hindiii Thermo Fisher Search Results


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  • 99
    Thermo Fisher hindii sites
    Representative <t>RFLP</t> patterns of the amplified 1100 bp fragment of actin gene obtained from T. vaginalis isolates. Lane 1, 5 and 8 digested by <t>HindII</t> ; Lane 2, 6 and 9 digested by Tru1 I; Lane 3, 7 and 10 digested by RsaI ; M: DNA ladder (50 bp)
    Hindii Sites, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 428 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher fastdigest hindiii
    Representative <t>RFLP</t> patterns of the amplified 1100 bp fragment of actin gene obtained from T. vaginalis isolates. Lane 1, 5 and 8 digested by <t>HindII</t> ; Lane 2, 6 and 9 digested by Tru1 I; Lane 3, 7 and 10 digested by RsaI ; M: DNA ladder (50 bp)
    Fastdigest Hindiii, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 115 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher hindiii
    Electrophoretic product of PvAMA-1 gene treated with EcoR I, PvuII, and <t>HindIII,</t> enzymes using RFLP-PCR technique including Columns 1,2 3 treated with EcoRI. Column 4 main bound of Pv AMA-1. Column 5 size marker with 1000bp. Columns 6, 7 and 8 treated with PvuII. Columns 9 10 treated with HindIII
    Hindiii, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 6954 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher ecori hindiii
    Electrophoretic product of PvAMA-1 gene treated with EcoR I, PvuII, and <t>HindIII,</t> enzymes using RFLP-PCR technique including Columns 1,2 3 treated with EcoRI. Column 4 main bound of Pv AMA-1. Column 5 size marker with 1000bp. Columns 6, 7 and 8 treated with PvuII. Columns 9 10 treated with HindIII
    Ecori Hindiii, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 85/100, based on 58 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher hindiii xbai
    Electrophoretic product of PvAMA-1 gene treated with EcoR I, PvuII, and <t>HindIII,</t> enzymes using RFLP-PCR technique including Columns 1,2 3 treated with EcoRI. Column 4 main bound of Pv AMA-1. Column 5 size marker with 1000bp. Columns 6, 7 and 8 treated with PvuII. Columns 9 10 treated with HindIII
    Hindiii Xbai, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 93/100, based on 66 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher hindiii restrictases
    Electrophoretic product of PvAMA-1 gene treated with EcoR I, PvuII, and <t>HindIII,</t> enzymes using RFLP-PCR technique including Columns 1,2 3 treated with EcoRI. Column 4 main bound of Pv AMA-1. Column 5 size marker with 1000bp. Columns 6, 7 and 8 treated with PvuII. Columns 9 10 treated with HindIII
    Hindiii Restrictases, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 80/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    90
    Thermo Fisher xhoi hindiii
    Electrophoretic product of PvAMA-1 gene treated with EcoR I, PvuII, and <t>HindIII,</t> enzymes using RFLP-PCR technique including Columns 1,2 3 treated with EcoRI. Column 4 main bound of Pv AMA-1. Column 5 size marker with 1000bp. Columns 6, 7 and 8 treated with PvuII. Columns 9 10 treated with HindIII
    Xhoi Hindiii, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 90/100, based on 15 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Thermo Fisher lambda dna hindiii marker
    (A) Agarose gel electrophoresis of the recovered <t>DNA</t> from the Guaymas Basin sediment sample. Equal amount of sample (0.3 g) were extracted with three methods. The loading volume of the recovered DNA was 1 μL. M indicates <t>Lambda</t> <t>DNA/HindIII</t> <t>Marker</t> 2 (Thermo Fisher Scientific, USA). (B) Representation of DNA band intensity (A) as plotted in three dimensional image. (C) Standard curve for measuring the DNA concentration using CLIQS 1D Pro software as determined by the DNA band intensity on the agarose gel.
    Lambda Dna Hindiii Marker, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 36 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Thermo Fisher bamhi hindiii
    (A) Agarose gel electrophoresis of the recovered <t>DNA</t> from the Guaymas Basin sediment sample. Equal amount of sample (0.3 g) were extracted with three methods. The loading volume of the recovered DNA was 1 μL. M indicates <t>Lambda</t> <t>DNA/HindIII</t> <t>Marker</t> 2 (Thermo Fisher Scientific, USA). (B) Representation of DNA band intensity (A) as plotted in three dimensional image. (C) Standard curve for measuring the DNA concentration using CLIQS 1D Pro software as determined by the DNA band intensity on the agarose gel.
    Bamhi Hindiii, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 93/100, based on 66 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher hindiii bamhi digested pcdna3
    (A) Agarose gel electrophoresis of the recovered <t>DNA</t> from the Guaymas Basin sediment sample. Equal amount of sample (0.3 g) were extracted with three methods. The loading volume of the recovered DNA was 1 μL. M indicates <t>Lambda</t> <t>DNA/HindIII</t> <t>Marker</t> 2 (Thermo Fisher Scientific, USA). (B) Representation of DNA band intensity (A) as plotted in three dimensional image. (C) Standard curve for measuring the DNA concentration using CLIQS 1D Pro software as determined by the DNA band intensity on the agarose gel.
    Hindiii Bamhi Digested Pcdna3, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 85/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    86
    Thermo Fisher lamda hindiii
    Assessment of YGL250W , SOH1 , and BRE5 (A) Schematic representation of Chromosome II from the diploid W303 background which consists of two LYS2 heteroalleles ( lys2–5′nde I − and lys2–3′nde I − ). These were used to measure meiotic gene conversion (see Materials and Methods ). (B) Spot test of wild type, ygl250wΔ , soh1Δ , and bre5Δ on haploid selection plates and haploid selection plates without lysine to measure meiotic gene conversion. The reduction in meiotic gene conversion of ygl250wΔ , soh1Δ , and bre5Δ was further assessed by random spore analysis ( Table 1 ). (C) Southern blot of DNA isolated from wild type, soh1Δ , and ygl250wΔ SK1 strains containing the ectopic URA3 - ARG4 interval on Chromosome III. The DNA from the indicated times after initiation of sporulation were digested with XhoI then probed to detect COs and DSBs; mw1 represents the <t>λ-HindIII</t> molecular weight marker (Fermentas) and mw2 represents the 1-kb molecular weight marker (Fermentas). The full-sized Southern blots are presented in Figure S1 . For graphs (D–G), wild type, ygl250wΔ , and soh1Δ are represented by black diamonds, black circles, and white squares, respectively. The corresponding XhoI-digested Southern blots are presented in Figure 3 C. (D) Pre-meiotic DNA replication was assessed for synchronized meiotic cultures by fluorescence-activated cell sorting (FACS) and the change from 2c to 4c DNA content was plotted over time. (E) Nuclear divisions (MI and MII) of the synchronized meiotic cultures in (E) were assessed with fluorescence microscopy using 4′,6-diamidino-2-phenylindole (DAPI) staining to visualize nuclear division. (F) Molecular analysis for DSB (DSB1) signal/total lane signal from Southern blots of DNA extracted from synchronized meiotic cultures. (G) Molecular analysis for CO (CO2) signal/total lane signal from Southern blots of DNA extracted from synchronized meiotic cultures.
    Lamda Hindiii, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 86/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    86
    Thermo Fisher hindiii digested puc18
    Assessment of YGL250W , SOH1 , and BRE5 (A) Schematic representation of Chromosome II from the diploid W303 background which consists of two LYS2 heteroalleles ( lys2–5′nde I − and lys2–3′nde I − ). These were used to measure meiotic gene conversion (see Materials and Methods ). (B) Spot test of wild type, ygl250wΔ , soh1Δ , and bre5Δ on haploid selection plates and haploid selection plates without lysine to measure meiotic gene conversion. The reduction in meiotic gene conversion of ygl250wΔ , soh1Δ , and bre5Δ was further assessed by random spore analysis ( Table 1 ). (C) Southern blot of DNA isolated from wild type, soh1Δ , and ygl250wΔ SK1 strains containing the ectopic URA3 - ARG4 interval on Chromosome III. The DNA from the indicated times after initiation of sporulation were digested with XhoI then probed to detect COs and DSBs; mw1 represents the <t>λ-HindIII</t> molecular weight marker (Fermentas) and mw2 represents the 1-kb molecular weight marker (Fermentas). The full-sized Southern blots are presented in Figure S1 . For graphs (D–G), wild type, ygl250wΔ , and soh1Δ are represented by black diamonds, black circles, and white squares, respectively. The corresponding XhoI-digested Southern blots are presented in Figure 3 C. (D) Pre-meiotic DNA replication was assessed for synchronized meiotic cultures by fluorescence-activated cell sorting (FACS) and the change from 2c to 4c DNA content was plotted over time. (E) Nuclear divisions (MI and MII) of the synchronized meiotic cultures in (E) were assessed with fluorescence microscopy using 4′,6-diamidino-2-phenylindole (DAPI) staining to visualize nuclear division. (F) Molecular analysis for DSB (DSB1) signal/total lane signal from Southern blots of DNA extracted from synchronized meiotic cultures. (G) Molecular analysis for CO (CO2) signal/total lane signal from Southern blots of DNA extracted from synchronized meiotic cultures.
    Hindiii Digested Puc18, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 86/100, based on 9 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher xbai hindiii digested pcdna3 1zeo
    Assessment of YGL250W , SOH1 , and BRE5 (A) Schematic representation of Chromosome II from the diploid W303 background which consists of two LYS2 heteroalleles ( lys2–5′nde I − and lys2–3′nde I − ). These were used to measure meiotic gene conversion (see Materials and Methods ). (B) Spot test of wild type, ygl250wΔ , soh1Δ , and bre5Δ on haploid selection plates and haploid selection plates without lysine to measure meiotic gene conversion. The reduction in meiotic gene conversion of ygl250wΔ , soh1Δ , and bre5Δ was further assessed by random spore analysis ( Table 1 ). (C) Southern blot of DNA isolated from wild type, soh1Δ , and ygl250wΔ SK1 strains containing the ectopic URA3 - ARG4 interval on Chromosome III. The DNA from the indicated times after initiation of sporulation were digested with XhoI then probed to detect COs and DSBs; mw1 represents the <t>λ-HindIII</t> molecular weight marker (Fermentas) and mw2 represents the 1-kb molecular weight marker (Fermentas). The full-sized Southern blots are presented in Figure S1 . For graphs (D–G), wild type, ygl250wΔ , and soh1Δ are represented by black diamonds, black circles, and white squares, respectively. The corresponding XhoI-digested Southern blots are presented in Figure 3 C. (D) Pre-meiotic DNA replication was assessed for synchronized meiotic cultures by fluorescence-activated cell sorting (FACS) and the change from 2c to 4c DNA content was plotted over time. (E) Nuclear divisions (MI and MII) of the synchronized meiotic cultures in (E) were assessed with fluorescence microscopy using 4′,6-diamidino-2-phenylindole (DAPI) staining to visualize nuclear division. (F) Molecular analysis for DSB (DSB1) signal/total lane signal from Southern blots of DNA extracted from synchronized meiotic cultures. (G) Molecular analysis for CO (CO2) signal/total lane signal from Southern blots of DNA extracted from synchronized meiotic cultures.
    Xbai Hindiii Digested Pcdna3 1zeo, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 86/100, based on 9 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher ndei hindiii
    Assessment of YGL250W , SOH1 , and BRE5 (A) Schematic representation of Chromosome II from the diploid W303 background which consists of two LYS2 heteroalleles ( lys2–5′nde I − and lys2–3′nde I − ). These were used to measure meiotic gene conversion (see Materials and Methods ). (B) Spot test of wild type, ygl250wΔ , soh1Δ , and bre5Δ on haploid selection plates and haploid selection plates without lysine to measure meiotic gene conversion. The reduction in meiotic gene conversion of ygl250wΔ , soh1Δ , and bre5Δ was further assessed by random spore analysis ( Table 1 ). (C) Southern blot of DNA isolated from wild type, soh1Δ , and ygl250wΔ SK1 strains containing the ectopic URA3 - ARG4 interval on Chromosome III. The DNA from the indicated times after initiation of sporulation were digested with XhoI then probed to detect COs and DSBs; mw1 represents the <t>λ-HindIII</t> molecular weight marker (Fermentas) and mw2 represents the 1-kb molecular weight marker (Fermentas). The full-sized Southern blots are presented in Figure S1 . For graphs (D–G), wild type, ygl250wΔ , and soh1Δ are represented by black diamonds, black circles, and white squares, respectively. The corresponding XhoI-digested Southern blots are presented in Figure 3 C. (D) Pre-meiotic DNA replication was assessed for synchronized meiotic cultures by fluorescence-activated cell sorting (FACS) and the change from 2c to 4c DNA content was plotted over time. (E) Nuclear divisions (MI and MII) of the synchronized meiotic cultures in (E) were assessed with fluorescence microscopy using 4′,6-diamidino-2-phenylindole (DAPI) staining to visualize nuclear division. (F) Molecular analysis for DSB (DSB1) signal/total lane signal from Southern blots of DNA extracted from synchronized meiotic cultures. (G) Molecular analysis for CO (CO2) signal/total lane signal from Southern blots of DNA extracted from synchronized meiotic cultures.
    Ndei Hindiii, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 91/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher xbai hindiii digested prset a
