replicative Intermediates Allows Reliable Quantification and Fast Drug Efficacy Testing of Covalently Closed Circular DNA by PCR"

    Article Title: T5 Exonuclease Hydrolysis of Hepatitis B Virus Replicative Intermediates Allows Reliable Quantification and Fast Drug Efficacy Testing of Covalently Closed Circular DNA by PCR

    Journal: Journal of Virology

    doi: 10.1128/JVI.01117-18

    Identification of exonucleases selectively digesting rcDNA. (A) Properties of exonucleases tested in this study. +, strong activity; -, no significant activity; +/-, reduced activity; ss, single stranded; ds, double stranded; endo, endonuclease activity; dNMP, deoxyribonucleoside monophosphate; oligos, oligonucleotides. (B) Copies (3 × 10 8 ) of cell culture-derived viral DNA containing rcDNA and dslDNA were incubated for 1 h at 37°C with PSD (5 U), BAL-31 (5 U), Exo I (5 U), Exo V (5 U), and T5 Exo (5 U). Mung bean nuclease (5 U), EcoRI (5 U), and DNase I (5 U) were included as controls. After heat inactivation, the products were subjected to Southern blotting. The plasmid pUCX3.2 served as a marker for indicating the expected sizes of rcDNA and cccDNA.
    Figure Legend Snippet: Identification of exonucleases selectively digesting rcDNA. (A) Properties of exonucleases tested in this study. +, strong activity; -, no significant activity; +/-, reduced activity; ss, single stranded; ds, double stranded; endo, endonuclease activity; dNMP, deoxyribonucleoside monophosphate; oligos, oligonucleotides. (B) Copies (3 × 10 8 ) of cell culture-derived viral DNA containing rcDNA and dslDNA were incubated for 1 h at 37°C with PSD (5 U), BAL-31 (5 U), Exo I (5 U), Exo V (5 U), and T5 Exo (5 U). Mung bean nuclease (5 U), EcoRI (5 U), and DNase I (5 U) were included as controls. After heat inactivation, the products were subjected to Southern blotting. The plasmid pUCX3.2 served as a marker for indicating the expected sizes of rcDNA and cccDNA.

    Techniques Used: Activity Assay, Cell Culture, Derivative Assay, Incubation, Southern Blot, Plasmid Preparation, Marker

    25) Product Images from "Telomere length is highly heritable and independent of growth rate manipulated by temperature in field crickets"

    Article Title: Telomere length is highly heritable and independent of growth rate manipulated by temperature in field crickets

    Journal: bioRxiv

    doi: 10.1101/2020.05.29.123216

    Pulsed field gel electrophoresis showing TTAGG n telomere restriction fragments from the denatured chromosomes of six individual crickets (indicated by the numbers 1-6). XV molecular size ladders are shown on the outer lanes and shorter 1kb ladders are shown between crickets 2-3 and 4-5. The photo shows four aliquots of DNA sample per cricket and these aliquots were exposed to 0 (i.e. untreated), 20, 80, and 240 minutes of Bal 31 . Each time series clearly shows that Bal 31 decreases the molecular size distribution of TTAGG fragments. The absence of a banded distribution in the smears suggest that interstitial telomeric repeats were not present and that chromosome strands were intact. Taken together, these findings show that TTAGG n fragments were located at the chromosome-ends.
    Figure Legend Snippet: Pulsed field gel electrophoresis showing TTAGG n telomere restriction fragments from the denatured chromosomes of six individual crickets (indicated by the numbers 1-6). XV molecular size ladders are shown on the outer lanes and shorter 1kb ladders are shown between crickets 2-3 and 4-5. The photo shows four aliquots of DNA sample per cricket and these aliquots were exposed to 0 (i.e. untreated), 20, 80, and 240 minutes of Bal 31 . Each time series clearly shows that Bal 31 decreases the molecular size distribution of TTAGG fragments. The absence of a banded distribution in the smears suggest that interstitial telomeric repeats were not present and that chromosome strands were intact. Taken together, these findings show that TTAGG n fragments were located at the chromosome-ends.

    Techniques Used: Pulsed-Field Gel, Electrophoresis

    26) Product Images from "Cooperative kinking at distant sites in mechanically stressed DNA"

    Article Title: Cooperative kinking at distant sites in mechanically stressed DNA

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkr666

    DNA minicircles with 100, 106 and 108 bp, their purity and their susceptibility to Bal31 digestion. In a 10% denaturing PAGE gel (7 M urea), the denatured linear DNA fragments used as a marker, migrate much quicker than circular, covalently closed DNA minicircles in which the two strands cannot separate in space. Notice the presence of only one topoisomer in each category of minicircles and the sensitivity of minicircles with 100 bp to Bal31 nuclease.
    Figure Legend Snippet: DNA minicircles with 100, 106 and 108 bp, their purity and their susceptibility to Bal31 digestion. In a 10% denaturing PAGE gel (7 M urea), the denatured linear DNA fragments used as a marker, migrate much quicker than circular, covalently closed DNA minicircles in which the two strands cannot separate in space. Notice the presence of only one topoisomer in each category of minicircles and the sensitivity of minicircles with 100 bp to Bal31 nuclease.

    Techniques Used: Polyacrylamide Gel Electrophoresis, Marker

    27) Product Images from "Novel Virulent and Broad-Host-Range Erwinia amylovora Bacteriophages Reveal a High Degree of Mosaicism and a Relationship to Enterobacteriaceae Phages ▿ Phages ▿ †"

    Article Title: Novel Virulent and Broad-Host-Range Erwinia amylovora Bacteriophages Reveal a High Degree of Mosaicism and a Relationship to Enterobacteriaceae Phages ▿ Phages ▿ †

    Journal: Applied and Environmental Microbiology

    doi: 10.1128/AEM.03022-10

    Analysis of phage genome physical structures. Shown are fragment patterns of phage DNA, following time-limited BAL31 treatment and subsequent digestion with different endonucleases. Phage L1 was digested with MfeI, M7 with FspBI, S6 with three different
    Figure Legend Snippet: Analysis of phage genome physical structures. Shown are fragment patterns of phage DNA, following time-limited BAL31 treatment and subsequent digestion with different endonucleases. Phage L1 was digested with MfeI, M7 with FspBI, S6 with three different

