exo iii  (New England Biolabs)


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    Name:
    Exonuclease III
    Description:

    Catalog Number:
    M0206
    Price:
    252
    Category:
    DNA Modifying Enzymes
    Applications:
    DNA Manipulation
    Size:
    25000 units
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    Structured Review

    New England Biolabs exo iii
    Exonuclease III

    https://www.bioz.com/result/exo iii/product/New England Biolabs
    Average 97 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    exo iii - by Bioz Stars, 2021-09
    97/100 stars

    Images

    1) Product Images from "The replication of plastid minicircles involves rolling circle intermediates"

    Article Title: The replication of plastid minicircles involves rolling circle intermediates

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkp063

    Exo III-treated minicircular DNA revealed after PFGE and 2D-gel electrophoresis. Exo III-treated whole-DNA extract from H. triquetra was subjected to PFGE ( A ) and 2D-gel ( B ), and detected with 1.1-kb psbA NCR fragment [and psbA gene fragment in (B) only]. (A) The 6–8 kb minicircular DNA bands (APBs) were removed in the Exo III lane, while the putative 3-kb psbA minicircle band was diminished in intensity. (B) Hybridization signals of larger than 2 kb were removed after Exo III treatment as revealed in both 2D-gels detected by psbA NCR and gene fragments. Extra spots indicated by the arrows were observed in these two images when comparing with the controls. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).
    Figure Legend Snippet: Exo III-treated minicircular DNA revealed after PFGE and 2D-gel electrophoresis. Exo III-treated whole-DNA extract from H. triquetra was subjected to PFGE ( A ) and 2D-gel ( B ), and detected with 1.1-kb psbA NCR fragment [and psbA gene fragment in (B) only]. (A) The 6–8 kb minicircular DNA bands (APBs) were removed in the Exo III lane, while the putative 3-kb psbA minicircle band was diminished in intensity. (B) Hybridization signals of larger than 2 kb were removed after Exo III treatment as revealed in both 2D-gels detected by psbA NCR and gene fragments. Extra spots indicated by the arrows were observed in these two images when comparing with the controls. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).

    Techniques Used: Two-Dimensional Gel Electrophoresis, Electrophoresis, Hybridization

    Effects of DNA ligase/Klenow fragment on minicircular DNA. Total DNA from H. triquetra was treated with T4 DNA ligase, as well as Klenow fragment followed with T4 DNA ligase, prior to Exo III digestion. Treated DNAs were resolved by PFGE and detected the minicircular DNA signals by Southern Blot. The two upper arrows indicated APBs in the untreated control lane. The two lower arrows indicated the putative monomer of psbA minicircle signals of ∼2–3 kb. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).
    Figure Legend Snippet: Effects of DNA ligase/Klenow fragment on minicircular DNA. Total DNA from H. triquetra was treated with T4 DNA ligase, as well as Klenow fragment followed with T4 DNA ligase, prior to Exo III digestion. Treated DNAs were resolved by PFGE and detected the minicircular DNA signals by Southern Blot. The two upper arrows indicated APBs in the untreated control lane. The two lower arrows indicated the putative monomer of psbA minicircle signals of ∼2–3 kb. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).

    Techniques Used: Southern Blot

    2) Product Images from "The replication of plastid minicircles involves rolling circle intermediates"

    Article Title: The replication of plastid minicircles involves rolling circle intermediates

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkp063

    Exo III-treated minicircular DNA revealed after PFGE and 2D-gel electrophoresis. Exo III-treated whole-DNA extract from H. triquetra was subjected to PFGE ( A ) and 2D-gel ( B ), and detected with 1.1-kb psbA NCR fragment [and psbA gene fragment in (B) only]. (A) The 6–8 kb minicircular DNA bands (APBs) were removed in the Exo III lane, while the putative 3-kb psbA minicircle band was diminished in intensity. (B) Hybridization signals of larger than 2 kb were removed after Exo III treatment as revealed in both 2D-gels detected by psbA NCR and gene fragments. Extra spots indicated by the arrows were observed in these two images when comparing with the controls. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).
    Figure Legend Snippet: Exo III-treated minicircular DNA revealed after PFGE and 2D-gel electrophoresis. Exo III-treated whole-DNA extract from H. triquetra was subjected to PFGE ( A ) and 2D-gel ( B ), and detected with 1.1-kb psbA NCR fragment [and psbA gene fragment in (B) only]. (A) The 6–8 kb minicircular DNA bands (APBs) were removed in the Exo III lane, while the putative 3-kb psbA minicircle band was diminished in intensity. (B) Hybridization signals of larger than 2 kb were removed after Exo III treatment as revealed in both 2D-gels detected by psbA NCR and gene fragments. Extra spots indicated by the arrows were observed in these two images when comparing with the controls. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).

    Techniques Used: Two-Dimensional Gel Electrophoresis, Electrophoresis, Hybridization

    Effects of DNA ligase/Klenow fragment on minicircular DNA. Total DNA from H. triquetra was treated with T4 DNA ligase, as well as Klenow fragment followed with T4 DNA ligase, prior to Exo III digestion. Treated DNAs were resolved by PFGE and detected the minicircular DNA signals by Southern Blot. The two upper arrows indicated APBs in the untreated control lane. The two lower arrows indicated the putative monomer of psbA minicircle signals of ∼2–3 kb. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).
    Figure Legend Snippet: Effects of DNA ligase/Klenow fragment on minicircular DNA. Total DNA from H. triquetra was treated with T4 DNA ligase, as well as Klenow fragment followed with T4 DNA ligase, prior to Exo III digestion. Treated DNAs were resolved by PFGE and detected the minicircular DNA signals by Southern Blot. The two upper arrows indicated APBs in the untreated control lane. The two lower arrows indicated the putative monomer of psbA minicircle signals of ∼2–3 kb. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).

    Techniques Used: Southern Blot

    3) Product Images from "The replication of plastid minicircles involves rolling circle intermediates"

    Article Title: The replication of plastid minicircles involves rolling circle intermediates

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkp063

    Exo III-treated minicircular DNA revealed after PFGE and 2D-gel electrophoresis. Exo III-treated whole-DNA extract from H. triquetra was subjected to PFGE ( A ) and 2D-gel ( B ), and detected with 1.1-kb psbA NCR fragment [and psbA gene fragment in (B) only]. (A) The 6–8 kb minicircular DNA bands (APBs) were removed in the Exo III lane, while the putative 3-kb psbA minicircle band was diminished in intensity. (B) Hybridization signals of larger than 2 kb were removed after Exo III treatment as revealed in both 2D-gels detected by psbA NCR and gene fragments. Extra spots indicated by the arrows were observed in these two images when comparing with the controls. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).
    Figure Legend Snippet: Exo III-treated minicircular DNA revealed after PFGE and 2D-gel electrophoresis. Exo III-treated whole-DNA extract from H. triquetra was subjected to PFGE ( A ) and 2D-gel ( B ), and detected with 1.1-kb psbA NCR fragment [and psbA gene fragment in (B) only]. (A) The 6–8 kb minicircular DNA bands (APBs) were removed in the Exo III lane, while the putative 3-kb psbA minicircle band was diminished in intensity. (B) Hybridization signals of larger than 2 kb were removed after Exo III treatment as revealed in both 2D-gels detected by psbA NCR and gene fragments. Extra spots indicated by the arrows were observed in these two images when comparing with the controls. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).

    Techniques Used: Two-Dimensional Gel Electrophoresis, Electrophoresis, Hybridization

    Effects of DNA ligase/Klenow fragment on minicircular DNA. Total DNA from H. triquetra was treated with T4 DNA ligase, as well as Klenow fragment followed with T4 DNA ligase, prior to Exo III digestion. Treated DNAs were resolved by PFGE and detected the minicircular DNA signals by Southern Blot. The two upper arrows indicated APBs in the untreated control lane. The two lower arrows indicated the putative monomer of psbA minicircle signals of ∼2–3 kb. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).
    Figure Legend Snippet: Effects of DNA ligase/Klenow fragment on minicircular DNA. Total DNA from H. triquetra was treated with T4 DNA ligase, as well as Klenow fragment followed with T4 DNA ligase, prior to Exo III digestion. Treated DNAs were resolved by PFGE and detected the minicircular DNA signals by Southern Blot. The two upper arrows indicated APBs in the untreated control lane. The two lower arrows indicated the putative monomer of psbA minicircle signals of ∼2–3 kb. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).

    Techniques Used: Southern Blot

    4) Product Images from "Sensitive and Label-free Detection of Bacteria in Osteomyelitis through Exo III-Assisted Cascade Signal Amplification"

    Article Title: Sensitive and Label-free Detection of Bacteria in Osteomyelitis through Exo III-Assisted Cascade Signal Amplification

    Journal: ACS Omega

    doi: 10.1021/acsomega.1c01107

    Optimization of the whole sensing system. (a) Obtained fluorescence intensity of the sensing system when incubated with Exo III with different concentrations. (b) Fluorescence intensity of the sensing system with different concentrations of the primer. (c) Fluorescence intensity of the sensing system with different concentrations of circular DNA. (d) Fluorescence intensity of the sensing system with different concentrations of NMM.
    Figure Legend Snippet: Optimization of the whole sensing system. (a) Obtained fluorescence intensity of the sensing system when incubated with Exo III with different concentrations. (b) Fluorescence intensity of the sensing system with different concentrations of the primer. (c) Fluorescence intensity of the sensing system with different concentrations of circular DNA. (d) Fluorescence intensity of the sensing system with different concentrations of NMM.

    Techniques Used: Fluorescence, Incubation

    5) Product Images from "An Exonuclease III Protection-Based Electrochemical Method for Estrogen Receptor Assay"

    Article Title: An Exonuclease III Protection-Based Electrochemical Method for Estrogen Receptor Assay

    Journal: International Journal of Molecular Sciences

    doi: 10.3390/ijms140510298

    Square wave voltammograms obtained at the gold electrode immobilized with the DNA strands. ( A ) In the absence of ER, (a) before, and (b) after the DNA strands are digested by Exo III; ( B ) In the presence of 100 nM ER, (a) before, and (b) after the DNA strands are digested by Exo III. Curves c–e are for the control experiments by using 500 nM bovine serum albumin (BSA), thrombin and α-fetoprotein (AFP) instead of 100 nM ER. Buffer: 10 mM phosphate-buffered saline (PBS) buffer (PH 7.4).
    Figure Legend Snippet: Square wave voltammograms obtained at the gold electrode immobilized with the DNA strands. ( A ) In the absence of ER, (a) before, and (b) after the DNA strands are digested by Exo III; ( B ) In the presence of 100 nM ER, (a) before, and (b) after the DNA strands are digested by Exo III. Curves c–e are for the control experiments by using 500 nM bovine serum albumin (BSA), thrombin and α-fetoprotein (AFP) instead of 100 nM ER. Buffer: 10 mM phosphate-buffered saline (PBS) buffer (PH 7.4).