    Assessment of YGL250W , SOH1 , and BRE5 (A) Schematic representation of Chromosome II from the diploid W303 background which consists of two LYS2 heteroalleles ( lys2–5′nde I − and lys2–3′nde I − ). These were used to measure meiotic gene conversion (see Materials and Methods ). (B) Spot test of wild type, ygl250wΔ , soh1Δ , and bre5Δ on haploid selection plates and haploid selection plates without lysine to measure meiotic gene conversion. The reduction in meiotic gene conversion of ygl250wΔ , soh1Δ , and bre5Δ was further assessed by random spore analysis ( Table 1 ). (C) Southern blot of DNA isolated from wild type, soh1Δ , and ygl250wΔ SK1 strains containing the ectopic URA3 - ARG4 interval on Chromosome III. The DNA from the indicated times after initiation of sporulation were digested with XhoI then probed to detect COs and DSBs; mw1 represents the <t>λ-HindIII</t> molecular weight marker (Fermentas) and mw2 represents the 1-kb molecular weight marker (Fermentas). The full-sized Southern blots are presented in Figure S1 . For graphs (D–G), wild type, ygl250wΔ , and soh1Δ are represented by black diamonds, black circles, and white squares, respectively. The corresponding XhoI-digested Southern blots are presented in Figure 3 C. (D) Pre-meiotic DNA replication was assessed for synchronized meiotic cultures by fluorescence-activated cell sorting (FACS) and the change from 2c to 4c DNA content was plotted over time. (E) Nuclear divisions (MI and MII) of the synchronized meiotic cultures in (E) were assessed with fluorescence microscopy using 4′,6-diamidino-2-phenylindole (DAPI) staining to visualize nuclear division. (F) Molecular analysis for DSB (DSB1) signal/total lane signal from Southern blots of DNA extracted from synchronized meiotic cultures. (G) Molecular analysis for CO (CO2) signal/total lane signal from Southern blots of DNA extracted from synchronized meiotic cultures.
    Xbai Hindiii Digested Prset A, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 93/100, based on 7 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Thermo Fisher xhol hindiii
    Assessment of YGL250W , SOH1 , and BRE5 (A) Schematic representation of Chromosome II from the diploid W303 background which consists of two LYS2 heteroalleles ( lys2–5′nde I − and lys2–3′nde I − ). These were used to measure meiotic gene conversion (see Materials and Methods ). (B) Spot test of wild type, ygl250wΔ , soh1Δ , and bre5Δ on haploid selection plates and haploid selection plates without lysine to measure meiotic gene conversion. The reduction in meiotic gene conversion of ygl250wΔ , soh1Δ , and bre5Δ was further assessed by random spore analysis ( Table 1 ). (C) Southern blot of DNA isolated from wild type, soh1Δ , and ygl250wΔ SK1 strains containing the ectopic URA3 - ARG4 interval on Chromosome III. The DNA from the indicated times after initiation of sporulation were digested with XhoI then probed to detect COs and DSBs; mw1 represents the <t>λ-HindIII</t> molecular weight marker (Fermentas) and mw2 represents the 1-kb molecular weight marker (Fermentas). The full-sized Southern blots are presented in Figure S1 . For graphs (D–G), wild type, ygl250wΔ , and soh1Δ are represented by black diamonds, black circles, and white squares, respectively. The corresponding XhoI-digested Southern blots are presented in Figure 3 C. (D) Pre-meiotic DNA replication was assessed for synchronized meiotic cultures by fluorescence-activated cell sorting (FACS) and the change from 2c to 4c DNA content was plotted over time. (E) Nuclear divisions (MI and MII) of the synchronized meiotic cultures in (E) were assessed with fluorescence microscopy using 4′,6-diamidino-2-phenylindole (DAPI) staining to visualize nuclear division. (F) Molecular analysis for DSB (DSB1) signal/total lane signal from Southern blots of DNA extracted from synchronized meiotic cultures. (G) Molecular analysis for CO (CO2) signal/total lane signal from Southern blots of DNA extracted from synchronized meiotic cultures.
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    Thermo Fisher hindiii kpni cut pcdna5 frt to
    Assessment of YGL250W , SOH1 , and BRE5 (A) Schematic representation of Chromosome II from the diploid W303 background which consists of two LYS2 heteroalleles ( lys2–5′nde I − and lys2–3′nde I − ). These were used to measure meiotic gene conversion (see Materials and Methods ). (B) Spot test of wild type, ygl250wΔ , soh1Δ , and bre5Δ on haploid selection plates and haploid selection plates without lysine to measure meiotic gene conversion. The reduction in meiotic gene conversion of ygl250wΔ , soh1Δ , and bre5Δ was further assessed by random spore analysis ( Table 1 ). (C) Southern blot of DNA isolated from wild type, soh1Δ , and ygl250wΔ SK1 strains containing the ectopic URA3 - ARG4 interval on Chromosome III. The DNA from the indicated times after initiation of sporulation were digested with XhoI then probed to detect COs and DSBs; mw1 represents the <t>λ-HindIII</t> molecular weight marker (Fermentas) and mw2 represents the 1-kb molecular weight marker (Fermentas). The full-sized Southern blots are presented in Figure S1 . For graphs (D–G), wild type, ygl250wΔ , and soh1Δ are represented by black diamonds, black circles, and white squares, respectively. The corresponding XhoI-digested Southern blots are presented in Figure 3 C. (D) Pre-meiotic DNA replication was assessed for synchronized meiotic cultures by fluorescence-activated cell sorting (FACS) and the change from 2c to 4c DNA content was plotted over time. (E) Nuclear divisions (MI and MII) of the synchronized meiotic cultures in (E) were assessed with fluorescence microscopy using 4′,6-diamidino-2-phenylindole (DAPI) staining to visualize nuclear division. (F) Molecular analysis for DSB (DSB1) signal/total lane signal from Southern blots of DNA extracted from synchronized meiotic cultures. (G) Molecular analysis for CO (CO2) signal/total lane signal from Southern blots of DNA extracted from synchronized meiotic cultures.
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    Thermo Fisher bamhi hindiii digested prset a
    Assessment of YGL250W , SOH1 , and BRE5 (A) Schematic representation of Chromosome II from the diploid W303 background which consists of two LYS2 heteroalleles ( lys2–5′nde I − and lys2–3′nde I − ). These were used to measure meiotic gene conversion (see Materials and Methods ). (B) Spot test of wild type, ygl250wΔ , soh1Δ , and bre5Δ on haploid selection plates and haploid selection plates without lysine to measure meiotic gene conversion. The reduction in meiotic gene conversion of ygl250wΔ , soh1Δ , and bre5Δ was further assessed by random spore analysis ( Table 1 ). (C) Southern blot of DNA isolated from wild type, soh1Δ , and ygl250wΔ SK1 strains containing the ectopic URA3 - ARG4 interval on Chromosome III. The DNA from the indicated times after initiation of sporulation were digested with XhoI then probed to detect COs and DSBs; mw1 represents the <t>λ-HindIII</t> molecular weight marker (Fermentas) and mw2 represents the 1-kb molecular weight marker (Fermentas). The full-sized Southern blots are presented in Figure S1 . For graphs (D–G), wild type, ygl250wΔ , and soh1Δ are represented by black diamonds, black circles, and white squares, respectively. The corresponding XhoI-digested Southern blots are presented in Figure 3 C. (D) Pre-meiotic DNA replication was assessed for synchronized meiotic cultures by fluorescence-activated cell sorting (FACS) and the change from 2c to 4c DNA content was plotted over time. (E) Nuclear divisions (MI and MII) of the synchronized meiotic cultures in (E) were assessed with fluorescence microscopy using 4′,6-diamidino-2-phenylindole (DAPI) staining to visualize nuclear division. (F) Molecular analysis for DSB (DSB1) signal/total lane signal from Southern blots of DNA extracted from synchronized meiotic cultures. (G) Molecular analysis for CO (CO2) signal/total lane signal from Southern blots of DNA extracted from synchronized meiotic cultures.
    Bamhi Hindiii Digested Prset A, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 94/100, based on 10 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher hindiii xbai digested pyes2
    Assessment of YGL250W , SOH1 , and BRE5 (A) Schematic representation of Chromosome II from the diploid W303 background which consists of two LYS2 heteroalleles ( lys2–5′nde I − and lys2–3′nde I − ). These were used to measure meiotic gene conversion (see Materials and Methods ). (B) Spot test of wild type, ygl250wΔ , soh1Δ , and bre5Δ on haploid selection plates and haploid selection plates without lysine to measure meiotic gene conversion. The reduction in meiotic gene conversion of ygl250wΔ , soh1Δ , and bre5Δ was further assessed by random spore analysis ( Table 1 ). (C) Southern blot of DNA isolated from wild type, soh1Δ , and ygl250wΔ SK1 strains containing the ectopic URA3 - ARG4 interval on Chromosome III. The DNA from the indicated times after initiation of sporulation were digested with XhoI then probed to detect COs and DSBs; mw1 represents the <t>λ-HindIII</t> molecular weight marker (Fermentas) and mw2 represents the 1-kb molecular weight marker (Fermentas). The full-sized Southern blots are presented in Figure S1 . For graphs (D–G), wild type, ygl250wΔ , and soh1Δ are represented by black diamonds, black circles, and white squares, respectively. The corresponding XhoI-digested Southern blots are presented in Figure 3 C. (D) Pre-meiotic DNA replication was assessed for synchronized meiotic cultures by fluorescence-activated cell sorting (FACS) and the change from 2c to 4c DNA content was plotted over time. (E) Nuclear divisions (MI and MII) of the synchronized meiotic cultures in (E) were assessed with fluorescence microscopy using 4′,6-diamidino-2-phenylindole (DAPI) staining to visualize nuclear division. (F) Molecular analysis for DSB (DSB1) signal/total lane signal from Southern blots of DNA extracted from synchronized meiotic cultures. (G) Molecular analysis for CO (CO2) signal/total lane signal from Southern blots of DNA extracted from synchronized meiotic cultures.
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    Assessment of YGL250W , SOH1 , and BRE5 (A) Schematic representation of Chromosome II from the diploid W303 background which consists of two LYS2 heteroalleles ( lys2–5′nde I − and lys2–3′nde I − ). These were used to measure meiotic gene conversion (see Materials and Methods ). (B) Spot test of wild type, ygl250wΔ , soh1Δ , and bre5Δ on haploid selection plates and haploid selection plates without lysine to measure meiotic gene conversion. The reduction in meiotic gene conversion of ygl250wΔ , soh1Δ , and bre5Δ was further assessed by random spore analysis ( Table 1 ). (C) Southern blot of DNA isolated from wild type, soh1Δ , and ygl250wΔ SK1 strains containing the ectopic URA3 - ARG4 interval on Chromosome III. The DNA from the indicated times after initiation of sporulation were digested with XhoI then probed to detect COs and DSBs; mw1 represents the <t>λ-HindIII</t> molecular weight marker (Fermentas) and mw2 represents the 1-kb molecular weight marker (Fermentas). The full-sized Southern blots are presented in Figure S1 . For graphs (D–G), wild type, ygl250wΔ , and soh1Δ are represented by black diamonds, black circles, and white squares, respectively. The corresponding XhoI-digested Southern blots are presented in Figure 3 C. (D) Pre-meiotic DNA replication was assessed for synchronized meiotic cultures by fluorescence-activated cell sorting (FACS) and the change from 2c to 4c DNA content was plotted over time. (E) Nuclear divisions (MI and MII) of the synchronized meiotic cultures in (E) were assessed with fluorescence microscopy using 4′,6-diamidino-2-phenylindole (DAPI) staining to visualize nuclear division. (F) Molecular analysis for DSB (DSB1) signal/total lane signal from Southern blots of DNA extracted from synchronized meiotic cultures. (G) Molecular analysis for CO (CO2) signal/total lane signal from Southern blots of DNA extracted from synchronized meiotic cultures.
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    Assessment of YGL250W , SOH1 , and BRE5 (A) Schematic representation of Chromosome II from the diploid W303 background which consists of two LYS2 heteroalleles ( lys2–5′nde I − and lys2–3′nde I − ). These were used to measure meiotic gene conversion (see Materials and Methods ). (B) Spot test of wild type, ygl250wΔ , soh1Δ , and bre5Δ on haploid selection plates and haploid selection plates without lysine to measure meiotic gene conversion. The reduction in meiotic gene conversion of ygl250wΔ , soh1Δ , and bre5Δ was further assessed by random spore analysis ( Table 1 ). (C) Southern blot of DNA isolated from wild type, soh1Δ , and ygl250wΔ SK1 strains containing the ectopic URA3 - ARG4 interval on Chromosome III. The DNA from the indicated times after initiation of sporulation were digested with XhoI then probed to detect COs and DSBs; mw1 represents the <t>λ-HindIII</t> molecular weight marker (Fermentas) and mw2 represents the 1-kb molecular weight marker (Fermentas). The full-sized Southern blots are presented in Figure S1 . For graphs (D–G), wild type, ygl250wΔ , and soh1Δ are represented by black diamonds, black circles, and white squares, respectively. The corresponding XhoI-digested Southern blots are presented in Figure 3 C. (D) Pre-meiotic DNA replication was assessed for synchronized meiotic cultures by fluorescence-activated cell sorting (FACS) and the change from 2c to 4c DNA content was plotted over time. (E) Nuclear divisions (MI and MII) of the synchronized meiotic cultures in (E) were assessed with fluorescence microscopy using 4′,6-diamidino-2-phenylindole (DAPI) staining to visualize nuclear division. (F) Molecular analysis for DSB (DSB1) signal/total lane signal from Southern blots of DNA extracted from synchronized meiotic cultures. (G) Molecular analysis for CO (CO2) signal/total lane signal from Southern blots of DNA extracted from synchronized meiotic cultures.