    Techniques Used:

    28) Product Images from "Bending modes of DNA directly addressed by cryo-electron microscopy of DNA minicircles"

    Article Title: Bending modes of DNA directly addressed by cryo-electron microscopy of DNA minicircles

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkp137

    The formation and purification of 94-bp-long covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that after the ligation reaction of minicircles with nicks, new species appear on the gel (compare lanes 3 with 2). Only these new species resist digestion by Bal31 (lane 4) indicating that they are covalently closed DNA molecules. The electrophoretic migration of these new species is as expected for monomeric, dimeric and trimeric rings that acquired writhe due to ethidium bromide intercalation to covalently closed DNA molecules.
    Figure Legend Snippet: The formation and purification of 94-bp-long covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that after the ligation reaction of minicircles with nicks, new species appear on the gel (compare lanes 3 with 2). Only these new species resist digestion by Bal31 (lane 4) indicating that they are covalently closed DNA molecules. The electrophoretic migration of these new species is as expected for monomeric, dimeric and trimeric rings that acquired writhe due to ethidium bromide intercalation to covalently closed DNA molecules.

    Techniques Used: Purification, Agarose Gel Electrophoresis, Ligation, Migration

    Bal31 nuclease detects a destabilized duplex DNA structure in covalently closed supercoiled DNA molecules but does not reveal the presence of kinks in 94-bp covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that while negatively supercoiled DNA (lane 6) and nonligated nicked rings are completely digested by Bal31 nuclease (see lanes 2–4), the covalently closed monomeric, dimerc, trimeric and tetrameric circles (indicated with arrows) remain resistant to action of Bal31 nuclease (compare lanes 4 and 3).
    Figure Legend Snippet: Bal31 nuclease detects a destabilized duplex DNA structure in covalently closed supercoiled DNA molecules but does not reveal the presence of kinks in 94-bp covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that while negatively supercoiled DNA (lane 6) and nonligated nicked rings are completely digested by Bal31 nuclease (see lanes 2–4), the covalently closed monomeric, dimerc, trimeric and tetrameric circles (indicated with arrows) remain resistant to action of Bal31 nuclease (compare lanes 4 and 3).

    Techniques Used: Agarose Gel Electrophoresis

    29) Product Images from "Cooperative kinking at distant sites in mechanically stressed DNA"

    Article Title: Cooperative kinking at distant sites in mechanically stressed DNA

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkr666

    DNA minicircles with 100, 106 and 108 bp, their purity and their susceptibility to Bal31 digestion. In a 10% denaturing PAGE gel (7 M urea), the denatured linear DNA fragments used as a marker, migrate much quicker than circular, covalently closed DNA minicircles in which the two strands cannot separate in space. Notice the presence of only one topoisomer in each category of minicircles and the sensitivity of minicircles with 100 bp to Bal31 nuclease.
    Figure Legend Snippet: DNA minicircles with 100, 106 and 108 bp, their purity and their susceptibility to Bal31 digestion. In a 10% denaturing PAGE gel (7 M urea), the denatured linear DNA fragments used as a marker, migrate much quicker than circular, covalently closed DNA minicircles in which the two strands cannot separate in space. Notice the presence of only one topoisomer in each category of minicircles and the sensitivity of minicircles with 100 bp to Bal31 nuclease.

    Techniques Used: Polyacrylamide Gel Electrophoresis, Marker

    30) Product Images from "Design and characterization of decoy oligonucleotides containing locked nucleic acids"

    Article Title: Design and characterization of decoy oligonucleotides containing locked nucleic acids

    Journal: Nucleic Acids Research

    doi:

    Susceptibility to BAL-31 nuclease degradation of LNA-modified ODNs. LNA-modified [NF-κB(a), (b), (c), (c+b) and (b+c)] and control phosphodiester [NF-κB(d)] decoy molecules were incubated for different lengths of time, as indicated, with 0.5 U/ml BAL-31 and then submitted to electrophoretic separation on 2.5% (w/v) agarose gels. Detection and quantitation of the ethidium bromide stained bands were performed in a Molecular Analyst. Volume densities of the bands are expressed as percent ODN decoy remaining with respect to the relative time zero value and are shown as line graphs.
    Figure Legend Snippet: Susceptibility to BAL-31 nuclease degradation of LNA-modified ODNs. LNA-modified [NF-κB(a), (b), (c), (c+b) and (b+c)] and control phosphodiester [NF-κB(d)] decoy molecules were incubated for different lengths of time, as indicated, with 0.5 U/ml BAL-31 and then submitted to electrophoretic separation on 2.5% (w/v) agarose gels. Detection and quantitation of the ethidium bromide stained bands were performed in a Molecular Analyst. Volume densities of the bands are expressed as percent ODN decoy remaining with respect to the relative time zero value and are shown as line graphs.

    Techniques Used: Modification, Incubation, Quantitation Assay, Staining

    31) Product Images from "The mitochondrial genome of the pathogenic yeast Candida subhashii: GC-rich linear DNA with a protein covalently attached to the 5? termini"

    Article Title: The mitochondrial genome of the pathogenic yeast Candida subhashii: GC-rich linear DNA with a protein covalently attached to the 5? termini