    Techniques Used:

    6) Product Images from "In silico discovery of small molecules that inhibit RfaH recruitment to RNA polymerase"

    Article Title: In silico discovery of small molecules that inhibit RfaH recruitment to RNA polymerase

    Journal: Molecular microbiology

    doi: 10.1111/mmi.14093

    Probing RfaH-DNA interactions. A. A model of the RfaH-bound TEC. RNAP α (pale cyan), β (magenta) and β’ (orange) subunits, RfaH-NTD (gray), nucleic acids and Exo (green) are shown as cartoons. To provide an unobstructed view of the RfaH-NTD and the exposed nontemplate DNA, the EC is shown inan orientation that is opposite to the conventional left-to-right direction of transcription. The ops TEC scaffold used in these experiments is shown below, with nucleic acid chains colored and oriented as in the model; the ops element is in black. The upstream TA cross-linking motif is highlighted in yellow. B. Footprinting of the upstream RNAP boundary. The template strand DNA was 5’-end labeled with [γ 32 P]-ATP. After the addition of Exo III, aliquots were quenched at the indicated times (0 represents an untreated DNA control) and analyzed on a 12% denaturing gel; a representative of three independent experiments is shown. Numbers indicate the distance from the RNAP active site (yellow circle). .
    Figure Legend Snippet: Probing RfaH-DNA interactions. A. A model of the RfaH-bound TEC. RNAP α (pale cyan), β (magenta) and β’ (orange) subunits, RfaH-NTD (gray), nucleic acids and Exo (green) are shown as cartoons. To provide an unobstructed view of the RfaH-NTD and the exposed nontemplate DNA, the EC is shown inan orientation that is opposite to the conventional left-to-right direction of transcription. The ops TEC scaffold used in these experiments is shown below, with nucleic acid chains colored and oriented as in the model; the ops element is in black. The upstream TA cross-linking motif is highlighted in yellow. B. Footprinting of the upstream RNAP boundary. The template strand DNA was 5’-end labeled with [γ 32 P]-ATP. After the addition of Exo III, aliquots were quenched at the indicated times (0 represents an untreated DNA control) and analyzed on a 12% denaturing gel; a representative of three independent experiments is shown. Numbers indicate the distance from the RNAP active site (yellow circle). .

    Techniques Used: Footprinting, Labeling

    7) Product Images from "Locking the non-template DNA to control transcription"

    Article Title: Locking the non-template DNA to control transcription

    Journal: Molecular microbiology

    doi: 10.1111/mmi.13983

    Protection of the upstream DNA by RfaH and RNAP variants. A. U11 ECs were assembled with [γ 32 P]-ATP labeled T strand with the WT or altered RNAP. Exo III was added following incubation with RfaH (or storage buffer) and the reactions were analyzed as above. B. The WT U11 ECs assembled as in A were probed with Exo III in the presence of selected RfaH variants. AAAA is a quadruple mutant in which HTTT residues 65–68 were replaced with four alanines.
    Figure Legend Snippet: Protection of the upstream DNA by RfaH and RNAP variants. A. U11 ECs were assembled with [γ 32 P]-ATP labeled T strand with the WT or altered RNAP. Exo III was added following incubation with RfaH (or storage buffer) and the reactions were analyzed as above. B. The WT U11 ECs assembled as in A were probed with Exo III in the presence of selected RfaH variants. AAAA is a quadruple mutant in which HTTT residues 65–68 were replaced with four alanines.

    Techniques Used: Labeling, Incubation, Mutagenesis

    The upstream protection is independent of the ops DNA sequence. The scrambled ops ECs were assembled on a scaffold shown on top; the NT strand residues in magenta match those in ops . The wild-type full-length RfaH does not bind to this EC because contacts with the ops element are required to induce RfaH domain dissociation to expose the RNAP-binding site on the NTD. The isolated NTD binds to any EC and serves and a model of activated, post-recruitment conformation of RfaH. The assembled with the T DNA strand 5′-end labeled with [γ 32 P]-ATP EC was incubated with RfaH or NTD (at 100 nM) and treated with Exo III. The reactions were analyzed on a 12% urea-acrylamide gel. Numbers indicate the distance from the RNAP active site.
    Figure Legend Snippet: The upstream protection is independent of the ops DNA sequence. The scrambled ops ECs were assembled on a scaffold shown on top; the NT strand residues in magenta match those in ops . The wild-type full-length RfaH does not bind to this EC because contacts with the ops element are required to induce RfaH domain dissociation to expose the RNAP-binding site on the NTD. The isolated NTD binds to any EC and serves and a model of activated, post-recruitment conformation of RfaH. The assembled with the T DNA strand 5′-end labeled with [γ 32 P]-ATP EC was incubated with RfaH or NTD (at 100 nM) and treated with Exo III. The reactions were analyzed on a 12% urea-acrylamide gel. Numbers indicate the distance from the RNAP active site.

    Techniques Used: Sequencing, Binding Assay, Isolation, Labeling, Incubation, Acrylamide Gel Assay

    RfaH interactions with the EC. A. A model of the RfaH-bound EC. RNAP α (grey), β (blue), and β′ (yellow) subunits and RfaH NTD (magenta) are depicted by simplified molecular surfaces; β (with the GL labeled) and RfaH NTD are rendered semi-transparent. Nucleic acids are shown as cartoons, two Mg 2+ ions in the active site – as cyan spheres, and an incoming NTP – as red sticks. B and C. Footprinting of the upstream (B) and downstream (C) RNAP boundary in G8 and U11 ops ECs. Exo III was added to ECs assembled on synthetic scaffolds in which RNAP is halted at G8 and U11 positions in the ops element (NT sequence G 1 GCGGTAG 8 CGT 11 G); the probed DNA strand (T, panel B; NT, panel C) was 5′-end labeled with [γ 32 P]-ATP. RfaH was present at 100 nM where indicated. Aliquots were quenched at the indicated times (0 represents an untreated DNA control) and analyzed on a 12 % denaturing gel; a representative of three independent experiments is shown. Numbers indicate the distance from the RNAP active site.
    Figure Legend Snippet: RfaH interactions with the EC. A. A model of the RfaH-bound EC. RNAP α (grey), β (blue), and β′ (yellow) subunits and RfaH NTD (magenta) are depicted by simplified molecular surfaces; β (with the GL labeled) and RfaH NTD are rendered semi-transparent. Nucleic acids are shown as cartoons, two Mg 2+ ions in the active site – as cyan spheres, and an incoming NTP – as red sticks. B and C. Footprinting of the upstream (B) and downstream (C) RNAP boundary in G8 and U11 ops ECs. Exo III was added to ECs assembled on synthetic scaffolds in which RNAP is halted at G8 and U11 positions in the ops element (NT sequence G 1 GCGGTAG 8 CGT 11 G); the probed DNA strand (T, panel B; NT, panel C) was 5′-end labeled with [γ 32 P]-ATP. RfaH was present at 100 nM where indicated. Aliquots were quenched at the indicated times (0 represents an untreated DNA control) and analyzed on a 12 % denaturing gel; a representative of three independent experiments is shown. Numbers indicate the distance from the RNAP active site.

    Techniques Used: Labeling, Footprinting, Sequencing

    8) Product Images from "The universally-conserved transcription factor RfaH is recruited to a hairpin structure of the non-template DNA strand"

    Article Title: The universally-conserved transcription factor RfaH is recruited to a hairpin structure of the non-template DNA strand

    Journal: eLife

    doi: 10.7554/eLife.36349

    RfaH recruitment to RNAP transcribing through the ops element. ( A ) Schematic of Exo III footprinting of free and RfaH-bound TECs. Numbers indicate the upstream footprint boundaries relative to the RNA 3’ end. ( B ) The G8 TEC was assembled on the scaffold, with RNA and template (T) DNA strands labeled with [γ 32 P]-ATP and T4 polynucleotide kinase (PNK), and walked in one-nucleotide steps to C9, G10, and U11 positions in the presence of the matching NTP substrates. ( C ) RfaH was added to 50 nM, where indicated. Following the addition of Exo III, the reactions were quenched at indicated times (0 represents an untreated DNA control) and analyzed on a 12% urea-acrylamide (19:1) gel in 0.5X TBE. Numbers indicate the distance from the RNA 3’ end. Hypothetical TEC structures are shown below. G8 and C9 complexes are predominantly post-translocated, as indicated by 14 bp protection of the upstream DNA. In G10 TEC, the pre-translocated state (15 bp protection) is observed, and in U11 an additional backtracked state (16 bp protection). Exo III may counteract backtracking; the sensitivity of the nascent RNA in G10 and U11 TECs to GreB-assisted cleavage ( Nedialkov et al., 2018 ) was used to infer the translocation states shown in the schematics.
    Figure Legend Snippet: RfaH recruitment to RNAP transcribing through the ops element. ( A ) Schematic of Exo III footprinting of free and RfaH-bound TECs. Numbers indicate the upstream footprint boundaries relative to the RNA 3’ end. ( B ) The G8 TEC was assembled on the scaffold, with RNA and template (T) DNA strands labeled with [γ 32 P]-ATP and T4 polynucleotide kinase (PNK), and walked in one-nucleotide steps to C9, G10, and U11 positions in the presence of the matching NTP substrates. ( C ) RfaH was added to 50 nM, where indicated. Following the addition of Exo III, the reactions were quenched at indicated times (0 represents an untreated DNA control) and analyzed on a 12% urea-acrylamide (19:1) gel in 0.5X TBE. Numbers indicate the distance from the RNA 3’ end. Hypothetical TEC structures are shown below. G8 and C9 complexes are predominantly post-translocated, as indicated by 14 bp protection of the upstream DNA. In G10 TEC, the pre-translocated state (15 bp protection) is observed, and in U11 an additional backtracked state (16 bp protection). Exo III may counteract backtracking; the sensitivity of the nascent RNA in G10 and U11 TECs to GreB-assisted cleavage ( Nedialkov et al., 2018 ) was used to infer the translocation states shown in the schematics.