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    Publicly available caucasian control individual genotyping frequencies versus pooling frequencies for <t>Affymetrix</t> <t>Genechip</t> <t>HindIII</t> and Illumina HumanHap300 arrays. The data are the 15 645 SNPs in common between the Affymetrix Genechip HindIII and Illumina HumanHap300 arrays. The frequency of the sample of 271 publicly available caucasian controls is on the y -axis, with the pooling frequencies from the N =384 pooled case/controls on the x -axis. The broken line is y = x . The solid line is the regression line.
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    Publicly available caucasian control individual genotyping frequencies versus pooling frequencies for <t>Affymetrix</t> <t>Genechip</t> <t>HindIII</t> and Illumina HumanHap300 arrays. The data are the 15 645 SNPs in common between the Affymetrix Genechip HindIII and Illumina HumanHap300 arrays. The frequency of the sample of 271 publicly available caucasian controls is on the y -axis, with the pooling frequencies from the N =384 pooled case/controls on the x -axis. The broken line is y = x . The solid line is the regression line.
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    Publicly available caucasian control individual genotyping frequencies versus pooling frequencies for <t>Affymetrix</t> <t>Genechip</t> <t>HindIII</t> and Illumina HumanHap300 arrays. The data are the 15 645 SNPs in common between the Affymetrix Genechip HindIII and Illumina HumanHap300 arrays. The frequency of the sample of 271 publicly available caucasian controls is on the y -axis, with the pooling frequencies from the N =384 pooled case/controls on the x -axis. The broken line is y = x . The solid line is the regression line.
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    Publicly available caucasian control individual genotyping frequencies versus pooling frequencies for <t>Affymetrix</t> <t>Genechip</t> <t>HindIII</t> and Illumina HumanHap300 arrays. The data are the 15 645 SNPs in common between the Affymetrix Genechip HindIII and Illumina HumanHap300 arrays. The frequency of the sample of 271 publicly available caucasian controls is on the y -axis, with the pooling frequencies from the N =384 pooled case/controls on the x -axis. The broken line is y = x . The solid line is the regression line.
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    Publicly available caucasian control individual genotyping frequencies versus pooling frequencies for <t>Affymetrix</t> <t>Genechip</t> <t>HindIII</t> and Illumina HumanHap300 arrays. The data are the 15 645 SNPs in common between the Affymetrix Genechip HindIII and Illumina HumanHap300 arrays. The frequency of the sample of 271 publicly available caucasian controls is on the y -axis, with the pooling frequencies from the N =384 pooled case/controls on the x -axis. The broken line is y = x . The solid line is the regression line.
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    Publicly available caucasian control individual genotyping frequencies versus pooling frequencies for <t>Affymetrix</t> <t>Genechip</t> <t>HindIII</t> and Illumina HumanHap300 arrays. The data are the 15 645 SNPs in common between the Affymetrix Genechip HindIII and Illumina HumanHap300 arrays. The frequency of the sample of 271 publicly available caucasian controls is on the y -axis, with the pooling frequencies from the N =384 pooled case/controls on the x -axis. The broken line is y = x . The solid line is the regression line.
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    Some Twinkle mutants cause replication stalling. 2DNAGE samples for all panels consisted of purified <t>mtDNA</t> digested with <t>HincII</t> and probed with a radiolabeled cytochrome b gene fragment (nt 14 846–15 357). The detected fragment includes the non-coding region of mtDNA also including the cytochrome b , ND6 , part of the ND5 gene and intervening tRNA genes (nt 13 636–1006). ( A ) The first two panels on the left show the RIs of HEK293 cells and the interpretation based primarily on earlier 2DNAGE analysis of mtDNA RIs ( 7 , 9 ). Abbreviations: 1n, 3.9 kb non-replicating HincII fragment. (b) bubble arcs. MtDNA bubble arcs are usually very sensitive to RNase H due to the presence of patches of RNA–DNA especially on the lagging strand; these therefore also fall in the category of RITOLS as do various other RIs. Here, y and y′ indicate ascending and descending parts of the y arc and (dy) indicates double-Y structures. These will eventually form resolution intermediates resembling Holliday junctions (HJL-Holliday junction like molecules). The two panels on the right show a comparison of RIs of non-induced and fully induced cells expressing Twinkle wt. The only notable difference in this case is a reproducible reduction in one of the HJL RIs as indicated. ( B ) A normal pattern of RIs similar to non-expressing cells was observed in cells expressing Twinky or the ▵Linker variant. ( C ) K421A, G575D and ▵70–343 show similar patterns of replication stalling, with increased bubble (b) and descending Y-arc (y′) intensities, a sharpening and lengthening of the bubble arc and loss of RITOLS (ovals). The right-most panel shows the same exposure 2D gel pattern of the non-induced ▵70–343 line showing the typical HincII fragment pattern, including abundant RITOLS. ( D ) A limited S1 digestion illustrates that stalled RIs observed in panel C are S1 insensitive (right two panels). The effectiveness of the S1 treatment is illustrated by the left two panels, showing the effect on Twinkle non-induced cells. Similar to the S1 treatment RNase H treatment shows that the stalling RIs observed with Twinkle mutants are largely insensitive to this enzyme (Supplementary Figure 2), showing that the observed stalled RIs in panel C are essentially dsDNA. Although the intensities in subfigures A–D cannot be directly compared due to differences in exposure time, each panel contains appropriate controls of similar exposure. For example, the exposures of the left two panels in C have been chosen to be similar in comparison to the right-most panels to properly illustrate the severity of the stalling phenotype.
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    Some Twinkle mutants cause replication stalling. 2DNAGE samples for all panels consisted of purified <t>mtDNA</t> digested with <t>HincII</t> and probed with a radiolabeled cytochrome b gene fragment (nt 14 846–15 357). The detected fragment includes the non-coding region of mtDNA also including the cytochrome b , ND6 , part of the ND5 gene and intervening tRNA genes (nt 13 636–1006). ( A ) The first two panels on the left show the RIs of HEK293 cells and the interpretation based primarily on earlier 2DNAGE analysis of mtDNA RIs ( 7 , 9 ). Abbreviations: 1n, 3.9 kb non-replicating HincII fragment. (b) bubble arcs. MtDNA bubble arcs are usually very sensitive to RNase H due to the presence of patches of RNA–DNA especially on the lagging strand; these therefore also fall in the category of RITOLS as do various other RIs. Here, y and y′ indicate ascending and descending parts of the y arc and (dy) indicates double-Y structures. These will eventually form resolution intermediates resembling Holliday junctions (HJL-Holliday junction like molecules). The two panels on the right show a comparison of RIs of non-induced and fully induced cells expressing Twinkle wt. The only notable difference in this case is a reproducible reduction in one of the HJL RIs as indicated. ( B ) A normal pattern of RIs similar to non-expressing cells was observed in cells expressing Twinky or the ▵Linker variant. ( C ) K421A, G575D and ▵70–343 show similar patterns of replication stalling, with increased bubble (b) and descending Y-arc (y′) intensities, a sharpening and lengthening of the bubble arc and loss of RITOLS (ovals). The right-most panel shows the same exposure 2D gel pattern of the non-induced ▵70–343 line showing the typical HincII fragment pattern, including abundant RITOLS. ( D ) A limited S1 digestion illustrates that stalled RIs observed in panel C are S1 insensitive (right two panels). The effectiveness of the S1 treatment is illustrated by the left two panels, showing the effect on Twinkle non-induced cells. Similar to the S1 treatment RNase H treatment shows that the stalling RIs observed with Twinkle mutants are largely insensitive to this enzyme (Supplementary Figure 2), showing that the observed stalled RIs in panel C are essentially dsDNA. Although the intensities in subfigures A–D cannot be directly compared due to differences in exposure time, each panel contains appropriate controls of similar exposure. For example, the exposures of the left two panels in C have been chosen to be similar in comparison to the right-most panels to properly illustrate the severity of the stalling phenotype.
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    Some Twinkle mutants cause replication stalling. 2DNAGE samples for all panels consisted of purified <t>mtDNA</t> digested with <t>HincII</t> and probed with a radiolabeled cytochrome b gene fragment (nt 14 846–15 357). The detected fragment includes the non-coding region of mtDNA also including the cytochrome b , ND6 , part of the ND5 gene and intervening tRNA genes (nt 13 636–1006). ( A ) The first two panels on the left show the RIs of HEK293 cells and the interpretation based primarily on earlier 2DNAGE analysis of mtDNA RIs ( 7 , 9 ). Abbreviations: 1n, 3.9 kb non-replicating HincII fragment. (b) bubble arcs. MtDNA bubble arcs are usually very sensitive to RNase H due to the presence of patches of RNA–DNA especially on the lagging strand; these therefore also fall in the category of RITOLS as do various other RIs. Here, y and y′ indicate ascending and descending parts of the y arc and (dy) indicates double-Y structures. These will eventually form resolution intermediates resembling Holliday junctions (HJL-Holliday junction like molecules). The two panels on the right show a comparison of RIs of non-induced and fully induced cells expressing Twinkle wt. The only notable difference in this case is a reproducible reduction in one of the HJL RIs as indicated. ( B ) A normal pattern of RIs similar to non-expressing cells was observed in cells expressing Twinky or the ▵Linker variant. ( C ) K421A, G575D and ▵70–343 show similar patterns of replication stalling, with increased bubble (b) and descending Y-arc (y′) intensities, a sharpening and lengthening of the bubble arc and loss of RITOLS (ovals). The right-most panel shows the same exposure 2D gel pattern of the non-induced ▵70–343 line showing the typical HincII fragment pattern, including abundant RITOLS. ( D ) A limited S1 digestion illustrates that stalled RIs observed in panel C are S1 insensitive (right two panels). The effectiveness of the S1 treatment is illustrated by the left two panels, showing the effect on Twinkle non-induced cells. Similar to the S1 treatment RNase H treatment shows that the stalling RIs observed with Twinkle mutants are largely insensitive to this enzyme (Supplementary Figure 2), showing that the observed stalled RIs in panel C are essentially dsDNA. Although the intensities in subfigures A–D cannot be directly compared due to differences in exposure time, each panel contains appropriate controls of similar exposure. For example, the exposures of the left two panels in C have been chosen to be similar in comparison to the right-most panels to properly illustrate the severity of the stalling phenotype.
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    Some Twinkle mutants cause replication stalling. 2DNAGE samples for all panels consisted of purified <t>mtDNA</t> digested with <t>HincII</t> and probed with a radiolabeled cytochrome b gene fragment (nt 14 846–15 357). The detected fragment includes the non-coding region of mtDNA also including the cytochrome b , ND6 , part of the ND5 gene and intervening tRNA genes (nt 13 636–1006). ( A ) The first two panels on the left show the RIs of HEK293 cells and the interpretation based primarily on earlier 2DNAGE analysis of mtDNA RIs ( 7 , 9 ). Abbreviations: 1n, 3.9 kb non-replicating HincII fragment. (b) bubble arcs. MtDNA bubble arcs are usually very sensitive to RNase H due to the presence of patches of RNA–DNA especially on the lagging strand; these therefore also fall in the category of RITOLS as do various other RIs. Here, y and y′ indicate ascending and descending parts of the y arc and (dy) indicates double-Y structures. These will eventually form resolution intermediates resembling Holliday junctions (HJL-Holliday junction like molecules). The two panels on the right show a comparison of RIs of non-induced and fully induced cells expressing Twinkle wt. The only notable difference in this case is a reproducible reduction in one of the HJL RIs as indicated. ( B ) A normal pattern of RIs similar to non-expressing cells was observed in cells expressing Twinky or the ▵Linker variant. ( C ) K421A, G575D and ▵70–343 show similar patterns of replication stalling, with increased bubble (b) and descending Y-arc (y′) intensities, a sharpening and lengthening of the bubble arc and loss of RITOLS (ovals). The right-most panel shows the same exposure 2D gel pattern of the non-induced ▵70–343 line showing the typical HincII fragment pattern, including abundant RITOLS. ( D ) A limited S1 digestion illustrates that stalled RIs observed in panel C are S1 insensitive (right two panels). The effectiveness of the S1 treatment is illustrated by the left two panels, showing the effect on Twinkle non-induced cells. Similar to the S1 treatment RNase H treatment shows that the stalling RIs observed with Twinkle mutants are largely insensitive to this enzyme (Supplementary Figure 2), showing that the observed stalled RIs in panel C are essentially dsDNA. Although the intensities in subfigures A–D cannot be directly compared due to differences in exposure time, each panel contains appropriate controls of similar exposure. For example, the exposures of the left two panels in C have been chosen to be similar in comparison to the right-most panels to properly illustrate the severity of the stalling phenotype.
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    Image Search Results