    Journal: Microbiology

    doi: 10.1099/mic.0.038646-0

    The termini of the linear mtDNA are bound by a protein. (a) DNA samples were prepared from C. subhashii (see Methods) and separated by PFGE in a 1.5 % agarose gel. The gel was stained with 0.5 μg ml −1 ethidium bromide (EtBr) and transferred onto a nylon membrane. The blot was hybridized with radioactively labelled mtDNA from C. subhashii . Lane 1, isolated mtDNA; lanes 2 and 3, total cellular DNA prepared in agarose blocks treated or untreated with proteinase K, respectively. (b) Approximately 1 μg of isolated mtDNA was treated with exonuclease III (ExoIII) (left panel) or BAL-31 nuclease (right panel), as indicated. The mtDNA was then extracted from reactions, digested with Xba I endonuclease and electrophoretically separated. Note that the terminal fragments are sensitive to ExoIII but apparently not to BAL-31, indicating the possibility that the linear molecules have their 5′ termini blocked. L and R, positions of the 1527 and 2803 bp terminal restriction enzyme fragments, respectively; C, position of the internal control (a 1040 bp long linear blunt-ended DNA fragment) mixed with mtDNA prior to digestion with BAL-31 nuclease. (c) The mtDNA–protein complexes were isolated as described in Methods, digested with restriction endonucleases Cla I or Pvu I, and treated or not treated with proteinase K. The positions of terminal restriction enzyme fragments generated by Pvu I (833 and 2339 bp) and Cla I (547 bp) are indicated as L, R and L+R, respectively. Note that both terminal Cla I fragments have identical sizes, as this enzyme digests the C. subhashii mtDNA within TIRs.
    Figure Legend Snippet: The termini of the linear mtDNA are bound by a protein. (a) DNA samples were prepared from C. subhashii (see Methods) and separated by PFGE in a 1.5 % agarose gel. The gel was stained with 0.5 μg ml −1 ethidium bromide (EtBr) and transferred onto a nylon membrane. The blot was hybridized with radioactively labelled mtDNA from C. subhashii . Lane 1, isolated mtDNA; lanes 2 and 3, total cellular DNA prepared in agarose blocks treated or untreated with proteinase K, respectively. (b) Approximately 1 μg of isolated mtDNA was treated with exonuclease III (ExoIII) (left panel) or BAL-31 nuclease (right panel), as indicated. The mtDNA was then extracted from reactions, digested with Xba I endonuclease and electrophoretically separated. Note that the terminal fragments are sensitive to ExoIII but apparently not to BAL-31, indicating the possibility that the linear molecules have their 5′ termini blocked. L and R, positions of the 1527 and 2803 bp terminal restriction enzyme fragments, respectively; C, position of the internal control (a 1040 bp long linear blunt-ended DNA fragment) mixed with mtDNA prior to digestion with BAL-31 nuclease. (c) The mtDNA–protein complexes were isolated as described in Methods, digested with restriction endonucleases Cla I or Pvu I, and treated or not treated with proteinase K. The positions of terminal restriction enzyme fragments generated by Pvu I (833 and 2339 bp) and Cla I (547 bp) are indicated as L, R and L+R, respectively. Note that both terminal Cla I fragments have identical sizes, as this enzyme digests the C. subhashii mtDNA within TIRs.

    Techniques Used: Agarose Gel Electrophoresis, Staining, Isolation, Generated

    32) Product Images from "Bending modes of DNA directly addressed by cryo-electron microscopy of DNA minicircles"

    Article Title: Bending modes of DNA directly addressed by cryo-electron microscopy of DNA minicircles

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkp137

    The formation and purification of 94-bp-long covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that after the ligation reaction of minicircles with nicks, new species appear on the gel (compare lanes 3 with 2). Only these new species resist digestion by Bal31 (lane 4) indicating that they are covalently closed DNA molecules. The electrophoretic migration of these new species is as expected for monomeric, dimeric and trimeric rings that acquired writhe due to ethidium bromide intercalation to covalently closed DNA molecules.
    Figure Legend Snippet: The formation and purification of 94-bp-long covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that after the ligation reaction of minicircles with nicks, new species appear on the gel (compare lanes 3 with 2). Only these new species resist digestion by Bal31 (lane 4) indicating that they are covalently closed DNA molecules. The electrophoretic migration of these new species is as expected for monomeric, dimeric and trimeric rings that acquired writhe due to ethidium bromide intercalation to covalently closed DNA molecules.

    Techniques Used: Purification, Agarose Gel Electrophoresis, Ligation, Migration

    Bal31 nuclease detects a destabilized duplex DNA structure in covalently closed supercoiled DNA molecules but does not reveal the presence of kinks in 94-bp covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that while negatively supercoiled DNA (lane 6) and nonligated nicked rings are completely digested by Bal31 nuclease (see lanes 2–4), the covalently closed monomeric, dimerc, trimeric and tetrameric circles (indicated with arrows) remain resistant to action of Bal31 nuclease (compare lanes 4 and 3).
    Figure Legend Snippet: Bal31 nuclease detects a destabilized duplex DNA structure in covalently closed supercoiled DNA molecules but does not reveal the presence of kinks in 94-bp covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that while negatively supercoiled DNA (lane 6) and nonligated nicked rings are completely digested by Bal31 nuclease (see lanes 2–4), the covalently closed monomeric, dimerc, trimeric and tetrameric circles (indicated with arrows) remain resistant to action of Bal31 nuclease (compare lanes 4 and 3).

    Techniques Used: Agarose Gel Electrophoresis

    33) Product Images from "A shotgun antisense approach to the identification of novel essential genes in Pseudomonas aeruginosa"

    Article Title: A shotgun antisense approach to the identification of novel essential genes in Pseudomonas aeruginosa