    Techniques Used: Footprinting, Labeling, Translocation Assay

    9) Product Images from "Transient Reversal of RNA Polymerase II Active Site Closing Controls Fidelity of Transcription Elongation"

    Article Title: Transient Reversal of RNA Polymerase II Active Site Closing Controls Fidelity of Transcription Elongation

    Journal: Molecular cell

    doi: 10.1016/j.molcel.2008.04.017

    Translocation Properties of the WT and E1103G Pol II (A) Experimental setup. (B) Dynamics of DNA degradation by Exo III reveals equilibrium between pre- and posttranslocated states of the TEC. TEC9 variants were obtained by 5 min incubation of TEC8 with 10 μM ATP or 3′dATP. The RNAs are shown in the lower panel. Note the aberrantly high mobility of the 9 nt RNA containing 3′dAMP. (C) The E1103G mutation confers an equilibrium shift toward the pretranslocated state in the stalled TEC. TEC10 variants were obtained by 5 min incubation of TEC9 with 10 μM of CTP or 3′dCTP. (D) Forward translocation of Pol II in TEC9 containing terminating 3′dAMP is induced by the incoming substrate. CTP was added at the final concentrations indicated on top of the gel. (E) NTP-stabilized forward translocation of Pol II in TEC9 containing terminating 3′dAMP requires complementary substrate. CTP, UTP, GTP, and ATP were added at 1 μM.
    Figure Legend Snippet: Translocation Properties of the WT and E1103G Pol II (A) Experimental setup. (B) Dynamics of DNA degradation by Exo III reveals equilibrium between pre- and posttranslocated states of the TEC. TEC9 variants were obtained by 5 min incubation of TEC8 with 10 μM ATP or 3′dATP. The RNAs are shown in the lower panel. Note the aberrantly high mobility of the 9 nt RNA containing 3′dAMP. (C) The E1103G mutation confers an equilibrium shift toward the pretranslocated state in the stalled TEC. TEC10 variants were obtained by 5 min incubation of TEC9 with 10 μM of CTP or 3′dCTP. (D) Forward translocation of Pol II in TEC9 containing terminating 3′dAMP is induced by the incoming substrate. CTP was added at the final concentrations indicated on top of the gel. (E) NTP-stabilized forward translocation of Pol II in TEC9 containing terminating 3′dAMP requires complementary substrate. CTP, UTP, GTP, and ATP were added at 1 μM.

    Techniques Used: Translocation Assay, Incubation, Mutagenesis

    10) Product Images from "The replication of plastid minicircles involves rolling circle intermediates"

    Article Title: The replication of plastid minicircles involves rolling circle intermediates

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkp063

    Exo III-treated minicircular DNA revealed after PFGE and 2D-gel electrophoresis. Exo III-treated whole-DNA extract from H. triquetra was subjected to PFGE ( A ) and 2D-gel ( B ), and detected with 1.1-kb psbA NCR fragment [and psbA gene fragment in (B) only]. (A) The 6–8 kb minicircular DNA bands (APBs) were removed in the Exo III lane, while the putative 3-kb psbA minicircle band was diminished in intensity. (B) Hybridization signals of larger than 2 kb were removed after Exo III treatment as revealed in both 2D-gels detected by psbA NCR and gene fragments. Extra spots indicated by the arrows were observed in these two images when comparing with the controls. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).
    Figure Legend Snippet: Exo III-treated minicircular DNA revealed after PFGE and 2D-gel electrophoresis. Exo III-treated whole-DNA extract from H. triquetra was subjected to PFGE ( A ) and 2D-gel ( B ), and detected with 1.1-kb psbA NCR fragment [and psbA gene fragment in (B) only]. (A) The 6–8 kb minicircular DNA bands (APBs) were removed in the Exo III lane, while the putative 3-kb psbA minicircle band was diminished in intensity. (B) Hybridization signals of larger than 2 kb were removed after Exo III treatment as revealed in both 2D-gels detected by psbA NCR and gene fragments. Extra spots indicated by the arrows were observed in these two images when comparing with the controls. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).

    Techniques Used: Two-Dimensional Gel Electrophoresis, Electrophoresis, Hybridization

    Effects of DNA ligase/Klenow fragment on minicircular DNA. Total DNA from H. triquetra was treated with T4 DNA ligase, as well as Klenow fragment followed with T4 DNA ligase, prior to Exo III digestion. Treated DNAs were resolved by PFGE and detected the minicircular DNA signals by Southern Blot. The two upper arrows indicated APBs in the untreated control lane. The two lower arrows indicated the putative monomer of psbA minicircle signals of ∼2–3 kb. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).
    Figure Legend Snippet: Effects of DNA ligase/Klenow fragment on minicircular DNA. Total DNA from H. triquetra was treated with T4 DNA ligase, as well as Klenow fragment followed with T4 DNA ligase, prior to Exo III digestion. Treated DNAs were resolved by PFGE and detected the minicircular DNA signals by Southern Blot. The two upper arrows indicated APBs in the untreated control lane. The two lower arrows indicated the putative monomer of psbA minicircle signals of ∼2–3 kb. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).

    Techniques Used: Southern Blot

    11) Product Images from "Sensitive Detection of Small-Molecule Targets Using Cooperative Binding Split Aptamers and Enzyme-Assisted Target Recycling"

    Article Title: Sensitive Detection of Small-Molecule Targets Using Cooperative Binding Split Aptamers and Enzyme-Assisted Target Recycling

    Journal: Analytical chemistry

    doi: 10.1021/acs.analchem.7b03625

    Colorimetric CBSA-based EATR-amplified assay for naked-eye detection of cocaine. (A) AuNPs are coupled to the short fragment (COC-SF), creating steric repulsion between AuNPs that produces a visible red color. Upon addition of long fragment (COC-LF) and cocaine (B), the CBSA assembles, and the resulting duplexed abasic site can then (C) be digested by Exo III. (D) This cleaves off COC-SF while recycling COC-LF and cocaine for another round of assembly and digestion, until (E) all short fragments have been sheared from the AuNP. (F) These sheared AuNPs can now aggregate, producing a visible red-to-blue color change.
    Figure Legend Snippet: Colorimetric CBSA-based EATR-amplified assay for naked-eye detection of cocaine. (A) AuNPs are coupled to the short fragment (COC-SF), creating steric repulsion between AuNPs that produces a visible red color. Upon addition of long fragment (COC-LF) and cocaine (B), the CBSA assembles, and the resulting duplexed abasic site can then (C) be digested by Exo III. (D) This cleaves off COC-SF while recycling COC-LF and cocaine for another round of assembly and digestion, until (E) all short fragments have been sheared from the AuNP. (F) These sheared AuNPs can now aggregate, producing a visible red-to-blue color change.

    Techniques Used: Amplification

    Working principle of our CBSA-based, EATR-amplified fluorescence assay. (A) FQ–DIS-SF contains a fluorophore–quencher pair and remains dark in the absence of target. (B) In the presence of DIS, the CBSA assembles (C), creating a duplex that can be recognized and cleaved by Exo III at the abasic site (denoted by X). (D) The complex then disassembles, releasing the intact long fragment and target for additional rounds of complex formation and digestion. The fluorophore is also released and produces a signal, which (E) becomes amplified over the course of many rounds of EATR.
    Figure Legend Snippet: Working principle of our CBSA-based, EATR-amplified fluorescence assay. (A) FQ–DIS-SF contains a fluorophore–quencher pair and remains dark in the absence of target. (B) In the presence of DIS, the CBSA assembles (C), creating a duplex that can be recognized and cleaved by Exo III at the abasic site (denoted by X). (D) The complex then disassembles, releasing the intact long fragment and target for additional rounds of complex formation and digestion. The fluorophore is also released and produces a signal, which (E) becomes amplified over the course of many rounds of EATR.

    Techniques Used: Amplification, Fluorescence

    Time-course of Exo III-mediated cleavage of DIS–CBSA-4536. (A) PAGE analysis of digestion products from DIS–CBSA-4536. Reactions consisted of 1 μ M DIS-SF, 1 μ M DIS-LF, and 0.004 U/ μ L Exo III with or without 500 μ M DIS after 5, 10, 15, 30, 60, and 120 min of digestion. Cleavage of short (B) and long (C) fragments was quantified by (Int 0 – Int)/Int 0 × 100%, where Int is the band intensity after the addition of Exo III and Int 0 is the band intensity before the addition of Exo III.
    Figure Legend Snippet: Time-course of Exo III-mediated cleavage of DIS–CBSA-4536. (A) PAGE analysis of digestion products from DIS–CBSA-4536. Reactions consisted of 1 μ M DIS-SF, 1 μ M DIS-LF, and 0.004 U/ μ L Exo III with or without 500 μ M DIS after 5, 10, 15, 30, 60, and 120 min of digestion. Cleavage of short (B) and long (C) fragments was quantified by (Int 0 – Int)/Int 0 × 100%, where Int is the band intensity after the addition of Exo III and Int 0 is the band intensity before the addition of Exo III.

    Techniques Used: Polyacrylamide Gel Electrophoresis

    CBSA-based EATR-amplified fluorescence assay for sensitive detection of DIS. (A) Fluorescence time-course measurements after adding 0.01 U/ μ L of Exo III to our CBSA in the presence of different concentrations of DIS (0, 0.5, 1, 2, 5, 10, 20, 50, 100, and 200 μ ).
    Figure Legend Snippet: CBSA-based EATR-amplified fluorescence assay for sensitive detection of DIS. (A) Fluorescence time-course measurements after adding 0.01 U/ μ L of Exo III to our CBSA in the presence of different concentrations of DIS (0, 0.5, 1, 2, 5, 10, 20, 50, 100, and 200 μ ).

    Techniques Used: Amplification, Fluorescence

    Cocaine-binding CBSA with a poly(A) 5 -protected long fragment and different derivatives of the short fragment. Arrows indicate suitable digestion sites for Exo III.
    Figure Legend Snippet: Cocaine-binding CBSA with a poly(A) 5 -protected long fragment and different derivatives of the short fragment. Arrows indicate suitable digestion sites for Exo III.

    Techniques Used: Binding Assay

    Design of DIS-binding CBSA. (A) The single-stranded DIS binding aptamer (DISS.1-AT) was truncated at stem 1 and stem 2 to generate the parent split aptamers. (B) Two sets of parent split aptamers were (C) merged and modified to incorporate (D) a C3 spacer abasic site (denoted by X) that resides opposite a thymine nucleotide when the CBSA duplex is assembled. A fluorophore (Cy5) and a quencher (Iowa Black RQ) were, respectively, modified at the 3′ and 5′ terminus of the short CBSA fragment. A poly(A) 5 overhang was added on the 3′ termini of each CBSA fragment to prevent nonspecific Exo III digestion.
    Figure Legend Snippet: Design of DIS-binding CBSA. (A) The single-stranded DIS binding aptamer (DISS.1-AT) was truncated at stem 1 and stem 2 to generate the parent split aptamers. (B) Two sets of parent split aptamers were (C) merged and modified to incorporate (D) a C3 spacer abasic site (denoted by X) that resides opposite a thymine nucleotide when the CBSA duplex is assembled. A fluorophore (Cy5) and a quencher (Iowa Black RQ) were, respectively, modified at the 3′ and 5′ terminus of the short CBSA fragment. A poly(A) 5 overhang was added on the 3′ termini of each CBSA fragment to prevent nonspecific Exo III digestion.