    Representative RFLP patterns of the amplified 1100 bp fragment of actin gene obtained from T. vaginalis isolates. Lane 1, 5 and 8 digested by HindII ; Lane 2, 6 and 9 digested by Tru1 I; Lane 3, 7 and 10 digested by RsaI ; M: DNA ladder (50 bp)

    Journal: Iranian Journal of Parasitology

    Article Title: Genotyping, Drug Susceptibility and Prevalence Survey of Trichomonas vaginalis among Women Attending Gynecology Clinics in Hamadan, Western Iran, in 2014–2015

    doi:

    Figure Lengend Snippet: Representative RFLP patterns of the amplified 1100 bp fragment of actin gene obtained from T. vaginalis isolates. Lane 1, 5 and 8 digested by HindII ; Lane 2, 6 and 9 digested by Tru1 I; Lane 3, 7 and 10 digested by RsaI ; M: DNA ladder (50 bp)

    Article Snippet: RFLP analysis was performed by three restriction enzyme, HindII , Tru1I and RsaI (Fermentas, Thermo Scientific, USA), according to the manufacturer’s instructions.

    Techniques: Amplification

    The pAVOIAF{#1–#2–#3–#4} vector. ( A ) Vector map of pAVOIAF{#1–#2–#3–#4}. The vector is based on the pUC57-Kan vector, from which only the kanamycin resistance cassette and the origin of replication remain. The four-slot cloning site together with the 3’ and 5’ piggyBac terminal repeats is located between the AatII and PciI sites. The light gray band on the inside indicates the transgene. ( B ) Scheme of the four-slot cloning site. Each slot consists of an 18 bp spacer that translates into the amino acids Phe-Arg-Glu-Asp-Asp-Tyr and thus was termed FREDDY spacer. The slots can be accessed individually by unique restriction enzyme site pairs (XmaI/SpeI for #1, HindIII/XbaI for #2, XhoI/NheI for #3 and AflII/AvrII for #4). They are embedded into five PmeI restriction enzyme sites that allow a simple one-enzyme control digestion to determine the size of the sequences that were inserted into the slots. For convenience, the downstream restriction enzyme sites for each slot (SpeI for #1, XbaI for #2, NheI for #3 and AvrII #4) result in identical sticky ends, facilitating cloning procedures that cannot utilize the suggested restriction enzyme site pairs. Extends of the genetic elements are not to scale. ORF, open-reading frame.

    Journal: eLife

    Article Title: A universal vector concept for a direct genotyping of transgenic organisms and a systematic creation of homozygous lines

    doi: 10.7554/eLife.31677

    Figure Lengend Snippet: The pAVOIAF{#1–#2–#3–#4} vector. ( A ) Vector map of pAVOIAF{#1–#2–#3–#4}. The vector is based on the pUC57-Kan vector, from which only the kanamycin resistance cassette and the origin of replication remain. The four-slot cloning site together with the 3’ and 5’ piggyBac terminal repeats is located between the AatII and PciI sites. The light gray band on the inside indicates the transgene. ( B ) Scheme of the four-slot cloning site. Each slot consists of an 18 bp spacer that translates into the amino acids Phe-Arg-Glu-Asp-Asp-Tyr and thus was termed FREDDY spacer. The slots can be accessed individually by unique restriction enzyme site pairs (XmaI/SpeI for #1, HindIII/XbaI for #2, XhoI/NheI for #3 and AflII/AvrII for #4). They are embedded into five PmeI restriction enzyme sites that allow a simple one-enzyme control digestion to determine the size of the sequences that were inserted into the slots. For convenience, the downstream restriction enzyme sites for each slot (SpeI for #1, XbaI for #2, NheI for #3 and AvrII #4) result in identical sticky ends, facilitating cloning procedures that cannot utilize the suggested restriction enzyme site pairs. Extends of the genetic elements are not to scale. ORF, open-reading frame.