    Journal: BMC Microbiology

    doi: 10.1186/1471-2180-14-24

    Construction and screening of PAO1 SALs. (A) Genomic DNA was isolated from P. aeruginosa PAO1 and nebulized to obtain sheared fragments of 200–800 bp. After treatment with exonuclease BAL-31 and Klenow polymerase, the genomic DNA fragments were cloned into the E. coli strain JM109, downstream of the arabinose-inducible promoter P BAD of the pHERD20T vector. (B) E. coli transformants, representing the PAO1 shotgun antisense library (SAL), were arrayed in 96-well microplates and (C) mated with P. aeruginosa PAO1 in the presence of a helper strain (triparental mating). (D) SAL recipient PAO1 exconjugants were selected by spotting on PIA plates supplemented with Cb both in the absence and in the presence of the P BAD inducer arabinose. Recipient PAO1 exconjugant spots were inspected for growth defects following 24 h of incubation at 37°C. (E) The identity of the genomic fragments eliciting growth defects (lethal effects, indicated by a lack of a spot: only with inducer, e.g. clones A4, A8, B5, and E4, and with and without an inducer, e.g. clones A2 and E6; growth impairment, indicated as gray spots: only with an inducer, e.g. clones C2, A6, and B6, and with and without an inducer, e.g. C3 and B8) was determined by sequencing the inserts in the corresponding clones of E. coli SAL.
    Figure Legend Snippet: Construction and screening of PAO1 SALs. (A) Genomic DNA was isolated from P. aeruginosa PAO1 and nebulized to obtain sheared fragments of 200–800 bp. After treatment with exonuclease BAL-31 and Klenow polymerase, the genomic DNA fragments were cloned into the E. coli strain JM109, downstream of the arabinose-inducible promoter P BAD of the pHERD20T vector. (B) E. coli transformants, representing the PAO1 shotgun antisense library (SAL), were arrayed in 96-well microplates and (C) mated with P. aeruginosa PAO1 in the presence of a helper strain (triparental mating). (D) SAL recipient PAO1 exconjugants were selected by spotting on PIA plates supplemented with Cb both in the absence and in the presence of the P BAD inducer arabinose. Recipient PAO1 exconjugant spots were inspected for growth defects following 24 h of incubation at 37°C. (E) The identity of the genomic fragments eliciting growth defects (lethal effects, indicated by a lack of a spot: only with inducer, e.g. clones A4, A8, B5, and E4, and with and without an inducer, e.g. clones A2 and E6; growth impairment, indicated as gray spots: only with an inducer, e.g. clones C2, A6, and B6, and with and without an inducer, e.g. C3 and B8) was determined by sequencing the inserts in the corresponding clones of E. coli SAL.

    Techniques Used: Isolation, Clone Assay, Plasmid Preparation, Incubation, Sequencing

    34) Product Images from "Structural diversity of supercoiled DNA"

    Article Title: Structural diversity of supercoiled DNA

    Journal: Nature Communications

    doi: 10.1038/ncomms9440

    Mapping Bal-31 cleavage. To determine whether Bal-31 cleavage occurs at multiple sites or at a preferred site, the Δ Lk =−6 topoisomer was cleaved with Bal-31 and various restriction enzymes. ( a ) Products were separated by agarose gel electrophoresis. Left, (lanes 1–5), control reactions, mc336 (approximately equal mixture of Δ Lk =−2 and Δ Lk =−3 topoisomers) with combinations of the various restriction enzymes (as indicated) to generate fragments of known DNA lengths. Right, (lanes 6–9), Δ Lk =−6 topoisomer cleaved first with Bal-31, followed by a restriction enzyme (as indicated). Mr 1 : 100 bp DNA ladder, Mr 2 : Low molecular weight DNA ladder. ( b ) Map of the minicircle sequence showing the positions of the restriction enzymes used, the estimated location of Bal-31 cleavage (with parentheses indicating the range), and the location of the observed base-pair breaking in MD simulation of the Δ Lk =−3 topoisomer.
    Figure Legend Snippet: Mapping Bal-31 cleavage. To determine whether Bal-31 cleavage occurs at multiple sites or at a preferred site, the Δ Lk =−6 topoisomer was cleaved with Bal-31 and various restriction enzymes. ( a ) Products were separated by agarose gel electrophoresis. Left, (lanes 1–5), control reactions, mc336 (approximately equal mixture of Δ Lk =−2 and Δ Lk =−3 topoisomers) with combinations of the various restriction enzymes (as indicated) to generate fragments of known DNA lengths. Right, (lanes 6–9), Δ Lk =−6 topoisomer cleaved first with Bal-31, followed by a restriction enzyme (as indicated). Mr 1 : 100 bp DNA ladder, Mr 2 : Low molecular weight DNA ladder. ( b ) Map of the minicircle sequence showing the positions of the restriction enzymes used, the estimated location of Bal-31 cleavage (with parentheses indicating the range), and the location of the observed base-pair breaking in MD simulation of the Δ Lk =−3 topoisomer.

    Techniques Used: Agarose Gel Electrophoresis, Molecular Weight, Sequencing

    Effect of supercoiling on DNA base accessibility. ( a ) Minicircle DNA incubated with nuclease Bal-31. Over time, samples were removed, quenched by the addition of stop buffer and the products analysed by polyacrylamide gel electrophoresis. Mr: 100 bp DNA ladder, L: linearized 336 bp DNA, N: nicked 336 bp minicircle. ( b ) Graphic representation of the data shown in ( a ) Fitted lines are for visualization purposes only. ( c ) MD simulation of the Δ Lk =−3 topoisomer in explicit solvent. Splayed bases were found at a sharp bend of a needle conformation. This may be a potential atomistic explanation for Bal-31 susceptibility of negatively supercoiled topoisomers.
    Figure Legend Snippet: Effect of supercoiling on DNA base accessibility. ( a ) Minicircle DNA incubated with nuclease Bal-31. Over time, samples were removed, quenched by the addition of stop buffer and the products analysed by polyacrylamide gel electrophoresis. Mr: 100 bp DNA ladder, L: linearized 336 bp DNA, N: nicked 336 bp minicircle. ( b ) Graphic representation of the data shown in ( a ) Fitted lines are for visualization purposes only. ( c ) MD simulation of the Δ Lk =−3 topoisomer in explicit solvent. Splayed bases were found at a sharp bend of a needle conformation. This may be a potential atomistic explanation for Bal-31 susceptibility of negatively supercoiled topoisomers.

    Techniques Used: Incubation, Polyacrylamide Gel Electrophoresis

    35) Product Images from "Telomere length is highly heritable and independent of growth rate manipulated by temperature in field crickets"

    Article Title: Telomere length is highly heritable and independent of growth rate manipulated by temperature in field crickets

    Journal: bioRxiv

    doi: 10.1101/2020.05.29.123216

    Pulsed field gel electrophoresis showing TTAGG n telomere restriction fragments from the denatured chromosomes of six individual crickets (indicated by the numbers 1-6). XV molecular size ladders are shown on the outer lanes and shorter 1kb ladders are shown between crickets 2-3 and 4-5. The photo shows four aliquots of DNA sample per cricket and these aliquots were exposed to 0 (i.e. untreated), 20, 80, and 240 minutes of Bal 31 . Each time series clearly shows that Bal 31 decreases the molecular size distribution of TTAGG fragments. The absence of a banded distribution in the smears suggest that interstitial telomeric repeats were not present and that chromosome strands were intact. Taken together, these findings show that TTAGG n fragments were located at the chromosome-ends.
    Figure Legend Snippet: Pulsed field gel electrophoresis showing TTAGG n telomere restriction fragments from the denatured chromosomes of six individual crickets (indicated by the numbers 1-6). XV molecular size ladders are shown on the outer lanes and shorter 1kb ladders are shown between crickets 2-3 and 4-5. The photo shows four aliquots of DNA sample per cricket and these aliquots were exposed to 0 (i.e. untreated), 20, 80, and 240 minutes of Bal 31 . Each time series clearly shows that Bal 31 decreases the molecular size distribution of TTAGG fragments. The absence of a banded distribution in the smears suggest that interstitial telomeric repeats were not present and that chromosome strands were intact. Taken together, these findings show that TTAGG n fragments were located at the chromosome-ends.