    Techniques Used: Binding Assay, Modification

    12) Product Images from "The replication of plastid minicircles involves rolling circle intermediates"

    Article Title: The replication of plastid minicircles involves rolling circle intermediates

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkp063

    Exo III-treated minicircular DNA revealed after PFGE and 2D-gel electrophoresis. Exo III-treated whole-DNA extract from H. triquetra was subjected to PFGE ( A ) and 2D-gel ( B ), and detected with 1.1-kb psbA NCR fragment [and psbA gene fragment in (B) only]. (A) The 6–8 kb minicircular DNA bands (APBs) were removed in the Exo III lane, while the putative 3-kb psbA minicircle band was diminished in intensity. (B) Hybridization signals of larger than 2 kb were removed after Exo III treatment as revealed in both 2D-gels detected by psbA NCR and gene fragments. Extra spots indicated by the arrows were observed in these two images when comparing with the controls. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).
    Figure Legend Snippet: Exo III-treated minicircular DNA revealed after PFGE and 2D-gel electrophoresis. Exo III-treated whole-DNA extract from H. triquetra was subjected to PFGE ( A ) and 2D-gel ( B ), and detected with 1.1-kb psbA NCR fragment [and psbA gene fragment in (B) only]. (A) The 6–8 kb minicircular DNA bands (APBs) were removed in the Exo III lane, while the putative 3-kb psbA minicircle band was diminished in intensity. (B) Hybridization signals of larger than 2 kb were removed after Exo III treatment as revealed in both 2D-gels detected by psbA NCR and gene fragments. Extra spots indicated by the arrows were observed in these two images when comparing with the controls. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).

    Techniques Used: Two-Dimensional Gel Electrophoresis, Electrophoresis, Hybridization

    Effects of DNA ligase/Klenow fragment on minicircular DNA. Total DNA from H. triquetra was treated with T4 DNA ligase, as well as Klenow fragment followed with T4 DNA ligase, prior to Exo III digestion. Treated DNAs were resolved by PFGE and detected the minicircular DNA signals by Southern Blot. The two upper arrows indicated APBs in the untreated control lane. The two lower arrows indicated the putative monomer of psbA minicircle signals of ∼2–3 kb. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).
    Figure Legend Snippet: Effects of DNA ligase/Klenow fragment on minicircular DNA. Total DNA from H. triquetra was treated with T4 DNA ligase, as well as Klenow fragment followed with T4 DNA ligase, prior to Exo III digestion. Treated DNAs were resolved by PFGE and detected the minicircular DNA signals by Southern Blot. The two upper arrows indicated APBs in the untreated control lane. The two lower arrows indicated the putative monomer of psbA minicircle signals of ∼2–3 kb. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).

    Techniques Used: Southern Blot

    13) Product Images from "Exonuclease III-Regulated Target Cyclic Amplification-Based Single Nucleotide Polymorphism Detection Using Ultrathin Ternary Chalcogenide Nanosheets"

    Article Title: Exonuclease III-Regulated Target Cyclic Amplification-Based Single Nucleotide Polymorphism Detection Using Ultrathin Ternary Chalcogenide Nanosheets

    Journal: Frontiers in Chemistry

    doi: 10.3389/fchem.2019.00844

    (A) Fluorescence spectra of P/MT + Exo III + Ta 2 NiS 5 (black), P/WT + Exo III + Ta 2 NiS 5 (red), P/MT + Ta 2 NiS 5 (pink), and P/WT + Ta 2 NiS 5 (green). (B) The fluorescence intensity ratio (FP/MT/FP/WT) at 610 nm for P/MT + Exo III and P/WT + Exo III in the absence (black) and presence (red) of Ta 2 NiS 5 nanosheets. (C) Fluorescence intensity of P/MT + Exo III (black) and P/WT + Exo III (red) in the presence of Ta 2 NiS 5 nanosheets with different final concentrations of 2.5, 5.0, 7.5, 10.0, and 12.5 μg ml −1 (P = 1 μM; MT = 100 nM; WT = 100 nM; Exo III = 0.25 U μl −1 ). (D) The fluorescence intensity ratio (FP/MT/FP/WT) at 610 nm in the presence of Ta2NiS5 nanosheets with different final concentrations of 2.5, 5.0, 7.5, 10.0, and 12.5 μg ml −1 (P = 1 μM; MT = 100 nM; WT = 100 nM; Exo III = 0.25 U μl −1) . The excitation wavelength is 590 nm.
    Figure Legend Snippet: (A) Fluorescence spectra of P/MT + Exo III + Ta 2 NiS 5 (black), P/WT + Exo III + Ta 2 NiS 5 (red), P/MT + Ta 2 NiS 5 (pink), and P/WT + Ta 2 NiS 5 (green). (B) The fluorescence intensity ratio (FP/MT/FP/WT) at 610 nm for P/MT + Exo III and P/WT + Exo III in the absence (black) and presence (red) of Ta 2 NiS 5 nanosheets. (C) Fluorescence intensity of P/MT + Exo III (black) and P/WT + Exo III (red) in the presence of Ta 2 NiS 5 nanosheets with different final concentrations of 2.5, 5.0, 7.5, 10.0, and 12.5 μg ml −1 (P = 1 μM; MT = 100 nM; WT = 100 nM; Exo III = 0.25 U μl −1 ). (D) The fluorescence intensity ratio (FP/MT/FP/WT) at 610 nm in the presence of Ta2NiS5 nanosheets with different final concentrations of 2.5, 5.0, 7.5, 10.0, and 12.5 μg ml −1 (P = 1 μM; MT = 100 nM; WT = 100 nM; Exo III = 0.25 U μl −1) . The excitation wavelength is 590 nm.

    Techniques Used: Fluorescence

    (A) The fluorescence spectra of P (1 μM) in the presence of different concentrations of mutant-type target (0, 0.001, 0.01, 0.1, 1, 10, and 100 nM) and Exo III (0.25 U μl −1 ) with addition of Ta 2 NiS 5 nanosheets (5.0 μg ml −1 ). (B) Relationship between fluorescence intensity at 610 nm and the concentrations of mutant-type target. Inset: Calibration curve for detection of mutant-type target. (C) Fluorescence spectra of different percentage of mutant-type target in mixed DNA samples (MT/(MT + WT) was 0, 5, 10, 20, 40, 60, 80, and 100%). (D) Fluorescence intensity at 610 nm as a function of allele frequency. The total concentration of the mutant and wild-type target is 100 pM. The excitation wavelength is 590 nm.
    Figure Legend Snippet: (A) The fluorescence spectra of P (1 μM) in the presence of different concentrations of mutant-type target (0, 0.001, 0.01, 0.1, 1, 10, and 100 nM) and Exo III (0.25 U μl −1 ) with addition of Ta 2 NiS 5 nanosheets (5.0 μg ml −1 ). (B) Relationship between fluorescence intensity at 610 nm and the concentrations of mutant-type target. Inset: Calibration curve for detection of mutant-type target. (C) Fluorescence spectra of different percentage of mutant-type target in mixed DNA samples (MT/(MT + WT) was 0, 5, 10, 20, 40, 60, 80, and 100%). (D) Fluorescence intensity at 610 nm as a function of allele frequency. The total concentration of the mutant and wild-type target is 100 pM. The excitation wavelength is 590 nm.

    Techniques Used: Fluorescence, Mutagenesis, Concentration Assay

    14) Product Images from "Mechanism of RNA polymerase II bypass of oxidative cyclopurine DNA lesions"

    Article Title: Mechanism of RNA polymerase II bypass of oxidative cyclopurine DNA lesions

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    doi: 10.1073/pnas.1415186112

    Translocation barrier beyond the CydA lesion. ( A ) Rear-end footprinting experimental setup. The figure illustrates the predicted translocation register of lesion-stalled TECs. The length of the template DNA strand protected from Exo III digestion differs
    Figure Legend Snippet: Translocation barrier beyond the CydA lesion. ( A ) Rear-end footprinting experimental setup. The figure illustrates the predicted translocation register of lesion-stalled TECs. The length of the template DNA strand protected from Exo III digestion differs

    Techniques Used: Translocation Assay, Footprinting

    15) Product Images from "Versatile transcription control based on reversible dCas9 binding"

    Article Title: Versatile transcription control based on reversible dCas9 binding

    Journal: RNA

    doi: 10.1261/rna.071613.119

    Blockage of various RNAPs by sgRNA5:dCas9 complexes. ( A ) Cy5-labeled DNA template encoding U5 was transcribed by SP6, T3, T7, or Eco RNAP, and reaction aliquots were removed and digested using Exonuclease III. The tDNA strand was visualized on a denaturing urea-polyacrylamide gel by detecting Cy5 (magenta star), and all nucleic acid species were visualized via staining with SYBR gold (green halos). Each species is given a letter that is used to identify its band. Band A: full-length DNA template; B: full-length U5; C: protected ntDNA fragment; D: blockage product; E: sgRNA5; and F: protected tDNA fragment. ( B ) Urea-polyacrylamide gels analyzing transcription reactions using SP6, T3, and T7 RNAPs, with Cy5 (magenta) and SYBR gold (green) scans overlaid. Reactions were performed with, from left to right , SP6 RNAP in the absence of a blockade, and SP6, T3, and T7 RNAPs in the presence of a blockade. Nucleic acid bands are labeled as indicated in panel A . The time points represented by the gray triangle are 5, 10, 15, 30, 60, and 90 min, left to right . ( C ) Quantification of the number of molecules of blockage product (band D) produced per molecule of RNAP for the reactions containing SP6, T7, and T3 RNAPs. ( Inset ) Quantification of molecules of full-length U5 (band B) produced per molecule of RNAP. ( D ) Urea-polyacrylamide gel analyzing a transcription reaction using Eco RNAP, with Cy5 (magenta) and SYBR gold (green) scans overlaid. Bands are labeled as in panel B . ( Right ) Band F displayed at higher contrast, showing new bands (F*) that are dependent on the presence of both Exo III and RNAP. ( E ) Quantification of the number of molecules of full-length U5 and blockage product produced per molecule of Eco RNAP.
    Figure Legend Snippet: Blockage of various RNAPs by sgRNA5:dCas9 complexes. ( A ) Cy5-labeled DNA template encoding U5 was transcribed by SP6, T3, T7, or Eco RNAP, and reaction aliquots were removed and digested using Exonuclease III. The tDNA strand was visualized on a denaturing urea-polyacrylamide gel by detecting Cy5 (magenta star), and all nucleic acid species were visualized via staining with SYBR gold (green halos). Each species is given a letter that is used to identify its band. Band A: full-length DNA template; B: full-length U5; C: protected ntDNA fragment; D: blockage product; E: sgRNA5; and F: protected tDNA fragment. ( B ) Urea-polyacrylamide gels analyzing transcription reactions using SP6, T3, and T7 RNAPs, with Cy5 (magenta) and SYBR gold (green) scans overlaid. Reactions were performed with, from left to right , SP6 RNAP in the absence of a blockade, and SP6, T3, and T7 RNAPs in the presence of a blockade. Nucleic acid bands are labeled as indicated in panel A . The time points represented by the gray triangle are 5, 10, 15, 30, 60, and 90 min, left to right . ( C ) Quantification of the number of molecules of blockage product (band D) produced per molecule of RNAP for the reactions containing SP6, T7, and T3 RNAPs. ( Inset ) Quantification of molecules of full-length U5 (band B) produced per molecule of RNAP. ( D ) Urea-polyacrylamide gel analyzing a transcription reaction using Eco RNAP, with Cy5 (magenta) and SYBR gold (green) scans overlaid. Bands are labeled as in panel B . ( Right ) Band F displayed at higher contrast, showing new bands (F*) that are dependent on the presence of both Exo III and RNAP. ( E ) Quantification of the number of molecules of full-length U5 and blockage product produced per molecule of Eco RNAP.