    Article Snippet: Molecular biology: the pGS[#P’#O(LA)-mEmerald] and pAGOC{#P’#O(LA)-mEmerald} vectors A hybrid sequence, consisting of (i) a HindIII site, (ii) the modular fluorescent protein expression cassette as described above and (iii) a XbaI site, was de novo synthetized and inserted into the unique SfiI site of pMK-RQ (Thermo Fisher Scientific).

    Techniques: Plasmid Preparation, Clone Assay

    Analysis of plasmid copy number. Strains were retransformed with pSEL1 and pSEL2 encoding rhomboid-Rv1337 and were grown to mid-log phase. Samples were normalized based on OD 600 and were column purified along with HindIII-digested pUC19 plasmid as an internal purification control. The samples were then analyzed on an agarose gel.

    Journal: Protein Science : A Publication of the Protein Society

    Article Title: Genetic selection system for improving recombinant membrane protein expression in E. coli

    doi: 10.1002/pro.39

    Figure Lengend Snippet: Analysis of plasmid copy number. Strains were retransformed with pSEL1 and pSEL2 encoding rhomboid-Rv1337 and were grown to mid-log phase. Samples were normalized based on OD 600 and were column purified along with HindIII-digested pUC19 plasmid as an internal purification control. The samples were then analyzed on an agarose gel.

    Article Snippet: This fragment was then placed between the KpnI and HindIII sites in the MCS of pBAD/HisA (Invitrogen).

    Techniques: Plasmid Preparation, Purification, Agarose Gel Electrophoresis

    Chromatin conformation in GATA2 locus is significantly changed upon ATRA induction a Diagram showing ChIP-seq, ATAC-seq, and 4C-seq results of GATA2 promoter and upstream regions. (upper panel) HindIII cutting sites, gene annotation, and control-specific ATAC-seq peaks. Transcription factors with enriched motifs in the peak are labeled. (middle panel) Genome browser views of ChIP-seq and ATAC-seq results of control and ATRA-treated HL-60 cells; semitransparent cyan color labels the bait site used in 4C-seq, and semitransparent yellow color labels the putative enhancers recognized in 4C-seq. (lower panel) 4C-seq results of control and ATRA-treated HL-60 cells using the bait mentioned above. Red dots indicate 4C-seq RPMs in the ATRA-treated cells, and blue dots indicate RPMs in the control cells; only fragments with significant interactions with the bait are shown. Log2-foldchange of RPMs is shown in the lowest bar graph. b DNA FISH showing the loss of the chromatin loop between the GATA2 promoter and enhancer. BAC probes containing the promoter (red) and enhancer (green) were more separated in the ATRA-treated cells (right) than in the control cells (left). c Loci of promoter and enhancer interacted more in the control cells than in the ATRA-treated cells. p -Value was determined by Mann–Whitney U -test: ** p -value

    Journal: Cell Death & Disease

    Article Title: Alterations of specific chromatin conformation affect ATRA-induced leukemia cell differentiation

    doi: 10.1038/s41419-017-0173-6

    Figure Lengend Snippet: Chromatin conformation in GATA2 locus is significantly changed upon ATRA induction a Diagram showing ChIP-seq, ATAC-seq, and 4C-seq results of GATA2 promoter and upstream regions. (upper panel) HindIII cutting sites, gene annotation, and control-specific ATAC-seq peaks. Transcription factors with enriched motifs in the peak are labeled. (middle panel) Genome browser views of ChIP-seq and ATAC-seq results of control and ATRA-treated HL-60 cells; semitransparent cyan color labels the bait site used in 4C-seq, and semitransparent yellow color labels the putative enhancers recognized in 4C-seq. (lower panel) 4C-seq results of control and ATRA-treated HL-60 cells using the bait mentioned above. Red dots indicate 4C-seq RPMs in the ATRA-treated cells, and blue dots indicate RPMs in the control cells; only fragments with significant interactions with the bait are shown. Log2-foldchange of RPMs is shown in the lowest bar graph. b DNA FISH showing the loss of the chromatin loop between the GATA2 promoter and enhancer. BAC probes containing the promoter (red) and enhancer (green) were more separated in the ATRA-treated cells (right) than in the control cells (left). c Loci of promoter and enhancer interacted more in the control cells than in the ATRA-treated cells. p -Value was determined by Mann–Whitney U -test: ** p -value

    Article Snippet: HindIII digestion was performed at 37 °C overnight, and then, a proximity ligation was performed at 4 °C for 4 h. After reversing the cross-linking with a Proteinase K (Ambion, USA) treatment, the DNA was purified using a phenol–chloroform (Solarbio) extraction with ethanol precipitation.

    Techniques: Chromatin Immunoprecipitation, Labeling, Fluorescence In Situ Hybridization, BAC Assay, MANN-WHITNEY

    5′–3′ loop of ZBTB16 was lost upon ATRA induction a Diagram showing ChIP-seq, ATAC-seq and 4C-seq results of ZBTB16. (upper panel) HindIII cutting sites, gene annotation and control-specific ATAC-seq peaks. Transcription factors with enriched motifs in the peak are labeled. (middle panel) Genome browser views of ChIP-seq and ATAC-seq results of ATRA-treated and control HL-60 cells; semitransparent cyan color labels the bait site used in 4C-seq, and semitransparent yellow color labels the site with a strong interaction with the bait recognized in 4C-seq. (lower panel) 4C-seq results of ATRA-treated and control HL-60 cells using the bait mentioned above. Red dots indicate 4C-seq RPMs in the ATRA-treated cells, and blue dots indicate RPMs in the control cells; only fragments with significant interactions with the bait are shown. Log2-foldchange of RPMs are shown in the lowest bar graph. b Model depicting the change in the chromatin architecture around the Gata2 gene. c Model depicting the change in the chromatin architecture around the Zbtb16 gene

    Journal: Cell Death & Disease

    Article Title: Alterations of specific chromatin conformation affect ATRA-induced leukemia cell differentiation

    doi: 10.1038/s41419-017-0173-6

    Figure Lengend Snippet: 5′–3′ loop of ZBTB16 was lost upon ATRA induction a Diagram showing ChIP-seq, ATAC-seq and 4C-seq results of ZBTB16. (upper panel) HindIII cutting sites, gene annotation and control-specific ATAC-seq peaks. Transcription factors with enriched motifs in the peak are labeled. (middle panel) Genome browser views of ChIP-seq and ATAC-seq results of ATRA-treated and control HL-60 cells; semitransparent cyan color labels the bait site used in 4C-seq, and semitransparent yellow color labels the site with a strong interaction with the bait recognized in 4C-seq. (lower panel) 4C-seq results of ATRA-treated and control HL-60 cells using the bait mentioned above. Red dots indicate 4C-seq RPMs in the ATRA-treated cells, and blue dots indicate RPMs in the control cells; only fragments with significant interactions with the bait are shown. Log2-foldchange of RPMs are shown in the lowest bar graph. b Model depicting the change in the chromatin architecture around the Gata2 gene. c Model depicting the change in the chromatin architecture around the Zbtb16 gene

    Article Snippet: HindIII digestion was performed at 37 °C overnight, and then, a proximity ligation was performed at 4 °C for 4 h. After reversing the cross-linking with a Proteinase K (Ambion, USA) treatment, the DNA was purified using a phenol–chloroform (Solarbio) extraction with ethanol precipitation.

    Techniques: Chromatin Immunoprecipitation, Labeling

    (a) Schematic representation of recombinant vector containing designed construct and (b) 1% agarose gel electrophoresis of recombinant vector [Lane 1: recombinant vector digested by HindIII and SacI and Lane 2: 1 kb DNA ladder SM0331 (Fermentas, Lithuania)]

    Journal: Advanced Biomedical Research

    Article Title: In Vitro Evaluation of Vegf-Pseudomonas Exotoxin: A Conjugated on Tumor Cells

    doi: 10.4103/2277-9175.218691

    Figure Lengend Snippet: (a) Schematic representation of recombinant vector containing designed construct and (b) 1% agarose gel electrophoresis of recombinant vector [Lane 1: recombinant vector digested by HindIII and SacI and Lane 2: 1 kb DNA ladder SM0331 (Fermentas, Lithuania)]

    Article Snippet: The construct was digested by SacI and HindIII restriction enzymes (Thermo Scientific, Lithuania) and subcloned into pET-28a expression vector (MilliporeMillipore, USA) by T4 DNA ligase (Thermo Scientific, Lithuania).

    Techniques: Recombinant, Plasmid Preparation, Construct, Agarose Gel Electrophoresis

    Depletion of Mcl-1 downregulates HR and upregulates NHEJ. ( A ) Schematic diagram of HR and NHEJ reporter systems. HR reporter is composed of 2 defective GFP genes that can be rescued only by HR, resulting in GFP fluorescence. In the NHEJ reporter, GFP is interrupted by an adenoviral exon (Ad2) and can be restored upon HindIII digestion and NHEJ repair. ( B ) HR activity was compared in WT and Mcl1 –/– MEFs. Data represent the mean ± SD, n = 3 per group. ** P

    Journal: The Journal of Clinical Investigation

    Article Title: Targeting Mcl-1 enhances DNA replication stress sensitivity to cancer therapy

    doi: 10.1172/JCI92742

    Figure Lengend Snippet: Depletion of Mcl-1 downregulates HR and upregulates NHEJ. ( A ) Schematic diagram of HR and NHEJ reporter systems. HR reporter is composed of 2 defective GFP genes that can be rescued only by HR, resulting in GFP fluorescence. In the NHEJ reporter, GFP is interrupted by an adenoviral exon (Ad2) and can be restored upon HindIII digestion and NHEJ repair. ( B ) HR activity was compared in WT and Mcl1 –/– MEFs. Data represent the mean ± SD, n = 3 per group. ** P

    Article Snippet: First, NHEJ substrate GFP-Pem1-Ad2 plasmids were linearized by restriction enzyme HindIII (Thermo Fisher Scientific).

    Techniques: Non-Homologous End Joining, Fluorescence, Activity Assay

    Control digestions of recombinant plasmid DNA using HindIII (Fermentas) enzyme. (A) pIL253:PptcB:MOG35-55, (B) pIL253:PptcB:MBP85-97, (C) pIL253:PptcB:PLP139-151. Expected DNA fragments after restriction analysis for correct constructs: 3897 bp, 845 bp (marked by red arrows), 1kb DNA ladder (M) 1kb DNA Ladder (Fermentas) used as DNA size reference marker.

    Journal: Medical Science Monitor : International Medical Journal of Experimental and Clinical Research

    Article Title: Oral Administration of Lactococcus lactis Expressing Synthetic Genes of Myelin Antigens in Decreasing Experimental Autoimmune Encephalomyelitis in Rats

    doi: 10.12659/MSM.892764

    Figure Lengend Snippet: Control digestions of recombinant plasmid DNA using HindIII (Fermentas) enzyme. (A) pIL253:PptcB:MOG35-55, (B) pIL253:PptcB:MBP85-97, (C) pIL253:PptcB:PLP139-151. Expected DNA fragments after restriction analysis for correct constructs: 3897 bp, 845 bp (marked by red arrows), 1kb DNA ladder (M) 1kb DNA Ladder (Fermentas) used as DNA size reference marker.