    Techniques Used: Pulsed-Field Gel, Electrophoresis

    36) Product Images from "Evolution of linear chromosomes and multipartite genomes in yeast mitochondria"

    Article Title: Evolution of linear chromosomes and multipartite genomes in yeast mitochondria

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkq1345

    Multipartite linear-mapping genomes in C. labiduridarum and C. frijolesensis ( A ) PFGE separated samples of C. labiduridarum NRRL Y-27940 (lane 1) and C. frijolesensis NRRL Y-48060 (lane 2) were blotted onto a nylon membrane and hybridized with the radioactively labeled probes P-668 and H-1030 (regions hybridizing with both probes are shown as dashed lines). Presumed master (I) and two smaller chromosomes (II and III) are indicated. Note that the master chromosome occurs in four isomers (i.e. L III − R III − L II − R II (shown in the scheme), L III − R III − R II − L II , R III − L III − L II − R II and R III − L III − R II − L II . ‘L’ and ‘R’ indicate the left and the right telomere, respectively). The C. frijolesensis mtDNA (∼1 µg) was digested with BAL-31 nuclease ( B ) or exonuclease III (ExoIII) ( C ) as indicated. After nuclease inactivation, the DNA was digested with EcoRV, separated in 0.9% (w/v) agarose gel. The Southern blots were hybridized with the P-668 and EH-1350 probes specific for the left and the right arm of the master chromosome, respectively (see ‘Materials and Methods’ section). Arrows show the positions of the left (L) and right (R) terminal fragments and their fusions (R + R, R + L and L + L). Note that after ExoIII treatment the telomeric fragments form two subpopulations that differ in their sensitivity to the ExoIII treatment. This indicates that the linear mtDNA molecules possess an open structure with 5′ overhang or blunt end or covalently closed t-hairpin. ( D ) The C. frijolesensis mtDNA was treated with antarctic phosphatase and labeled with [γ 32 P]ATP and T4 polynucleotide kinase. The mtDNA was then digested with restriction endonuclease EcoRV (lane 1) or BglII (lane 2) and separated in 0.8% (w/v) agarose gel (left panel). The gel was fixed in 10% (v/v) methanol/10% (v/v) acetic acid for 30 min, dried overnight and autoradiographed (right panel). Arrows indicate the position of telomeric fragments containing the open structures accessible to terminal labeling.
    Figure Legend Snippet: Multipartite linear-mapping genomes in C. labiduridarum and C. frijolesensis ( A ) PFGE separated samples of C. labiduridarum NRRL Y-27940 (lane 1) and C. frijolesensis NRRL Y-48060 (lane 2) were blotted onto a nylon membrane and hybridized with the radioactively labeled probes P-668 and H-1030 (regions hybridizing with both probes are shown as dashed lines). Presumed master (I) and two smaller chromosomes (II and III) are indicated. Note that the master chromosome occurs in four isomers (i.e. L III − R III − L II − R II (shown in the scheme), L III − R III − R II − L II , R III − L III − L II − R II and R III − L III − R II − L II . ‘L’ and ‘R’ indicate the left and the right telomere, respectively). The C. frijolesensis mtDNA (∼1 µg) was digested with BAL-31 nuclease ( B ) or exonuclease III (ExoIII) ( C ) as indicated. After nuclease inactivation, the DNA was digested with EcoRV, separated in 0.9% (w/v) agarose gel. The Southern blots were hybridized with the P-668 and EH-1350 probes specific for the left and the right arm of the master chromosome, respectively (see ‘Materials and Methods’ section). Arrows show the positions of the left (L) and right (R) terminal fragments and their fusions (R + R, R + L and L + L). Note that after ExoIII treatment the telomeric fragments form two subpopulations that differ in their sensitivity to the ExoIII treatment. This indicates that the linear mtDNA molecules possess an open structure with 5′ overhang or blunt end or covalently closed t-hairpin. ( D ) The C. frijolesensis mtDNA was treated with antarctic phosphatase and labeled with [γ 32 P]ATP and T4 polynucleotide kinase. The mtDNA was then digested with restriction endonuclease EcoRV (lane 1) or BglII (lane 2) and separated in 0.8% (w/v) agarose gel (left panel). The gel was fixed in 10% (v/v) methanol/10% (v/v) acetic acid for 30 min, dried overnight and autoradiographed (right panel). Arrows indicate the position of telomeric fragments containing the open structures accessible to terminal labeling.

    Techniques Used: Labeling, Agarose Gel Electrophoresis

    Circular- and linear-mapping genome isomers in mitochondria of C. viswanathii. ( A ) The mtDNA samples were digested with BamHI (lane 1) or Eco91I (lane 2) and separated in 1% (w/v) agarose gel. The Southern blot was hybridized with radioactively labeled oligonucleotide probe Oligo-32 derived from the large palindrome (shown as dashed arrows). The solid arrows show positions of the palindrome and the presumed terminal fragments of resolved linear molecules capped with t-hairpins. Scheme shows presumed circular- (I) and linear-mapping (II) genome isomers. ( B ) Isolated mtDNA was treated or untreated with BAL-31 nuclease (0.2 U for 5 min). The mtDNA was then extracted from the reaction, digested with BamHI or Eco91I endonuclease, and electrophoretically separated. Note that the fragments containing presumed t-hairpins were sensitive to BAL-31 nuclease (indicated by asterisk).
    Figure Legend Snippet: Circular- and linear-mapping genome isomers in mitochondria of C. viswanathii. ( A ) The mtDNA samples were digested with BamHI (lane 1) or Eco91I (lane 2) and separated in 1% (w/v) agarose gel. The Southern blot was hybridized with radioactively labeled oligonucleotide probe Oligo-32 derived from the large palindrome (shown as dashed arrows). The solid arrows show positions of the palindrome and the presumed terminal fragments of resolved linear molecules capped with t-hairpins. Scheme shows presumed circular- (I) and linear-mapping (II) genome isomers. ( B ) Isolated mtDNA was treated or untreated with BAL-31 nuclease (0.2 U for 5 min). The mtDNA was then extracted from the reaction, digested with BamHI or Eco91I endonuclease, and electrophoretically separated. Note that the fragments containing presumed t-hairpins were sensitive to BAL-31 nuclease (indicated by asterisk).