    Techniques Used: Labeling, Staining, Produced

    Binding and displacement of blockades containing mismatched sgRNAs. ( A ) Schematic of a dCas9 blockade with mismatches (blue, indicated by an arrow) between the DNA target sequence and the 5′ end of the sgRNA. A perfectly matched trap DNA is added as a competitor, displacing the blockade from the original target. ( B ) Cy5 scan of a native polyacrylamide gel showing WT or 4MM sgRNA1:dCas9:DNA complexes after the addition of different concentrations of trap DNA. Traps containing target sequences perfectly matched to the WT and 4MM sgRNAs are compared in their ability to displace blockades. The fraction intact is quantified on the right . ( C ) Close-up of the gel in B showing the difference in the migration position between WT and 4MM complexes ( left ). Close-up of the DNA region of an independent native polyacrylamide gel after SYBR gold staining reveals that the difference in migration occurs with the sgRNA:dCas9 complex alone ( right ). ( D ) Schematics of digestion of sgRNA:dCas9-bound DNA with Exo III and λ Exo. In Exo III digestions, the tDNA is detected via Cy5 labeling (magenta star), while in both digestions, all nucleic acid species are detected via SYBR gold staining (green halo). ( E ) Comparison of exonuclease digestions of DNA with WT sgRNA:dCas9 blockades and blockades containing one to four mismatches, with Cy5 (magenta) and SYBR gold (green) scans overlaid. In the Exo III gel, a digestion of DNA with an sgRNA5:dCas9 blockade is shown for comparison.
    Figure Legend Snippet: Binding and displacement of blockades containing mismatched sgRNAs. ( A ) Schematic of a dCas9 blockade with mismatches (blue, indicated by an arrow) between the DNA target sequence and the 5′ end of the sgRNA. A perfectly matched trap DNA is added as a competitor, displacing the blockade from the original target. ( B ) Cy5 scan of a native polyacrylamide gel showing WT or 4MM sgRNA1:dCas9:DNA complexes after the addition of different concentrations of trap DNA. Traps containing target sequences perfectly matched to the WT and 4MM sgRNAs are compared in their ability to displace blockades. The fraction intact is quantified on the right . ( C ) Close-up of the gel in B showing the difference in the migration position between WT and 4MM complexes ( left ). Close-up of the DNA region of an independent native polyacrylamide gel after SYBR gold staining reveals that the difference in migration occurs with the sgRNA:dCas9 complex alone ( right ). ( D ) Schematics of digestion of sgRNA:dCas9-bound DNA with Exo III and λ Exo. In Exo III digestions, the tDNA is detected via Cy5 labeling (magenta star), while in both digestions, all nucleic acid species are detected via SYBR gold staining (green halo). ( E ) Comparison of exonuclease digestions of DNA with WT sgRNA:dCas9 blockades and blockades containing one to four mismatches, with Cy5 (magenta) and SYBR gold (green) scans overlaid. In the Exo III gel, a digestion of DNA with an sgRNA5:dCas9 blockade is shown for comparison.

    Techniques Used: Binding Assay, Sequencing, Migration, Staining, Labeling

    16) Product Images from "The replication of plastid minicircles involves rolling circle intermediates"

    Article Title: The replication of plastid minicircles involves rolling circle intermediates

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkp063

    Exo III-treated minicircular DNA revealed after PFGE and 2D-gel electrophoresis. Exo III-treated whole-DNA extract from H. triquetra was subjected to PFGE ( A ) and 2D-gel ( B ), and detected with 1.1-kb psbA NCR fragment [and psbA gene fragment in (B) only]. (A) The 6–8 kb minicircular DNA bands (APBs) were removed in the Exo III lane, while the putative 3-kb psbA minicircle band was diminished in intensity. (B) Hybridization signals of larger than 2 kb were removed after Exo III treatment as revealed in both 2D-gels detected by psbA NCR and gene fragments. Extra spots indicated by the arrows were observed in these two images when comparing with the controls. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).
    Figure Legend Snippet: Exo III-treated minicircular DNA revealed after PFGE and 2D-gel electrophoresis. Exo III-treated whole-DNA extract from H. triquetra was subjected to PFGE ( A ) and 2D-gel ( B ), and detected with 1.1-kb psbA NCR fragment [and psbA gene fragment in (B) only]. (A) The 6–8 kb minicircular DNA bands (APBs) were removed in the Exo III lane, while the putative 3-kb psbA minicircle band was diminished in intensity. (B) Hybridization signals of larger than 2 kb were removed after Exo III treatment as revealed in both 2D-gels detected by psbA NCR and gene fragments. Extra spots indicated by the arrows were observed in these two images when comparing with the controls. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).

    Techniques Used: Two-Dimensional Gel Electrophoresis, Electrophoresis, Hybridization

    Effects of DNA ligase/Klenow fragment on minicircular DNA. Total DNA from H. triquetra was treated with T4 DNA ligase, as well as Klenow fragment followed with T4 DNA ligase, prior to Exo III digestion. Treated DNAs were resolved by PFGE and detected the minicircular DNA signals by Southern Blot. The two upper arrows indicated APBs in the untreated control lane. The two lower arrows indicated the putative monomer of psbA minicircle signals of ∼2–3 kb. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).
    Figure Legend Snippet: Effects of DNA ligase/Klenow fragment on minicircular DNA. Total DNA from H. triquetra was treated with T4 DNA ligase, as well as Klenow fragment followed with T4 DNA ligase, prior to Exo III digestion. Treated DNAs were resolved by PFGE and detected the minicircular DNA signals by Southern Blot. The two upper arrows indicated APBs in the untreated control lane. The two lower arrows indicated the putative monomer of psbA minicircle signals of ∼2–3 kb. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).

    Techniques Used: Southern Blot

    17) Product Images from "The replication of plastid minicircles involves rolling circle intermediates"

    Article Title: The replication of plastid minicircles involves rolling circle intermediates

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkp063

    Exo III-treated minicircular DNA revealed after PFGE and 2D-gel electrophoresis. Exo III-treated whole-DNA extract from H. triquetra was subjected to PFGE ( A ) and 2D-gel ( B ), and detected with 1.1-kb psbA NCR fragment [and psbA gene fragment in (B) only]. (A) The 6–8 kb minicircular DNA bands (APBs) were removed in the Exo III lane, while the putative 3-kb psbA minicircle band was diminished in intensity. (B) Hybridization signals of larger than 2 kb were removed after Exo III treatment as revealed in both 2D-gels detected by psbA NCR and gene fragments. Extra spots indicated by the arrows were observed in these two images when comparing with the controls. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).
    Figure Legend Snippet: Exo III-treated minicircular DNA revealed after PFGE and 2D-gel electrophoresis. Exo III-treated whole-DNA extract from H. triquetra was subjected to PFGE ( A ) and 2D-gel ( B ), and detected with 1.1-kb psbA NCR fragment [and psbA gene fragment in (B) only]. (A) The 6–8 kb minicircular DNA bands (APBs) were removed in the Exo III lane, while the putative 3-kb psbA minicircle band was diminished in intensity. (B) Hybridization signals of larger than 2 kb were removed after Exo III treatment as revealed in both 2D-gels detected by psbA NCR and gene fragments. Extra spots indicated by the arrows were observed in these two images when comparing with the controls. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).

    Techniques Used: Two-Dimensional Gel Electrophoresis, Electrophoresis, Hybridization

    Effects of DNA ligase/Klenow fragment on minicircular DNA. Total DNA from H. triquetra was treated with T4 DNA ligase, as well as Klenow fragment followed with T4 DNA ligase, prior to Exo III digestion. Treated DNAs were resolved by PFGE and detected the minicircular DNA signals by Southern Blot. The two upper arrows indicated APBs in the untreated control lane. The two lower arrows indicated the putative monomer of psbA minicircle signals of ∼2–3 kb. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).
    Figure Legend Snippet: Effects of DNA ligase/Klenow fragment on minicircular DNA. Total DNA from H. triquetra was treated with T4 DNA ligase, as well as Klenow fragment followed with T4 DNA ligase, prior to Exo III digestion. Treated DNAs were resolved by PFGE and detected the minicircular DNA signals by Southern Blot. The two upper arrows indicated APBs in the untreated control lane. The two lower arrows indicated the putative monomer of psbA minicircle signals of ∼2–3 kb. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).

    Techniques Used: Southern Blot

    18) Product Images from "The universally-conserved transcription factor RfaH is recruited to a hairpin structure of the non-template DNA strand"

    Article Title: The universally-conserved transcription factor RfaH is recruited to a hairpin structure of the non-template DNA strand

    Journal: eLife

    doi: 10.7554/eLife.36349

    RfaH recruitment to RNAP transcribing through the ops element. ( A ) Schematic of Exo III footprinting of free and RfaH-bound TECs. Numbers indicate the upstream footprint boundaries relative to the RNA 3’ end. ( B ) The G8 TEC was assembled on the scaffold, with RNA and template (T) DNA strands labeled with [γ 32 P]-ATP and T4 polynucleotide kinase (PNK), and walked in one-nucleotide steps to C9, G10, and U11 positions in the presence of the matching NTP substrates. ( C ) RfaH was added to 50 nM, where indicated. Following the addition of Exo III, the reactions were quenched at indicated times (0 represents an untreated DNA control) and analyzed on a 12% urea-acrylamide (19:1) gel in 0.5X TBE. Numbers indicate the distance from the RNA 3’ end. Hypothetical TEC structures are shown below. G8 and C9 complexes are predominantly post-translocated, as indicated by 14 bp protection of the upstream DNA. In G10 TEC, the pre-translocated state (15 bp protection) is observed, and in U11 an additional backtracked state (16 bp protection). Exo III may counteract backtracking; the sensitivity of the nascent RNA in G10 and U11 TECs to GreB-assisted cleavage ( Nedialkov et al., 2018 ) was used to infer the translocation states shown in the schematics.
    Figure Legend Snippet: RfaH recruitment to RNAP transcribing through the ops element. ( A ) Schematic of Exo III footprinting of free and RfaH-bound TECs. Numbers indicate the upstream footprint boundaries relative to the RNA 3’ end. ( B ) The G8 TEC was assembled on the scaffold, with RNA and template (T) DNA strands labeled with [γ 32 P]-ATP and T4 polynucleotide kinase (PNK), and walked in one-nucleotide steps to C9, G10, and U11 positions in the presence of the matching NTP substrates. ( C ) RfaH was added to 50 nM, where indicated. Following the addition of Exo III, the reactions were quenched at indicated times (0 represents an untreated DNA control) and analyzed on a 12% urea-acrylamide (19:1) gel in 0.5X TBE. Numbers indicate the distance from the RNA 3’ end. Hypothetical TEC structures are shown below. G8 and C9 complexes are predominantly post-translocated, as indicated by 14 bp protection of the upstream DNA. In G10 TEC, the pre-translocated state (15 bp protection) is observed, and in U11 an additional backtracked state (16 bp protection). Exo III may counteract backtracking; the sensitivity of the nascent RNA in G10 and U11 TECs to GreB-assisted cleavage ( Nedialkov et al., 2018 ) was used to infer the translocation states shown in the schematics.