    Article Snippet: From confirmed proper recombinant cells, plasmid DNA was isolated using the Plasmid Mini Kit (A & A Biotechnology) and subjected to restriction analysis with HindIII enzyme (Fermentas).

    Techniques: Recombinant, Plasmid Preparation, Construct, Marker

    Identification of multiple copies of IS 1630 in M. fermentans strains. Genomic DNA from M. fermentans PG18 (lanes 2 and 4) or II-29/1 (lanes 3 and 5) was digested with Eco RI (lanes 2 and 3) or Hin dIII (lanes 4 and 5), transferred to a nylon membrane, and hybridized with DIG-labeled oligonucleotide probe 5 (see Materials and Methods). Lane 1 contains DIG-labeled lambda Hin dIII markers (Boehringer), the sizes of which are indicated in kilobase pairs.

    Journal: Journal of Bacteriology

    Article Title: IS1630 of Mycoplasma fermentans, a Novel IS30-Type Insertion Element That Targets and Duplicates Inverted Repeats of Variable Length and Sequence during Insertion

    doi:

    Figure Lengend Snippet: Identification of multiple copies of IS 1630 in M. fermentans strains. Genomic DNA from M. fermentans PG18 (lanes 2 and 4) or II-29/1 (lanes 3 and 5) was digested with Eco RI (lanes 2 and 3) or Hin dIII (lanes 4 and 5), transferred to a nylon membrane, and hybridized with DIG-labeled oligonucleotide probe 5 (see Materials and Methods). Lane 1 contains DIG-labeled lambda Hin dIII markers (Boehringer), the sizes of which are indicated in kilobase pairs.

    Article Snippet: Bgl II-, Hin dIII-, or Xba I-digested chromosomal DNA from M. fermentans PG18 or Bgl II-digested total DNA from M. fermentans II-29/1 was ligated to Bam HI-, Hin dIII-, or Xba I-digested cloning vector pZero 2 or pZero2.1 under conditions recommended by the vector supplier (Invitrogen).

    Techniques: Labeling

    Confirmation of genomic composition of 3 independent recombinant VV-ΔTk viruses. ( A ) An ethidium bromide stained DNA gel of genomic HindIII restriction digests of viral DNA isolated from parental VV (Wyeth Strain), 3 clones of VV-ΔTk- yfp-gpt and 3 clones of VV-ΔTk'. Arrows indicate the Tk insertion site (VV-Wyeth), and the unique bands that result from insertion of the yfp-gpt cassette (VV-ΔTk- yfp-gpt ). ( B ) Southern hybridization of the DNA gel in A identifying the yfp insert present in the genome of the VV-ΔTk- yfp-gpt clones, but not in parental VV-Wyeth or the VV-ΔTk' clones. ( C ) DNAStar sequence alignment at the yfp-gpt insertion site of DNA isolated from the 3 VV-ΔTk' clones post Cre passage.

    Journal: PLoS ONE

    Article Title: A Selectable and Excisable Marker System for the Rapid Creation of Recombinant Poxviruses

    doi: 10.1371/journal.pone.0024643

    Figure Lengend Snippet: Confirmation of genomic composition of 3 independent recombinant VV-ΔTk viruses. ( A ) An ethidium bromide stained DNA gel of genomic HindIII restriction digests of viral DNA isolated from parental VV (Wyeth Strain), 3 clones of VV-ΔTk- yfp-gpt and 3 clones of VV-ΔTk'. Arrows indicate the Tk insertion site (VV-Wyeth), and the unique bands that result from insertion of the yfp-gpt cassette (VV-ΔTk- yfp-gpt ). ( B ) Southern hybridization of the DNA gel in A identifying the yfp insert present in the genome of the VV-ΔTk- yfp-gpt clones, but not in parental VV-Wyeth or the VV-ΔTk' clones. ( C ) DNAStar sequence alignment at the yfp-gpt insertion site of DNA isolated from the 3 VV-ΔTk' clones post Cre passage.

    Article Snippet: Briefly, 7 µg of DNA was digested overnight with HindIII restriction enzyme (Invitrogen) at 37°C.

    Techniques: Recombinant, Staining, Isolation, Clone Assay, Hybridization, Sequencing

    Identification of recombination vector pET 32b (+)-UL16 by restriction enzymes digestion . Lane M1, DNA marker; Lane 1, PCR products from pET 32b(+)-UL16; Lane 2, recombination plasmid pET 32b(+)-UL16 was digested with two restriction enzymes HindIII and XhoI; Lane M2, DNA marker.

    Journal: Virology Journal

    Article Title: Expression and characterization of UL16 gene from duck enteritis virus

    doi: 10.1186/1743-422X-8-413

    Figure Lengend Snippet: Identification of recombination vector pET 32b (+)-UL16 by restriction enzymes digestion . Lane M1, DNA marker; Lane 1, PCR products from pET 32b(+)-UL16; Lane 2, recombination plasmid pET 32b(+)-UL16 was digested with two restriction enzymes HindIII and XhoI; Lane M2, DNA marker.

    Article Snippet: The T-clone plasmid, pMD18-T- UL16, was digested with the endonucleases HindIII and XhoI, and the UL16 target sequence was subcloned into the same multicloning sites of pET32b (+) (Invitrogen).

    Techniques: Plasmid Preparation, Positron Emission Tomography, Marker, Polymerase Chain Reaction

    U178G/U179G replicates but fails to exit local leaves following rub-inoculation. Total RNA was collected from: (A) inoculated leaves, (B) upper systemically infected leaves, or (C) petioles of inoculated leaves of 10 plants inoculated with U178G/U179G or wild type PSTVd (WT, one plant, positive control). Mock inoculation (M) was a negative control. (A) RNA blot assay indicates U178G/U179G replication in rub-inoculated leaves. (B) RNA blot assay indicates U178G/U179G is unable to traffic to upper leaves following rub inoculation. (C) RT-PCR indicates U178G/U179G is not present in petioles and fails to exit inoculated leaves. In A and B, the region of the blot corresponding to circular progeny genomes is shown. Loading controls were ribosomal RNA (rRNA) (A and B) and RT-PCR of actin mRNA (C), detected by ethidium bromide staining. Images are representative of 10 (A and B) and three (C) independent experiments.

    Journal: PLoS Pathogens

    Article Title: A three-dimensional RNA motif mediates directional trafficking of Potato spindle tuber viroid from epidermal to palisade mesophyll cells in Nicotiana benthamiana

    doi: 10.1371/journal.ppat.1008147

    Figure Lengend Snippet: U178G/U179G replicates but fails to exit local leaves following rub-inoculation. Total RNA was collected from: (A) inoculated leaves, (B) upper systemically infected leaves, or (C) petioles of inoculated leaves of 10 plants inoculated with U178G/U179G or wild type PSTVd (WT, one plant, positive control). Mock inoculation (M) was a negative control. (A) RNA blot assay indicates U178G/U179G replication in rub-inoculated leaves. (B) RNA blot assay indicates U178G/U179G is unable to traffic to upper leaves following rub inoculation. (C) RT-PCR indicates U178G/U179G is not present in petioles and fails to exit inoculated leaves. In A and B, the region of the blot corresponding to circular progeny genomes is shown. Loading controls were ribosomal RNA (rRNA) (A and B) and RT-PCR of actin mRNA (C), detected by ethidium bromide staining. Images are representative of 10 (A and B) and three (C) independent experiments.

    Article Snippet: Plasmid pRZ6-2-Int and mutant derivatives were linearized with Hind III and employed as templates to generate (+)-PSTVd wild type and mutant in vitro transcripts using the T7 Megascript kit (ThermoFisher Scientific).

    Techniques: Infection, Positive Control, Negative Control, Northern blot, Reverse Transcription Polymerase Chain Reaction, Staining

    Stability of U178G/U179G is similar to wild type PSTVd. (A) RNA blot of in vitro degradation assays performed at 28°C in buffer (20 mM Tris-HCl pH 7.5, 150 mM NaCl, 10 mM phenylmethylsulfonyl fluoride), or uninfected N . benthamiana leaf extract prepared with the same buffer. (B) Percentage of remaining PSTVd wild type (WT) and U178G/U179G RNA over time was determined using Quantity One software. Data are representative of three independent experiments.

    Journal: PLoS Pathogens

    Article Title: A three-dimensional RNA motif mediates directional trafficking of Potato spindle tuber viroid from epidermal to palisade mesophyll cells in Nicotiana benthamiana

    doi: 10.1371/journal.ppat.1008147

    Figure Lengend Snippet: Stability of U178G/U179G is similar to wild type PSTVd. (A) RNA blot of in vitro degradation assays performed at 28°C in buffer (20 mM Tris-HCl pH 7.5, 150 mM NaCl, 10 mM phenylmethylsulfonyl fluoride), or uninfected N . benthamiana leaf extract prepared with the same buffer. (B) Percentage of remaining PSTVd wild type (WT) and U178G/U179G RNA over time was determined using Quantity One software. Data are representative of three independent experiments.

    Article Snippet: Plasmid pRZ6-2-Int and mutant derivatives were linearized with Hind III and employed as templates to generate (+)-PSTVd wild type and mutant in vitro transcripts using the T7 Megascript kit (ThermoFisher Scientific).

    Techniques: Northern blot, In Vitro, Software

    Transient, stable and amplificational replication of the HPV11wt and HPV11E8- genomes in U2OS cells. The mock-transfected cells were used as a negative control (A, lane 7, B, lane 23 and C, lane 10). The linearized HPV11 genome of 100 copies (A, lane 8, B, lane 24 and C, lane 11) and DpnI fragments (C, lane 12) was used as size markers, also indicated with arrows. (A) U2OS cells were transfected with 500 ng of the HPV11wt (lanes 1-3) or E8- (lanes 4-6) genome. Extrachromosomal DNA was extracted via Hirt lysis at 3, 4 and 5 days post-transfection, digested with HindIII and with DpnI. The replication signal was detected via the Southern blot method with a radiolabelled HPV11 genome probe. (B) U2OS cells were transfected with 500 ng of the HPV11wt (lanes 1-11) or HPV11E8- (lanes 12-22) genome, together with 2 μg of the linearized pBabe-Neo construct. The transfected cells were selected with the antibiotic G418, and at 10 days post-transfection, the cells were split and cultivated under either subconfluent (lanes 2-6 and 13-17) or confluent conditions (lanes 7-11 and 18-22). Total DNA was extracted at the time points indicated at top of the figure, and 3 μg of each sample was analyzed as indicated in A . (C) Effect of E8˄E1 and E8˄E2 proteins on viral genome replication. U2OS cells were transfected with 500 ng of the HPV11E8- genome with increasing amounts of either the E8˄E1 or E8˄E2 expression plasmid. Total DNA was extracted at 4 days post-transfection, and 3 μg of each sample was analyzed as indicated in A . (D) The quantitation of HPV11wt and E8- genome DNA replication signals at different time points at subconfluent and confluent culture conditions. The signals were normalized to 10 th day time point. Shown is one of two independent experiments.