    Techniques Used: Agarose Gel Electrophoresis, Southern Blot, Labeling, Derivative Assay, Isolation

    37) Product Images from "Selaginella moellendorffii telomeres: conserved and unique features in an ancient land plant lineage"

    Article Title: Selaginella moellendorffii telomeres: conserved and unique features in an ancient land plant lineage

    Journal: Frontiers in Plant Science

    doi: 10.3389/fpls.2012.00161

    Telomere length analysis in Selaginella moellendorffii . (A) Comparative terminal restriction fragment (TRF) analysis of S. moellendorffii (lane 1) and A. thaliana (lane 2) telomeres. Molecular weight markers are shown on the left. (B) Bal 31 digestion of S. moellendorffii telomeric DNA. Lane 1, Tru 1I digestion of genomic DNA without prior Bal treatment (0 min). Lanes 2–6, Tru 1I digestion of genomic DNA with Bal 31 tr31eatment for 15, 30, 45, 60, and 90 min, respectively. Asterisks indicate cross-hybridizing interstitial telomeric DNA bands, which are not sensitive to Bal 31 digestion for up to 90 min.
    Figure Legend Snippet: Telomere length analysis in Selaginella moellendorffii . (A) Comparative terminal restriction fragment (TRF) analysis of S. moellendorffii (lane 1) and A. thaliana (lane 2) telomeres. Molecular weight markers are shown on the left. (B) Bal 31 digestion of S. moellendorffii telomeric DNA. Lane 1, Tru 1I digestion of genomic DNA without prior Bal treatment (0 min). Lanes 2–6, Tru 1I digestion of genomic DNA with Bal 31 tr31eatment for 15, 30, 45, 60, and 90 min, respectively. Asterisks indicate cross-hybridizing interstitial telomeric DNA bands, which are not sensitive to Bal 31 digestion for up to 90 min.

    Techniques Used: Molecular Weight

    38) Product Images from "Bending modes of DNA directly addressed by cryo-electron microscopy of DNA minicircles"

    Article Title: Bending modes of DNA directly addressed by cryo-electron microscopy of DNA minicircles

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkp137

    The formation and purification of 94-bp-long covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that after the ligation reaction of minicircles with nicks, new species appear on the gel (compare lanes 3 with 2). Only these new species resist digestion by Bal31 (lane 4) indicating that they are covalently closed DNA molecules. The electrophoretic migration of these new species is as expected for monomeric, dimeric and trimeric rings that acquired writhe due to ethidium bromide intercalation to covalently closed DNA molecules.
    Figure Legend Snippet: The formation and purification of 94-bp-long covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that after the ligation reaction of minicircles with nicks, new species appear on the gel (compare lanes 3 with 2). Only these new species resist digestion by Bal31 (lane 4) indicating that they are covalently closed DNA molecules. The electrophoretic migration of these new species is as expected for monomeric, dimeric and trimeric rings that acquired writhe due to ethidium bromide intercalation to covalently closed DNA molecules.

    Techniques Used: Purification, Agarose Gel Electrophoresis, Ligation, Migration

    Bal31 nuclease detects a destabilized duplex DNA structure in covalently closed supercoiled DNA molecules but does not reveal the presence of kinks in 94-bp covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that while negatively supercoiled DNA (lane 6) and nonligated nicked rings are completely digested by Bal31 nuclease (see lanes 2–4), the covalently closed monomeric, dimerc, trimeric and tetrameric circles (indicated with arrows) remain resistant to action of Bal31 nuclease (compare lanes 4 and 3).
    Figure Legend Snippet: Bal31 nuclease detects a destabilized duplex DNA structure in covalently closed supercoiled DNA molecules but does not reveal the presence of kinks in 94-bp covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that while negatively supercoiled DNA (lane 6) and nonligated nicked rings are completely digested by Bal31 nuclease (see lanes 2–4), the covalently closed monomeric, dimerc, trimeric and tetrameric circles (indicated with arrows) remain resistant to action of Bal31 nuclease (compare lanes 4 and 3).

    Techniques Used: Agarose Gel Electrophoresis

    39) Product Images from "Brochothrix thermosphacta Bacteriophages Feature Heterogeneous and Highly Mosaic Genomes and Utilize Unique Prophage Insertion Sites ▿ Bacteriophages Feature Heterogeneous and Highly Mosaic Genomes and Utilize Unique Prophage Insertion Sites ▿ †"

    Article Title: Brochothrix thermosphacta Bacteriophages Feature Heterogeneous and Highly Mosaic Genomes and Utilize Unique Prophage Insertion Sites ▿ Bacteriophages Feature Heterogeneous and Highly Mosaic Genomes and Utilize Unique Prophage Insertion Sites ▿ †