    Techniques Used: Footprinting, Labeling, Translocation Assay

    19) Product Images from "Telomere-associated proteins add deoxynucleotides to terminal proteins during replication of the telomeres of linear chromosomes and plasmids in Streptomyces"

    Article Title: Telomere-associated proteins add deoxynucleotides to terminal proteins during replication of the telomeres of linear chromosomes and plasmids in Streptomyces

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkv302

    Characterization of the elongation products produced. ( A ) Different forms of products of oligonucleotide elongation . Form 1 . Tpg (large filled circle) linked to 1–3 radioactive dCMP (small filled circle). Form 2 . Tpg linked to radioactive dCMP and non-radioactive dNMP (open filled circle) with the total length not exceeding 7 nt. Form 3 . Tpg linked to13 nt-oligomer (Palindrome I) complexed with the template strand (shaded circles), the length of which is not to scale. Form 4 . Tpg attached to the 13-nt oligonucleotide (which fold back to form a hairpin) without the template, representing the denaturation product of Form 3 . Form 5 . Tpg linked to 13-nt oligomer duplexed with the template strand of 14–16 nt, representing T7 endonuclease I digestion product of Form 3 . Form 1 ′– 5 ′ proteinase K digestion products of Form 1 – 5 . Tpg was removed from the oligonucleotides. ( B ) The products of the first step of end patching based on TO65 . The template was 65- (left and middle panels) or 89-nt (right panel) telomere DNA. The reaction conditions were as in Figure 2B except that the product was eluted from the G-25 spin column in deionized water. The eluted product was subjected to proteinase K (‘PK’) digestion, Exo III (‘EX’) digestion, or heat denaturation (‘Δ’). Left panel . Only radioactive dCTP (‘dC’) was included. Middle panel . Radioactive dCTP and non-radioactive dATP, dTTP and dGTP (‘all dN’) were included. The different forms of products identified in the SDS gels were marked. The sizes of marker proteins (in kD) are indicated to the right. ( C ) The products produced on templates of different lengths. Upper panel. The reactions were carried under the same conditions as described in (B) with all dNTPs added. The lengths of the template strands used are as indicated. In three cases, the incubation time was increased from 10 m to 30 and 60 m as indicated. Some products were further treated with T7 endonuclease I (‘Endo’). Lower panel . The products were heat-denatured before being subjected to electrophoresis. The various forms of the products are marked in the gels. The product, which was eluted for further analysis in (D) is marked by an asterisk. ( D ) Sizing the length of the oligonucleotide . Form 3 produced from the 89 nt template (marked with an asterisk in (C) was eluted from the gel slice in 0.1% SDS by soaking. The eluent was concentrated by ethanol precipitation, re-dissolved in 1 N NaOH, and incubated at 37°C for 2 h. After neutralization with HCl, the sample was electrophoresed in a 15% urea–PAGE at 20 V/cm together with three size markers—single-stranded DNA of TO13, TO16 and TO35 as indicated. The positions of the size markers were determined by UV absorption and EtBr staining. The position of the products was determined by autoradiography.
    Figure Legend Snippet: Characterization of the elongation products produced. ( A ) Different forms of products of oligonucleotide elongation . Form 1 . Tpg (large filled circle) linked to 1–3 radioactive dCMP (small filled circle). Form 2 . Tpg linked to radioactive dCMP and non-radioactive dNMP (open filled circle) with the total length not exceeding 7 nt. Form 3 . Tpg linked to13 nt-oligomer (Palindrome I) complexed with the template strand (shaded circles), the length of which is not to scale. Form 4 . Tpg attached to the 13-nt oligonucleotide (which fold back to form a hairpin) without the template, representing the denaturation product of Form 3 . Form 5 . Tpg linked to 13-nt oligomer duplexed with the template strand of 14–16 nt, representing T7 endonuclease I digestion product of Form 3 . Form 1 ′– 5 ′ proteinase K digestion products of Form 1 – 5 . Tpg was removed from the oligonucleotides. ( B ) The products of the first step of end patching based on TO65 . The template was 65- (left and middle panels) or 89-nt (right panel) telomere DNA. The reaction conditions were as in Figure 2B except that the product was eluted from the G-25 spin column in deionized water. The eluted product was subjected to proteinase K (‘PK’) digestion, Exo III (‘EX’) digestion, or heat denaturation (‘Δ’). Left panel . Only radioactive dCTP (‘dC’) was included. Middle panel . Radioactive dCTP and non-radioactive dATP, dTTP and dGTP (‘all dN’) were included. The different forms of products identified in the SDS gels were marked. The sizes of marker proteins (in kD) are indicated to the right. ( C ) The products produced on templates of different lengths. Upper panel. The reactions were carried under the same conditions as described in (B) with all dNTPs added. The lengths of the template strands used are as indicated. In three cases, the incubation time was increased from 10 m to 30 and 60 m as indicated. Some products were further treated with T7 endonuclease I (‘Endo’). Lower panel . The products were heat-denatured before being subjected to electrophoresis. The various forms of the products are marked in the gels. The product, which was eluted for further analysis in (D) is marked by an asterisk. ( D ) Sizing the length of the oligonucleotide . Form 3 produced from the 89 nt template (marked with an asterisk in (C) was eluted from the gel slice in 0.1% SDS by soaking. The eluent was concentrated by ethanol precipitation, re-dissolved in 1 N NaOH, and incubated at 37°C for 2 h. After neutralization with HCl, the sample was electrophoresed in a 15% urea–PAGE at 20 V/cm together with three size markers—single-stranded DNA of TO13, TO16 and TO35 as indicated. The positions of the size markers were determined by UV absorption and EtBr staining. The position of the products was determined by autoradiography.

    Techniques Used: Produced, Marker, Incubation, Electrophoresis, Ethanol Precipitation, Neutralization, Polyacrylamide Gel Electrophoresis, Staining, Autoradiography

    20) Product Images from "The replication of plastid minicircles involves rolling circle intermediates"

    Article Title: The replication of plastid minicircles involves rolling circle intermediates

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkp063

    Exo III-treated minicircular DNA revealed after PFGE and 2D-gel electrophoresis. Exo III-treated whole-DNA extract from H. triquetra was subjected to PFGE ( A ) and 2D-gel ( B ), and detected with 1.1-kb psbA NCR fragment [and psbA gene fragment in (B) only]. (A) The 6–8 kb minicircular DNA bands (APBs) were removed in the Exo III lane, while the putative 3-kb psbA minicircle band was diminished in intensity. (B) Hybridization signals of larger than 2 kb were removed after Exo III treatment as revealed in both 2D-gels detected by psbA NCR and gene fragments. Extra spots indicated by the arrows were observed in these two images when comparing with the controls. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).
    Figure Legend Snippet: Exo III-treated minicircular DNA revealed after PFGE and 2D-gel electrophoresis. Exo III-treated whole-DNA extract from H. triquetra was subjected to PFGE ( A ) and 2D-gel ( B ), and detected with 1.1-kb psbA NCR fragment [and psbA gene fragment in (B) only]. (A) The 6–8 kb minicircular DNA bands (APBs) were removed in the Exo III lane, while the putative 3-kb psbA minicircle band was diminished in intensity. (B) Hybridization signals of larger than 2 kb were removed after Exo III treatment as revealed in both 2D-gels detected by psbA NCR and gene fragments. Extra spots indicated by the arrows were observed in these two images when comparing with the controls. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).

    Techniques Used: Two-Dimensional Gel Electrophoresis, Electrophoresis, Hybridization

    Effects of DNA ligase/Klenow fragment on minicircular DNA. Total DNA from H. triquetra was treated with T4 DNA ligase, as well as Klenow fragment followed with T4 DNA ligase, prior to Exo III digestion. Treated DNAs were resolved by PFGE and detected the minicircular DNA signals by Southern Blot. The two upper arrows indicated APBs in the untreated control lane. The two lower arrows indicated the putative monomer of psbA minicircle signals of ∼2–3 kb. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).
    Figure Legend Snippet: Effects of DNA ligase/Klenow fragment on minicircular DNA. Total DNA from H. triquetra was treated with T4 DNA ligase, as well as Klenow fragment followed with T4 DNA ligase, prior to Exo III digestion. Treated DNAs were resolved by PFGE and detected the minicircular DNA signals by Southern Blot. The two upper arrows indicated APBs in the untreated control lane. The two lower arrows indicated the putative monomer of psbA minicircle signals of ∼2–3 kb. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).

    Techniques Used: Southern Blot

    21) Product Images from "Identification of an Intermediate in Hepatitis B Virus Covalently Closed Circular (CCC) DNA Formation and Sensitive and Selective CCC DNA Detection"

    Article Title: Identification of an Intermediate in Hepatitis B Virus Covalently Closed Circular (CCC) DNA Formation and Sensitive and Selective CCC DNA Detection

    Journal: Journal of Virology

    doi: 10.1128/JVI.00539-17

    Exo I III versus Exo T5 digestion of plasmid DNA. (A) Diagrams showing expected digestion results of various plasmid DNA species. A break in the circle denotes the nick on the DNA strand. (B and C) Plasmid pCI-HBc (2.5 ng) was mixed with 20 μl of mock PF DNA extracted from uninduced HepAD38 cells. The DNA mix was first treated with Nb.BbvCI (5 units) to nick the plasmid DNA specifically on the minus strand (B and C, lanes 5 to 8) or was left untreated (B and C, lanes 1 to 4) before digestion with Exo I III (5 units and 25 units, respectively) in two different buffers or with Exo T5 (5 units). The DNA samples were then resolved on an agarose gel, and HBc DNA was detected by Southern blotting using a riboprobe specific for the viral plus-strand (B) or minus-strand (C) DNA. The diagrams on the right of panel C depict the various DNA species and their migration on the gel. B3, 1× NEB buffer 3; BCS, 1× NEB buffer Cutsmart; PE, phenol extraction.
    Figure Legend Snippet: Exo I III versus Exo T5 digestion of plasmid DNA. (A) Diagrams showing expected digestion results of various plasmid DNA species. A break in the circle denotes the nick on the DNA strand. (B and C) Plasmid pCI-HBc (2.5 ng) was mixed with 20 μl of mock PF DNA extracted from uninduced HepAD38 cells. The DNA mix was first treated with Nb.BbvCI (5 units) to nick the plasmid DNA specifically on the minus strand (B and C, lanes 5 to 8) or was left untreated (B and C, lanes 1 to 4) before digestion with Exo I III (5 units and 25 units, respectively) in two different buffers or with Exo T5 (5 units). The DNA samples were then resolved on an agarose gel, and HBc DNA was detected by Southern blotting using a riboprobe specific for the viral plus-strand (B) or minus-strand (C) DNA. The diagrams on the right of panel C depict the various DNA species and their migration on the gel. B3, 1× NEB buffer 3; BCS, 1× NEB buffer Cutsmart; PE, phenol extraction.