    Journal: Virology Journal

    Article Title: The transcription map of HPV11 in U2OS cells adequately reflects the initial and stable replication phases of the viral genome

    doi: 10.1186/s12985-015-0292-6

    Figure Lengend Snippet: Transient, stable and amplificational replication of the HPV11wt and HPV11E8- genomes in U2OS cells. The mock-transfected cells were used as a negative control (A, lane 7, B, lane 23 and C, lane 10). The linearized HPV11 genome of 100 copies (A, lane 8, B, lane 24 and C, lane 11) and DpnI fragments (C, lane 12) was used as size markers, also indicated with arrows. (A) U2OS cells were transfected with 500 ng of the HPV11wt (lanes 1-3) or E8- (lanes 4-6) genome. Extrachromosomal DNA was extracted via Hirt lysis at 3, 4 and 5 days post-transfection, digested with HindIII and with DpnI. The replication signal was detected via the Southern blot method with a radiolabelled HPV11 genome probe. (B) U2OS cells were transfected with 500 ng of the HPV11wt (lanes 1-11) or HPV11E8- (lanes 12-22) genome, together with 2 μg of the linearized pBabe-Neo construct. The transfected cells were selected with the antibiotic G418, and at 10 days post-transfection, the cells were split and cultivated under either subconfluent (lanes 2-6 and 13-17) or confluent conditions (lanes 7-11 and 18-22). Total DNA was extracted at the time points indicated at top of the figure, and 3 μg of each sample was analyzed as indicated in A . (C) Effect of E8˄E1 and E8˄E2 proteins on viral genome replication. U2OS cells were transfected with 500 ng of the HPV11E8- genome with increasing amounts of either the E8˄E1 or E8˄E2 expression plasmid. Total DNA was extracted at 4 days post-transfection, and 3 μg of each sample was analyzed as indicated in A . (D) The quantitation of HPV11wt and E8- genome DNA replication signals at different time points at subconfluent and confluent culture conditions. The signals were normalized to 10 th day time point. Shown is one of two independent experiments.

    Article Snippet: Before Southern blot analysis, half of each extrachromosomal DNA sample or 3 μg of each total DNA sample was treated with the linearizing enzyme HindIII, and non-replicated, bacterially produced dam-methylated input DNA was fragmented with DpnI (Thermo Scientific).

    Techniques: Transfection, Negative Control, Lysis, Southern Blot, Construct, Expressing, Plasmid Preparation, Quantitation Assay

    Electrophoretic product of PvAMA-1 gene treated with EcoR I, PvuII, and HindIII, enzymes using RFLP-PCR technique including Columns 1,2 3 treated with EcoRI. Column 4 main bound of Pv AMA-1. Column 5 size marker with 1000bp. Columns 6, 7 and 8 treated with PvuII. Columns 9 10 treated with HindIII

    Journal: Iranian Journal of Parasitology

    Article Title: Allelic Variations of Plasmodium vivax Apical Membrane Antigen-1 (Pv AMA-1) in Malarious Areas of Southeastern Iran Using PCR-RFLP Technique

    doi:

    Figure Lengend Snippet: Electrophoretic product of PvAMA-1 gene treated with EcoR I, PvuII, and HindIII, enzymes using RFLP-PCR technique including Columns 1,2 3 treated with EcoRI. Column 4 main bound of Pv AMA-1. Column 5 size marker with 1000bp. Columns 6, 7 and 8 treated with PvuII. Columns 9 10 treated with HindIII

    Article Snippet: RFLP In order to determine the presence of different alleles of Pv AMA-1 gene in the region, PCR–RFLP technique was done to digest the gene using three restriction enzymes EcoR-1, Pvu-II and Hind3 (Thermo cat No #ER0271, #ER0631 and ferments cat No #ER0501 respectively) according to the manufacturer’s recommendations.

    Techniques: Polymerase Chain Reaction, Marker

    (A) Agarose gel electrophoresis of the recovered DNA from the Guaymas Basin sediment sample. Equal amount of sample (0.3 g) were extracted with three methods. The loading volume of the recovered DNA was 1 μL. M indicates Lambda DNA/HindIII Marker 2 (Thermo Fisher Scientific, USA). (B) Representation of DNA band intensity (A) as plotted in three dimensional image. (C) Standard curve for measuring the DNA concentration using CLIQS 1D Pro software as determined by the DNA band intensity on the agarose gel.

    Journal: Frontiers in Microbiology

    Article Title: A Modified SDS-Based DNA Extraction Method for High Quality Environmental DNA from Seafloor Environments

    doi: 10.3389/fmicb.2016.00986

    Figure Lengend Snippet: (A) Agarose gel electrophoresis of the recovered DNA from the Guaymas Basin sediment sample. Equal amount of sample (0.3 g) were extracted with three methods. The loading volume of the recovered DNA was 1 μL. M indicates Lambda DNA/HindIII Marker 2 (Thermo Fisher Scientific, USA). (B) Representation of DNA band intensity (A) as plotted in three dimensional image. (C) Standard curve for measuring the DNA concentration using CLIQS 1D Pro software as determined by the DNA band intensity on the agarose gel.

    Article Snippet: And the DNA concentration was also compared by CLIQS 1D Pro software (TotalLab, UK) based on the intensity of the DNA bands on the agarose gel, using Lambda DNA/HindIII Marker 2 (Thermo Fisher Scientific, USA) as the standard curve.

    Techniques: Agarose Gel Electrophoresis, Lambda DNA Preparation, Marker, Concentration Assay, Software

    Complementation in vitro among IN mutants analyzed using a PCR-based method. (A) Assays of integration into the top strand of a phage lambda DNA target. (B) Assays of integration into the bottom strand of a phage lambda DNA target. Full-length (1-288),

    Journal: Journal of Virology

    Article Title: Division of Labor within Human Immunodeficiency Virus Integrase Complexes: Determinants of Catalysis and Target DNA Capture †

    doi: 10.1128/JVI.79.24.15376-15387.2005

    Figure Lengend Snippet: Complementation in vitro among IN mutants analyzed using a PCR-based method. (A) Assays of integration into the top strand of a phage lambda DNA target. (B) Assays of integration into the bottom strand of a phage lambda DNA target. Full-length (1-288),

    Article Snippet: Purified IN variants were diluted to 10 pmol/μl in ISB (see above), and 30 pmol (or 15 pmol of each IN derivative in double mixtures or 15 pmol of 1-212 and 7.5 pmol of each of two 50-288 derivatives in triple mixtures) was incubated with 3 μg of lambda DNA/HindIII (target DNA; Invitrogen, Mass.) in 25 mM KCl, 10 mM β-ME, 30 mM MES (morpholineethanesulfonic acid; pH 6.7), 15 mM MnCl2 , 10% glycerol, and 0.1 mg/ml bovine serum albumin.

    Techniques: In Vitro, Polymerase Chain Reaction, Lambda DNA Preparation

    Complementation assays in vitro containing S119A indicate that the 50-288 partner is responsible for both target sequence recognition and catalysis. (A) Assays of integration into the top strand of a phage lambda DNA target. Full-length (1-288), C-terminally

    Journal: Journal of Virology

    Article Title: Division of Labor within Human Immunodeficiency Virus Integrase Complexes: Determinants of Catalysis and Target DNA Capture †

    doi: 10.1128/JVI.79.24.15376-15387.2005

    Figure Lengend Snippet: Complementation assays in vitro containing S119A indicate that the 50-288 partner is responsible for both target sequence recognition and catalysis. (A) Assays of integration into the top strand of a phage lambda DNA target. Full-length (1-288), C-terminally

    Article Snippet: Purified IN variants were diluted to 10 pmol/μl in ISB (see above), and 30 pmol (or 15 pmol of each IN derivative in double mixtures or 15 pmol of 1-212 and 7.5 pmol of each of two 50-288 derivatives in triple mixtures) was incubated with 3 μg of lambda DNA/HindIII (target DNA; Invitrogen, Mass.) in 25 mM KCl, 10 mM β-ME, 30 mM MES (morpholineethanesulfonic acid; pH 6.7), 15 mM MnCl2 , 10% glycerol, and 0.1 mg/ml bovine serum albumin.

    Techniques: In Vitro, Sequencing, Lambda DNA Preparation

    Complementation assays in vitro containing S119D indicate that the 50-288 partner is responsible for both target sequence recognition and catalysis. (A) Assays of integration into the top strand of a phage lambda DNA target. The S119D target site specificity

    Journal: Journal of Virology

    Article Title: Division of Labor within Human Immunodeficiency Virus Integrase Complexes: Determinants of Catalysis and Target DNA Capture †

    doi: 10.1128/JVI.79.24.15376-15387.2005

    Figure Lengend Snippet: Complementation assays in vitro containing S119D indicate that the 50-288 partner is responsible for both target sequence recognition and catalysis. (A) Assays of integration into the top strand of a phage lambda DNA target. The S119D target site specificity

    Article Snippet: Purified IN variants were diluted to 10 pmol/μl in ISB (see above), and 30 pmol (or 15 pmol of each IN derivative in double mixtures or 15 pmol of 1-212 and 7.5 pmol of each of two 50-288 derivatives in triple mixtures) was incubated with 3 μg of lambda DNA/HindIII (target DNA; Invitrogen, Mass.) in 25 mM KCl, 10 mM β-ME, 30 mM MES (morpholineethanesulfonic acid; pH 6.7), 15 mM MnCl2 , 10% glycerol, and 0.1 mg/ml bovine serum albumin.

    Techniques: In Vitro, Sequencing, Lambda DNA Preparation

    Assessment of YGL250W , SOH1 , and BRE5 (A) Schematic representation of Chromosome II from the diploid W303 background which consists of two LYS2 heteroalleles ( lys2–5′nde I − and lys2–3′nde I − ). These were used to measure meiotic gene conversion (see Materials and Methods ). (B) Spot test of wild type, ygl250wΔ , soh1Δ , and bre5Δ on haploid selection plates and haploid selection plates without lysine to measure meiotic gene conversion. The reduction in meiotic gene conversion of ygl250wΔ , soh1Δ , and bre5Δ was further assessed by random spore analysis ( Table 1 ). (C) Southern blot of DNA isolated from wild type, soh1Δ , and ygl250wΔ SK1 strains containing the ectopic URA3 - ARG4 interval on Chromosome III. The DNA from the indicated times after initiation of sporulation were digested with XhoI then probed to detect COs and DSBs; mw1 represents the λ-HindIII molecular weight marker (Fermentas) and mw2 represents the 1-kb molecular weight marker (Fermentas). The full-sized Southern blots are presented in Figure S1 . For graphs (D–G), wild type, ygl250wΔ , and soh1Δ are represented by black diamonds, black circles, and white squares, respectively. The corresponding XhoI-digested Southern blots are presented in Figure 3 C. (D) Pre-meiotic DNA replication was assessed for synchronized meiotic cultures by fluorescence-activated cell sorting (FACS) and the change from 2c to 4c DNA content was plotted over time. (E) Nuclear divisions (MI and MII) of the synchronized meiotic cultures in (E) were assessed with fluorescence microscopy using 4′,6-diamidino-2-phenylindole (DAPI) staining to visualize nuclear division. (F) Molecular analysis for DSB (DSB1) signal/total lane signal from Southern blots of DNA extracted from synchronized meiotic cultures. (G) Molecular analysis for CO (CO2) signal/total lane signal from Southern blots of DNA extracted from synchronized meiotic cultures.