    Journal: Journal of Bacteriology

    doi: 10.1128/JB.00709-10

    Structure analysis of the DNA molecules of phage A9, NF5, and BL3. (A) PFGE of full-length phage DNA. Size markers: λmix, lambda mix marker 19 (Fermentas); M1 and M2, PFG midrange markers I and II (NEB). BL3 and NF5 genomic DNAs run at slightly larger sizes than their unit genome lengths of 41.3 and 37 kb. A9 DNA exhibits approximately the same physical size as Listeria ). (B) Gel electrophoresis of PacI-digested A9 DNA after Bal31 nuclease treatment for the indicated time intervals. Terminal fragments of the nonpermuted genomes disappear over time (indicated by arrows). (C) Restriction enzyme profiles of PstI-digested NF5 DNA and SwaI-digested BL3 DNA; the putative pac fragments are indicated by black arrowheads. (D and E) EcoRI (NF5)- and Van91I (BL3)-digested genomic DNA after Bal31 nuclease treatment for the indicated time intervals. Arrows with square heads indicate restriction fragments which were not predicted in silico from a circular molecule ( pac fragment, 4.2 kb; smeary band, > 10 kb). The “submolar” pac fragment disappears after 10 min of Bal31 treatment. Arrows with pointed heads indicate restriction fragments “downstream” from the putative pac site-containing fragment. Their consecutive appearance within the respective genome is indicated by numbering and correlates with the order of disappearance, indicating incomplete permutation of the DNA molecules based on the limited number of genomes in each replication concatemer.
    Figure Legend Snippet: Structure analysis of the DNA molecules of phage A9, NF5, and BL3. (A) PFGE of full-length phage DNA. Size markers: λmix, lambda mix marker 19 (Fermentas); M1 and M2, PFG midrange markers I and II (NEB). BL3 and NF5 genomic DNAs run at slightly larger sizes than their unit genome lengths of 41.3 and 37 kb. A9 DNA exhibits approximately the same physical size as Listeria ). (B) Gel electrophoresis of PacI-digested A9 DNA after Bal31 nuclease treatment for the indicated time intervals. Terminal fragments of the nonpermuted genomes disappear over time (indicated by arrows). (C) Restriction enzyme profiles of PstI-digested NF5 DNA and SwaI-digested BL3 DNA; the putative pac fragments are indicated by black arrowheads. (D and E) EcoRI (NF5)- and Van91I (BL3)-digested genomic DNA after Bal31 nuclease treatment for the indicated time intervals. Arrows with square heads indicate restriction fragments which were not predicted in silico from a circular molecule ( pac fragment, 4.2 kb; smeary band, > 10 kb). The “submolar” pac fragment disappears after 10 min of Bal31 treatment. Arrows with pointed heads indicate restriction fragments “downstream” from the putative pac site-containing fragment. Their consecutive appearance within the respective genome is indicated by numbering and correlates with the order of disappearance, indicating incomplete permutation of the DNA molecules based on the limited number of genomes in each replication concatemer.

    Techniques Used: Marker, Nucleic Acid Electrophoresis, In Silico

    40) Product Images from "Production of DNA minicircles less than 250 base pairs through a novel concentrated DNA circularization assay enabling minicircle design with NF-κB inhibition activity"

    Article Title: Production of DNA minicircles less than 250 base pairs through a novel concentrated DNA circularization assay enabling minicircle design with NF-κB inhibition activity

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkw1034

    CNDCA-based production of supercoiled minicircle. Image of stained native polyacrylamide gel showing 95 bp minicircle topoisomers of decreased linking number (ΔLk values of −1 and −2) (lanes 4, 6) as indicated on the right and obtained by religation of the nicked minicircle in the presence of EtBr. The slowest migrating band corresponds to the relaxed minicircle that migrates expectedly at the same rate as the nicked minicircle (lanes 1 and 2). Digestion of minicircle samples by the nuclease Bal31 is indicated on the right. The position of the molecular mass markers in bp is indicated on the left.
    Figure Legend Snippet: CNDCA-based production of supercoiled minicircle. Image of stained native polyacrylamide gel showing 95 bp minicircle topoisomers of decreased linking number (ΔLk values of −1 and −2) (lanes 4, 6) as indicated on the right and obtained by religation of the nicked minicircle in the presence of EtBr. The slowest migrating band corresponds to the relaxed minicircle that migrates expectedly at the same rate as the nicked minicircle (lanes 1 and 2). Digestion of minicircle samples by the nuclease Bal31 is indicated on the right. The position of the molecular mass markers in bp is indicated on the left.

    Techniques Used: Staining

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    New England Biolabs bal 31 nuclease assay
    The character of hybridization signals revealed using Southern hybridization. The DIG-labeled probe specific to (TTAGG) n ( a ) and (TCAGG)n ( b ) sequence was hybridized with RsaI/HinfI-digested genomic DNAs of selected coleopteran species. ( c ) <t>Bal</t> 31 exonuclease digestion of genomic DNA of Anoplotrupes stercorosus and Clerus muttilarius. Time-course digestion intervals are indicated above the lanes.
    Bal 31 Nuclease Assay, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    The character of hybridization signals revealed using Southern hybridization. The DIG-labeled probe specific to (TTAGG) n ( a ) and (TCAGG)n ( b ) sequence was hybridized with RsaI/HinfI-digested genomic DNAs of selected coleopteran species. ( c ) Bal 31 exonuclease digestion of genomic DNA of Anoplotrupes stercorosus and Clerus muttilarius. Time-course digestion intervals are indicated above the lanes.

    Journal: Scientific Reports

    Article Title: Telomeric DNA sequences in beetle taxa vary with species richness

    doi: 10.1038/s41598-021-92705-y

    Figure Lengend Snippet: The character of hybridization signals revealed using Southern hybridization. The DIG-labeled probe specific to (TTAGG) n ( a ) and (TCAGG)n ( b ) sequence was hybridized with RsaI/HinfI-digested genomic DNAs of selected coleopteran species. ( c ) Bal 31 exonuclease digestion of genomic DNA of Anoplotrupes stercorosus and Clerus muttilarius. Time-course digestion intervals are indicated above the lanes.

    Article Snippet: Bal 31 nuclease assay To confirm the terminal positions of tested sequences on chromosome ends, the genomic DNA was subjected to BAL 31 exonuclease.

    Techniques: Hybridization, Labeling, Sequencing

    CNDCA-based production of supercoiled minicircle. Image of stained native polyacrylamide gel showing 95 bp minicircle topoisomers of decreased linking number (ΔLk values of −1 and −2) (lanes 4, 6) as indicated on the right and obtained by religation of the nicked minicircle in the presence of EtBr. The slowest migrating band corresponds to the relaxed minicircle that migrates expectedly at the same rate as the nicked minicircle (lanes 1 and 2). Digestion of minicircle samples by the nuclease Bal31 is indicated on the right. The position of the molecular mass markers in bp is indicated on the left.