    Techniques Used: Plasmid Preparation, Agarose Gel Electrophoresis, Southern Blot, Migration

    Exo I III versus Exo T5 digestion of HBV core and PF DNA. (A) Diagrams showing expected results of digestion with various HBV PF DNA species. Left, structures of known and potential HBV PF DNA species; middle and right, expected digestion products of the various DNA species. The DNA species in the rectangular box, with a covalently closed minus strand and an open plus strand, represents a potential intermediate during RC DNA to CCC DNA conversion that was identified in the current study (see the text for details). The black dot at the 5′ end of the minus strand of the PF-RC and PF-DSL DNA denotes the unknown modification of this end upon removal of the RT protein (deproteination; see the text for details). (B and C) HBV core DNA (0.3 μl) combined with mock PF DNA (20 μl) extracted from uninduced HepAD38 cells (B) or PF DNA (20 μl) extracted from induced HepAD38 cells (C) was treated with Exo I III (5 units and 25 units, respectively) (lanes 3 and 10) or Exo T5 (5 units) (lanes 6 and 13) in 1× NEB CutSmart buffer. Subsequently, MfeI-HF (10 units) was used to linearize CCC DNA (lanes 5, 7, 12, and 14) and Exo T5 (5 units) was used to digest the SS circular DNA (lanes 4 and 11). Heat treatment (95°C, 10 min) was used to denature RC DNA to SS linear DNA (lanes 2 and 9). The DNA samples were then resolved on an agarose gel, and the various HBV DNA species were detected by Southern blotting using a riboprobe specific for the plus-strand (lanes 1 to 7) or minus-strand (lanes 8 to 14) DNA. The diagrams on the sides depict the various DNA species and their migration on the gel. The positions of the various RC DNA species, CCC DNA species, and SS linear and circular DNA species are indicated by the schematic diagrams. Note that the linearized CCC DNA comigrates with the DSL DNA, a minor form present in both core DNA and PF DNA (lanes 1 and 8).
    Figure Legend Snippet: Exo I III versus Exo T5 digestion of HBV core and PF DNA. (A) Diagrams showing expected results of digestion with various HBV PF DNA species. Left, structures of known and potential HBV PF DNA species; middle and right, expected digestion products of the various DNA species. The DNA species in the rectangular box, with a covalently closed minus strand and an open plus strand, represents a potential intermediate during RC DNA to CCC DNA conversion that was identified in the current study (see the text for details). The black dot at the 5′ end of the minus strand of the PF-RC and PF-DSL DNA denotes the unknown modification of this end upon removal of the RT protein (deproteination; see the text for details). (B and C) HBV core DNA (0.3 μl) combined with mock PF DNA (20 μl) extracted from uninduced HepAD38 cells (B) or PF DNA (20 μl) extracted from induced HepAD38 cells (C) was treated with Exo I III (5 units and 25 units, respectively) (lanes 3 and 10) or Exo T5 (5 units) (lanes 6 and 13) in 1× NEB CutSmart buffer. Subsequently, MfeI-HF (10 units) was used to linearize CCC DNA (lanes 5, 7, 12, and 14) and Exo T5 (5 units) was used to digest the SS circular DNA (lanes 4 and 11). Heat treatment (95°C, 10 min) was used to denature RC DNA to SS linear DNA (lanes 2 and 9). The DNA samples were then resolved on an agarose gel, and the various HBV DNA species were detected by Southern blotting using a riboprobe specific for the plus-strand (lanes 1 to 7) or minus-strand (lanes 8 to 14) DNA. The diagrams on the sides depict the various DNA species and their migration on the gel. The positions of the various RC DNA species, CCC DNA species, and SS linear and circular DNA species are indicated by the schematic diagrams. Note that the linearized CCC DNA comigrates with the DSL DNA, a minor form present in both core DNA and PF DNA (lanes 1 and 8).

    Techniques Used: Countercurrent Chromatography, Modification, Agarose Gel Electrophoresis, Southern Blot, Migration

    Confirmation of the closed circular minus strand in the processed RC DNA by BmgBI or Nt.BbvCI and Exo I III digestion. (A and D) Diagrams showing expected results of digestion performed with various HBV PF DNA species. The short line intersecting the circle denotes the site of BmgBI digestion (A) or Nt.BbvCI nicking (D). The presence of the RNA (short gray line) at the 5′ end of the plus strand in RC DNA prevents BmgBI digestion (panel A; arrow blocked by a short line). The black dot at the 5′ end of the minus strand of the PF-RC DNA denotes the unknown modification of this end upon removal of the RT protein. The DNA species indicated in the rectangular box, with a covalently closed minus strand and an open plus strand, represents a potential intermediate during RC DNA to CCC DNA conversion that was identified in this study (see the text for details). (B and C) HBV core DNA (0.3 μl) combined with mock PF DNA (20 μl) extracted from uninduced HepAD38 cells (lanes 1 to 3) or PF DNA (lanes 4 to 6) extracted from induced HepAD38 cells was treated with BmgBI (5 units) in 1× NEB buffer 3 to linearize all supercoiled and nicked CCC DNA (lanes 2, 3, 5, and 6) or was mock treated (lanes 1 and 4). For lanes 3 and 6, the DNA samples were further digested with Exo I III after BmgBI treatment. The samples were then resolved on an agarose gel, and various HBV DNA species were detected by Southern blotting using a riboprobe specific for the viral plus-strand (B) or minus-strand (C) DNA. The diagrams on the right of panel C depict the various DNA species and their migration on the gel. (E) PF DNA extracted from induced HepAD38 cells was treated with Nt.BbvCI (5 units) in 1× NEB Cutsmart buffer to nick all CCC DNA (lanes 3, 4, 7, and 8) or mock treated (lanes 1 and 5). For lanes 4 and 8, the DNA samples were further digested with Exo I III after Nt.BbvCI treatment. The samples were then resolved on an agarose gel, and various HBV DNA species were detected by Southern blotting using a riboprobe specific for the viral plus-strand (lanes 1 to 4) or minus-strand (lanes 5 to 8) DNA. The diagrams on the right depict the various DNA species and their migration on the gel. Marker, the DNA marker lane. The size of the DNA markers is indicated (in kilobase pairs). The blank spaces between the lanes in panels B, C, and E indicate where other lanes from the same gel that were deemed nonessential for this work were cropped out during the preparation of the figure.
    Figure Legend Snippet: Confirmation of the closed circular minus strand in the processed RC DNA by BmgBI or Nt.BbvCI and Exo I III digestion. (A and D) Diagrams showing expected results of digestion performed with various HBV PF DNA species. The short line intersecting the circle denotes the site of BmgBI digestion (A) or Nt.BbvCI nicking (D). The presence of the RNA (short gray line) at the 5′ end of the plus strand in RC DNA prevents BmgBI digestion (panel A; arrow blocked by a short line). The black dot at the 5′ end of the minus strand of the PF-RC DNA denotes the unknown modification of this end upon removal of the RT protein. The DNA species indicated in the rectangular box, with a covalently closed minus strand and an open plus strand, represents a potential intermediate during RC DNA to CCC DNA conversion that was identified in this study (see the text for details). (B and C) HBV core DNA (0.3 μl) combined with mock PF DNA (20 μl) extracted from uninduced HepAD38 cells (lanes 1 to 3) or PF DNA (lanes 4 to 6) extracted from induced HepAD38 cells was treated with BmgBI (5 units) in 1× NEB buffer 3 to linearize all supercoiled and nicked CCC DNA (lanes 2, 3, 5, and 6) or was mock treated (lanes 1 and 4). For lanes 3 and 6, the DNA samples were further digested with Exo I III after BmgBI treatment. The samples were then resolved on an agarose gel, and various HBV DNA species were detected by Southern blotting using a riboprobe specific for the viral plus-strand (B) or minus-strand (C) DNA. The diagrams on the right of panel C depict the various DNA species and their migration on the gel. (E) PF DNA extracted from induced HepAD38 cells was treated with Nt.BbvCI (5 units) in 1× NEB Cutsmart buffer to nick all CCC DNA (lanes 3, 4, 7, and 8) or mock treated (lanes 1 and 5). For lanes 4 and 8, the DNA samples were further digested with Exo I III after Nt.BbvCI treatment. The samples were then resolved on an agarose gel, and various HBV DNA species were detected by Southern blotting using a riboprobe specific for the viral plus-strand (lanes 1 to 4) or minus-strand (lanes 5 to 8) DNA. The diagrams on the right depict the various DNA species and their migration on the gel. Marker, the DNA marker lane. The size of the DNA markers is indicated (in kilobase pairs). The blank spaces between the lanes in panels B, C, and E indicate where other lanes from the same gel that were deemed nonessential for this work were cropped out during the preparation of the figure.

    Techniques Used: Modification, Countercurrent Chromatography, Agarose Gel Electrophoresis, Southern Blot, Migration, Marker

    Detection of CCC DNA, PF-RC DNA, and PF-RC DNA with closed minus strand in HBV-infected HepG2-NTCP cells. (A) HBV PF DNA was extracted from infected HepG2-NTCP cells at the indicated days (day 0.5 [D0.5], D1, D1.5, and D2) postinfection. The PF DNA was either mock treated (lanes 1 to 4) or treated with Exo I III (lanes 5 to 8). (B) The HBV PF DNA extracted from infected HepG2-NTCP cells (lanes 1 to 4 [processed as described for panel A]), along with core and PF DNA extracted from induced HepD38 cells (lanes 5 and 6), was digested with BmgBI followed by Exo I III digestion. The samples were then resolved on an agarose gel, and various HBV DNA species were detected by Southern blotting using a riboprobe specific for the viral minus-strand DNA.
    Figure Legend Snippet: Detection of CCC DNA, PF-RC DNA, and PF-RC DNA with closed minus strand in HBV-infected HepG2-NTCP cells. (A) HBV PF DNA was extracted from infected HepG2-NTCP cells at the indicated days (day 0.5 [D0.5], D1, D1.5, and D2) postinfection. The PF DNA was either mock treated (lanes 1 to 4) or treated with Exo I III (lanes 5 to 8). (B) The HBV PF DNA extracted from infected HepG2-NTCP cells (lanes 1 to 4 [processed as described for panel A]), along with core and PF DNA extracted from induced HepD38 cells (lanes 5 and 6), was digested with BmgBI followed by Exo I III digestion. The samples were then resolved on an agarose gel, and various HBV DNA species were detected by Southern blotting using a riboprobe specific for the viral minus-strand DNA.