    Journal: Nucleic Acids Research

    Article Title: Production of DNA minicircles less than 250 base pairs through a novel concentrated DNA circularization assay enabling minicircle design with NF-κB inhibition activity

    doi: 10.1093/nar/gkw1034

    Figure Lengend Snippet: CNDCA-based production of supercoiled minicircle. Image of stained native polyacrylamide gel showing 95 bp minicircle topoisomers of decreased linking number (ΔLk values of −1 and −2) (lanes 4, 6) as indicated on the right and obtained by religation of the nicked minicircle in the presence of EtBr. The slowest migrating band corresponds to the relaxed minicircle that migrates expectedly at the same rate as the nicked minicircle (lanes 1 and 2). Digestion of minicircle samples by the nuclease Bal31 is indicated on the right. The position of the molecular mass markers in bp is indicated on the left.

    Article Snippet: We next used Bal31 nuclease to detect the presence of DNA distortions within our constrained minicircles.

    Techniques: Staining

    Susceptibility to BAL-31 nuclease degradation of LNA-modified ODNs. LNA-modified [NF-κB(a), (b), (c), (c+b) and (b+c)] and control phosphodiester [NF-κB(d)] decoy molecules were incubated for different lengths of time, as indicated, with 0.5 U/ml BAL-31 and then submitted to electrophoretic separation on 2.5% (w/v) agarose gels. Detection and quantitation of the ethidium bromide stained bands were performed in a Molecular Analyst. Volume densities of the bands are expressed as percent ODN decoy remaining with respect to the relative time zero value and are shown as line graphs.

    Journal: Nucleic Acids Research

    Article Title: Design and characterization of decoy oligonucleotides containing locked nucleic acids

    doi:

    Figure Lengend Snippet: Susceptibility to BAL-31 nuclease degradation of LNA-modified ODNs. LNA-modified [NF-κB(a), (b), (c), (c+b) and (b+c)] and control phosphodiester [NF-κB(d)] decoy molecules were incubated for different lengths of time, as indicated, with 0.5 U/ml BAL-31 and then submitted to electrophoretic separation on 2.5% (w/v) agarose gels. Detection and quantitation of the ethidium bromide stained bands were performed in a Molecular Analyst. Volume densities of the bands are expressed as percent ODN decoy remaining with respect to the relative time zero value and are shown as line graphs.

    Article Snippet: Double-stranded DNA and LNA ODNs (2.7 µM) were incubated either with 10 U/ml DNase I (EC 3.1.21.1) (Roche Diagnostics) in 50 mM Tris–HCl, pH 7.5, 1 mM MgCl2 and 0.1 mg/ml bovine serum albumin at 20°C or with 10 U/ml BAL-31 nuclease (New England Biolabs, Hitchin, UK) in a reaction buffer consisting of 0.6 M NaCl, 20 mM Tris–HCl, pH 8.0, 12 mM CaCl2 , 12 mM MgCl2 and 1 mM EDTA at 30°C.

    Techniques: Modification, Incubation, Quantitation Assay, Staining

    The formation and purification of 94-bp-long covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that after the ligation reaction of minicircles with nicks, new species appear on the gel (compare lanes 3 with 2). Only these new species resist digestion by Bal31 (lane 4) indicating that they are covalently closed DNA molecules. The electrophoretic migration of these new species is as expected for monomeric, dimeric and trimeric rings that acquired writhe due to ethidium bromide intercalation to covalently closed DNA molecules.

    Journal: Nucleic Acids Research

    Article Title: Bending modes of DNA directly addressed by cryo-electron microscopy of DNA minicircles

    doi: 10.1093/nar/gkp137

    Figure Lengend Snippet: The formation and purification of 94-bp-long covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that after the ligation reaction of minicircles with nicks, new species appear on the gel (compare lanes 3 with 2). Only these new species resist digestion by Bal31 (lane 4) indicating that they are covalently closed DNA molecules. The electrophoretic migration of these new species is as expected for monomeric, dimeric and trimeric rings that acquired writhe due to ethidium bromide intercalation to covalently closed DNA molecules.

    Article Snippet: Bal31 nuclease does not reveal the presence of kinks in covalently closed DNA minicircles As described in ‘Materials and Methods’ section, we have used Bal31 nuclease to purify covalently closed DNA minicircles from nicked circles that were incomplete products of the ligation reactions.

    Techniques: Purification, Agarose Gel Electrophoresis, Ligation, Migration

    Bal31 nuclease detects a destabilized duplex DNA structure in covalently closed supercoiled DNA molecules but does not reveal the presence of kinks in 94-bp covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that while negatively supercoiled DNA (lane 6) and nonligated nicked rings are completely digested by Bal31 nuclease (see lanes 2–4), the covalently closed monomeric, dimerc, trimeric and tetrameric circles (indicated with arrows) remain resistant to action of Bal31 nuclease (compare lanes 4 and 3).

    Journal: Nucleic Acids Research

    Article Title: Bending modes of DNA directly addressed by cryo-electron microscopy of DNA minicircles

    doi: 10.1093/nar/gkp137

    Figure Lengend Snippet: Bal31 nuclease detects a destabilized duplex DNA structure in covalently closed supercoiled DNA molecules but does not reveal the presence of kinks in 94-bp covalently closed DNA minicircles. A 2.5% agarose gel run in the presence of ethidium bromide (0.5 µg/ml) reveals that while negatively supercoiled DNA (lane 6) and nonligated nicked rings are completely digested by Bal31 nuclease (see lanes 2–4), the covalently closed monomeric, dimerc, trimeric and tetrameric circles (indicated with arrows) remain resistant to action of Bal31 nuclease (compare lanes 4 and 3).

    Article Snippet: Bal31 nuclease does not reveal the presence of kinks in covalently closed DNA minicircles As described in ‘Materials and Methods’ section, we have used Bal31 nuclease to purify covalently closed DNA minicircles from nicked circles that were incomplete products of the ligation reactions.

    Techniques: Agarose Gel Electrophoresis