    Techniques Used: Countercurrent Chromatography, Infection, Agarose Gel Electrophoresis, Southern Blot

    22) Product Images from "The replication of plastid minicircles involves rolling circle intermediates"

    Article Title: The replication of plastid minicircles involves rolling circle intermediates

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkp063

    Exo III-treated minicircular DNA revealed after PFGE and 2D-gel electrophoresis. Exo III-treated whole-DNA extract from H. triquetra was subjected to PFGE ( A ) and 2D-gel ( B ), and detected with 1.1-kb psbA NCR fragment [and psbA gene fragment in (B) only]. (A) The 6–8 kb minicircular DNA bands (APBs) were removed in the Exo III lane, while the putative 3-kb psbA minicircle band was diminished in intensity. (B) Hybridization signals of larger than 2 kb were removed after Exo III treatment as revealed in both 2D-gels detected by psbA NCR and gene fragments. Extra spots indicated by the arrows were observed in these two images when comparing with the controls. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).
    Figure Legend Snippet: Exo III-treated minicircular DNA revealed after PFGE and 2D-gel electrophoresis. Exo III-treated whole-DNA extract from H. triquetra was subjected to PFGE ( A ) and 2D-gel ( B ), and detected with 1.1-kb psbA NCR fragment [and psbA gene fragment in (B) only]. (A) The 6–8 kb minicircular DNA bands (APBs) were removed in the Exo III lane, while the putative 3-kb psbA minicircle band was diminished in intensity. (B) Hybridization signals of larger than 2 kb were removed after Exo III treatment as revealed in both 2D-gels detected by psbA NCR and gene fragments. Extra spots indicated by the arrows were observed in these two images when comparing with the controls. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).

    Techniques Used: Two-Dimensional Gel Electrophoresis, Electrophoresis, Hybridization

    Effects of DNA ligase/Klenow fragment on minicircular DNA. Total DNA from H. triquetra was treated with T4 DNA ligase, as well as Klenow fragment followed with T4 DNA ligase, prior to Exo III digestion. Treated DNAs were resolved by PFGE and detected the minicircular DNA signals by Southern Blot. The two upper arrows indicated APBs in the untreated control lane. The two lower arrows indicated the putative monomer of psbA minicircle signals of ∼2–3 kb. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).
    Figure Legend Snippet: Effects of DNA ligase/Klenow fragment on minicircular DNA. Total DNA from H. triquetra was treated with T4 DNA ligase, as well as Klenow fragment followed with T4 DNA ligase, prior to Exo III digestion. Treated DNAs were resolved by PFGE and detected the minicircular DNA signals by Southern Blot. The two upper arrows indicated APBs in the untreated control lane. The two lower arrows indicated the putative monomer of psbA minicircle signals of ∼2–3 kb. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).

    Techniques Used: Southern Blot

    23) Product Images from "The replication of plastid minicircles involves rolling circle intermediates"

    Article Title: The replication of plastid minicircles involves rolling circle intermediates

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkp063

    Exo III-treated minicircular DNA revealed after PFGE and 2D-gel electrophoresis. Exo III-treated whole-DNA extract from H. triquetra was subjected to PFGE ( A ) and 2D-gel ( B ), and detected with 1.1-kb psbA NCR fragment [and psbA gene fragment in (B) only]. (A) The 6–8 kb minicircular DNA bands (APBs) were removed in the Exo III lane, while the putative 3-kb psbA minicircle band was diminished in intensity. (B) Hybridization signals of larger than 2 kb were removed after Exo III treatment as revealed in both 2D-gels detected by psbA NCR and gene fragments. Extra spots indicated by the arrows were observed in these two images when comparing with the controls. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).
    Figure Legend Snippet: Exo III-treated minicircular DNA revealed after PFGE and 2D-gel electrophoresis. Exo III-treated whole-DNA extract from H. triquetra was subjected to PFGE ( A ) and 2D-gel ( B ), and detected with 1.1-kb psbA NCR fragment [and psbA gene fragment in (B) only]. (A) The 6–8 kb minicircular DNA bands (APBs) were removed in the Exo III lane, while the putative 3-kb psbA minicircle band was diminished in intensity. (B) Hybridization signals of larger than 2 kb were removed after Exo III treatment as revealed in both 2D-gels detected by psbA NCR and gene fragments. Extra spots indicated by the arrows were observed in these two images when comparing with the controls. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).

    Techniques Used: Two-Dimensional Gel Electrophoresis, Electrophoresis, Hybridization

    Effects of DNA ligase/Klenow fragment on minicircular DNA. Total DNA from H. triquetra was treated with T4 DNA ligase, as well as Klenow fragment followed with T4 DNA ligase, prior to Exo III digestion. Treated DNAs were resolved by PFGE and detected the minicircular DNA signals by Southern Blot. The two upper arrows indicated APBs in the untreated control lane. The two lower arrows indicated the putative monomer of psbA minicircle signals of ∼2–3 kb. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).
    Figure Legend Snippet: Effects of DNA ligase/Klenow fragment on minicircular DNA. Total DNA from H. triquetra was treated with T4 DNA ligase, as well as Klenow fragment followed with T4 DNA ligase, prior to Exo III digestion. Treated DNAs were resolved by PFGE and detected the minicircular DNA signals by Southern Blot. The two upper arrows indicated APBs in the untreated control lane. The two lower arrows indicated the putative monomer of psbA minicircle signals of ∼2–3 kb. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).

    Techniques Used: Southern Blot

    24) Product Images from "CAG/CTG Repeats Alter Affinity for the Histone Core and Positioning of DNA in the Nucleosome †"

    Article Title: CAG/CTG Repeats Alter Affinity for the Histone Core and Positioning of DNA in the Nucleosome †

    Journal: Biochemistry

    doi: 10.1021/bi301416v

    Exonuclease III digestion reveals weaker interactions between the ends of the DNA and the histone core. Exo III digestion of the CAG-containing strands of the S1 substrates are shown in panel A, while digestion of the CAG-containing strands of the HTT
    Figure Legend Snippet: Exonuclease III digestion reveals weaker interactions between the ends of the DNA and the histone core. Exo III digestion of the CAG-containing strands of the S1 substrates are shown in panel A, while digestion of the CAG-containing strands of the HTT

    Techniques Used:

    25) Product Images from "DNA microarrays with stem-loop DNA probes: preparation and applications"

    Article Title: DNA microarrays with stem-loop DNA probes: preparation and applications

    Journal: Nucleic Acids Research

    doi:

    Exonuclease III digests of Sanger sequencing ladders. ( A ) Outline of the experiments. ( B ) Image of the sequencing gel generated by software provided with the ALF sequencing instrument (Pharmacia Biotech, Uppsala, Sweden). Sanger sequencing ladders without and with exo III treatment. Arrows mark matching fragments. ( C ) Sequencing reads obtained before and after exo III digestion.
    Figure Legend Snippet: Exonuclease III digests of Sanger sequencing ladders. ( A ) Outline of the experiments. ( B ) Image of the sequencing gel generated by software provided with the ALF sequencing instrument (Pharmacia Biotech, Uppsala, Sweden). Sanger sequencing ladders without and with exo III treatment. Arrows mark matching fragments. ( C ) Sequencing reads obtained before and after exo III digestion.

    Techniques Used: Sequencing, Generated, Software

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    Article Snippet: .. Slides were exposed to 365-nm ultraviolet light using a Stratagene Stratalinker 1800 UV irradiator set to 5400 J, and incubated with 3 U/μl Exonuclease III (New England Biolabs) at 37 °C for 30 min. ..

    Incubation:

    Article Title: TERRA transcription destabilizes telomere integrity to initiate break-induced replication in human ALT cells
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    New England Biolabs exo iii
    <t>Exo</t> <t>III-treated</t> minicircular DNA revealed after PFGE and 2D-gel electrophoresis. Exo III-treated whole-DNA extract from H. triquetra was subjected to PFGE ( A ) and 2D-gel ( B ), and detected with 1.1-kb psbA NCR fragment [and psbA gene fragment in (B) only]. (A) The 6–8 kb minicircular DNA bands (APBs) were removed in the Exo III lane, while the putative 3-kb psbA minicircle band was diminished in intensity. (B) Hybridization signals of larger than 2 kb were removed after Exo III treatment as revealed in both 2D-gels detected by psbA NCR and gene fragments. Extra spots indicated by the arrows were observed in these two images when comparing with the controls. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).
    Exo Iii, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 97/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Exo III-treated minicircular DNA revealed after PFGE and 2D-gel electrophoresis. Exo III-treated whole-DNA extract from H. triquetra was subjected to PFGE ( A ) and 2D-gel ( B ), and detected with 1.1-kb psbA NCR fragment [and psbA gene fragment in (B) only]. (A) The 6–8 kb minicircular DNA bands (APBs) were removed in the Exo III lane, while the putative 3-kb psbA minicircle band was diminished in intensity. (B) Hybridization signals of larger than 2 kb were removed after Exo III treatment as revealed in both 2D-gels detected by psbA NCR and gene fragments. Extra spots indicated by the arrows were observed in these two images when comparing with the controls. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).

    Journal: Nucleic Acids Research

    Article Title: The replication of plastid minicircles involves rolling circle intermediates

    doi: 10.1093/nar/gkp063

    Figure Lengend Snippet: Exo III-treated minicircular DNA revealed after PFGE and 2D-gel electrophoresis. Exo III-treated whole-DNA extract from H. triquetra was subjected to PFGE ( A ) and 2D-gel ( B ), and detected with 1.1-kb psbA NCR fragment [and psbA gene fragment in (B) only]. (A) The 6–8 kb minicircular DNA bands (APBs) were removed in the Exo III lane, while the putative 3-kb psbA minicircle band was diminished in intensity. (B) Hybridization signals of larger than 2 kb were removed after Exo III treatment as revealed in both 2D-gels detected by psbA NCR and gene fragments. Extra spots indicated by the arrows were observed in these two images when comparing with the controls. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).

    Article Snippet: Total H. triquetra DNA (which contained minicircle DNA as detected in the Southern hybridization) were sequentially treated with Klenow, T4 DNA ligase and Exo III.

    Techniques: Two-Dimensional Gel Electrophoresis, Electrophoresis, Hybridization

    Effects of DNA ligase/Klenow fragment on minicircular DNA. Total DNA from H. triquetra was treated with T4 DNA ligase, as well as Klenow fragment followed with T4 DNA ligase, prior to Exo III digestion. Treated DNAs were resolved by PFGE and detected the minicircular DNA signals by Southern Blot. The two upper arrows indicated APBs in the untreated control lane. The two lower arrows indicated the putative monomer of psbA minicircle signals of ∼2–3 kb. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).

    Journal: Nucleic Acids Research

    Article Title: The replication of plastid minicircles involves rolling circle intermediates

    doi: 10.1093/nar/gkp063

    Figure Lengend Snippet: Effects of DNA ligase/Klenow fragment on minicircular DNA. Total DNA from H. triquetra was treated with T4 DNA ligase, as well as Klenow fragment followed with T4 DNA ligase, prior to Exo III digestion. Treated DNAs were resolved by PFGE and detected the minicircular DNA signals by Southern Blot. The two upper arrows indicated APBs in the untreated control lane. The two lower arrows indicated the putative monomer of psbA minicircle signals of ∼2–3 kb. DNA markers were linear DNAs from a commercial source (Invitrogen Corporation).

    Article Snippet: Total H. triquetra DNA (which contained minicircle DNA as detected in the Southern hybridization) were sequentially treated with Klenow, T4 DNA ligase and Exo III.

    Techniques: Southern Blot