    Journal: PLoS Genetics

    Article Title: Novel Roles for Selected Genes in Meiotic DNA Processing

    doi: 10.1371/journal.pgen.0030222

    Figure Lengend Snippet: Assessment of YGL250W , SOH1 , and BRE5 (A) Schematic representation of Chromosome II from the diploid W303 background which consists of two LYS2 heteroalleles ( lys2–5′nde I − and lys2–3′nde I − ). These were used to measure meiotic gene conversion (see Materials and Methods ). (B) Spot test of wild type, ygl250wΔ , soh1Δ , and bre5Δ on haploid selection plates and haploid selection plates without lysine to measure meiotic gene conversion. The reduction in meiotic gene conversion of ygl250wΔ , soh1Δ , and bre5Δ was further assessed by random spore analysis ( Table 1 ). (C) Southern blot of DNA isolated from wild type, soh1Δ , and ygl250wΔ SK1 strains containing the ectopic URA3 - ARG4 interval on Chromosome III. The DNA from the indicated times after initiation of sporulation were digested with XhoI then probed to detect COs and DSBs; mw1 represents the λ-HindIII molecular weight marker (Fermentas) and mw2 represents the 1-kb molecular weight marker (Fermentas). The full-sized Southern blots are presented in Figure S1 . For graphs (D–G), wild type, ygl250wΔ , and soh1Δ are represented by black diamonds, black circles, and white squares, respectively. The corresponding XhoI-digested Southern blots are presented in Figure 3 C. (D) Pre-meiotic DNA replication was assessed for synchronized meiotic cultures by fluorescence-activated cell sorting (FACS) and the change from 2c to 4c DNA content was plotted over time. (E) Nuclear divisions (MI and MII) of the synchronized meiotic cultures in (E) were assessed with fluorescence microscopy using 4′,6-diamidino-2-phenylindole (DAPI) staining to visualize nuclear division. (F) Molecular analysis for DSB (DSB1) signal/total lane signal from Southern blots of DNA extracted from synchronized meiotic cultures. (G) Molecular analysis for CO (CO2) signal/total lane signal from Southern blots of DNA extracted from synchronized meiotic cultures.

    Article Snippet: The DNA from the indicated times after initiation of sporulation were digested with XhoI and EcoRI then probed to detect NCOs, COs, and DSBs; mw1 represents the λ-HindIII molecular weight marker (Fermentas) and mw2 represents the 1-kb molecular weight marker (Fermentas).

    Techniques: Spot Test, Selection, Southern Blot, Isolation, Molecular Weight, Marker, Fluorescence, FACS, Microscopy, Staining

    Publicly available caucasian control individual genotyping frequencies versus pooling frequencies for Affymetrix Genechip HindIII and Illumina HumanHap300 arrays. The data are the 15 645 SNPs in common between the Affymetrix Genechip HindIII and Illumina HumanHap300 arrays. The frequency of the sample of 271 publicly available caucasian controls is on the y -axis, with the pooling frequencies from the N =384 pooled case/controls on the x -axis. The broken line is y = x . The solid line is the regression line.

    Journal: Nucleic Acids Research

    Article Title: Highly cost-efficient genome-wide association studies using DNA pools and dense SNP arrays

    doi: 10.1093/nar/gkm1060

    Figure Lengend Snippet: Publicly available caucasian control individual genotyping frequencies versus pooling frequencies for Affymetrix Genechip HindIII and Illumina HumanHap300 arrays. The data are the 15 645 SNPs in common between the Affymetrix Genechip HindIII and Illumina HumanHap300 arrays. The frequency of the sample of 271 publicly available caucasian controls is on the y -axis, with the pooling frequencies from the N =384 pooled case/controls on the x -axis. The broken line is y = x . The solid line is the regression line.

    Article Snippet: We compared and contrasted Affymetrix Genechip HindIII and Illumina HumanHap300 arrays on the same DNA pools and showed that the HumanHap300 arrays are substantially more efficient.

    Techniques:

    Affymetrix Genechip HindIII versus Illumina HumanHap300 array-specific error plots. The plots show the difference in allele frequency estimates for a pair of arrays for each type on the control pool (actual difference in frequency for each pool = 0). Affymetrix results are from a pair of 50K Genechip HindIII arrays and the Illumina results are from a pair of 300K HumanHap300 arrays. These results are for a single pair of arrays; in practice the array-specific error will be reduced through the use of multiple arrays.

    Journal: Nucleic Acids Research

    Article Title: Highly cost-efficient genome-wide association studies using DNA pools and dense SNP arrays

    doi: 10.1093/nar/gkm1060

    Figure Lengend Snippet: Affymetrix Genechip HindIII versus Illumina HumanHap300 array-specific error plots. The plots show the difference in allele frequency estimates for a pair of arrays for each type on the control pool (actual difference in frequency for each pool = 0). Affymetrix results are from a pair of 50K Genechip HindIII arrays and the Illumina results are from a pair of 300K HumanHap300 arrays. These results are for a single pair of arrays; in practice the array-specific error will be reduced through the use of multiple arrays.

    Article Snippet: We compared and contrasted Affymetrix Genechip HindIII and Illumina HumanHap300 arrays on the same DNA pools and showed that the HumanHap300 arrays are substantially more efficient.

    Techniques:

    Power curves for individual genotyping and pooling. Power is for 2000 cases, 2000 controls. ‘30x HumanHap300’ assumes 6 Illumina HumanHap300 arrays per N =400 pool. ‘15x HumanHap300’ assumes 3 Illumina HumanHap300 arrays per N =400 pool. ‘10x HumanHap300’ assumes 2 Illumina HumanHap300 arrays per N =400 pool. ‘15x Genechip HindIII’ assumes 3 Affymetrix Genechip HindIII arrays per N =400 pool. PSD is taken to be 0.009 for Illumina HumanHap arrays, 0.024 for Affymetrix Genechip HindIII arrays. Assumptions for power calculation are a multiplicative disease model, marker allele frequency and disease allele frequency both =0.4, complete linkage disequilibrium between marker and disease alleles, alpha = 0.0000001 (i.e. 500 000 tests), disease prevalence 0.01.

    Journal: Nucleic Acids Research

    Article Title: Highly cost-efficient genome-wide association studies using DNA pools and dense SNP arrays

    doi: 10.1093/nar/gkm1060

    Figure Lengend Snippet: Power curves for individual genotyping and pooling. Power is for 2000 cases, 2000 controls. ‘30x HumanHap300’ assumes 6 Illumina HumanHap300 arrays per N =400 pool. ‘15x HumanHap300’ assumes 3 Illumina HumanHap300 arrays per N =400 pool. ‘10x HumanHap300’ assumes 2 Illumina HumanHap300 arrays per N =400 pool. ‘15x Genechip HindIII’ assumes 3 Affymetrix Genechip HindIII arrays per N =400 pool. PSD is taken to be 0.009 for Illumina HumanHap arrays, 0.024 for Affymetrix Genechip HindIII arrays. Assumptions for power calculation are a multiplicative disease model, marker allele frequency and disease allele frequency both =0.4, complete linkage disequilibrium between marker and disease alleles, alpha = 0.0000001 (i.e. 500 000 tests), disease prevalence 0.01.

    Article Snippet: We compared and contrasted Affymetrix Genechip HindIII and Illumina HumanHap300 arrays on the same DNA pools and showed that the HumanHap300 arrays are substantially more efficient.

    Techniques: Marker

    Some Twinkle mutants cause replication stalling. 2DNAGE samples for all panels consisted of purified mtDNA digested with HincII and probed with a radiolabeled cytochrome b gene fragment (nt 14 846–15 357). The detected fragment includes the non-coding region of mtDNA also including the cytochrome b , ND6 , part of the ND5 gene and intervening tRNA genes (nt 13 636–1006). ( A ) The first two panels on the left show the RIs of HEK293 cells and the interpretation based primarily on earlier 2DNAGE analysis of mtDNA RIs ( 7 , 9 ). Abbreviations: 1n, 3.9 kb non-replicating HincII fragment. (b) bubble arcs. MtDNA bubble arcs are usually very sensitive to RNase H due to the presence of patches of RNA–DNA especially on the lagging strand; these therefore also fall in the category of RITOLS as do various other RIs. Here, y and y′ indicate ascending and descending parts of the y arc and (dy) indicates double-Y structures. These will eventually form resolution intermediates resembling Holliday junctions (HJL-Holliday junction like molecules). The two panels on the right show a comparison of RIs of non-induced and fully induced cells expressing Twinkle wt. The only notable difference in this case is a reproducible reduction in one of the HJL RIs as indicated. ( B ) A normal pattern of RIs similar to non-expressing cells was observed in cells expressing Twinky or the ▵Linker variant. ( C ) K421A, G575D and ▵70–343 show similar patterns of replication stalling, with increased bubble (b) and descending Y-arc (y′) intensities, a sharpening and lengthening of the bubble arc and loss of RITOLS (ovals). The right-most panel shows the same exposure 2D gel pattern of the non-induced ▵70–343 line showing the typical HincII fragment pattern, including abundant RITOLS. ( D ) A limited S1 digestion illustrates that stalled RIs observed in panel C are S1 insensitive (right two panels). The effectiveness of the S1 treatment is illustrated by the left two panels, showing the effect on Twinkle non-induced cells. Similar to the S1 treatment RNase H treatment shows that the stalling RIs observed with Twinkle mutants are largely insensitive to this enzyme (Supplementary Figure 2), showing that the observed stalled RIs in panel C are essentially dsDNA. Although the intensities in subfigures A–D cannot be directly compared due to differences in exposure time, each panel contains appropriate controls of similar exposure. For example, the exposures of the left two panels in C have been chosen to be similar in comparison to the right-most panels to properly illustrate the severity of the stalling phenotype.

    Journal: Nucleic Acids Research

    Article Title: Expression of catalytic mutants of the mtDNA helicase Twinkle and polymerase POLG causes distinct replication stalling phenotypes

    doi: 10.1093/nar/gkm215

    Figure Lengend Snippet: Some Twinkle mutants cause replication stalling. 2DNAGE samples for all panels consisted of purified mtDNA digested with HincII and probed with a radiolabeled cytochrome b gene fragment (nt 14 846–15 357). The detected fragment includes the non-coding region of mtDNA also including the cytochrome b , ND6 , part of the ND5 gene and intervening tRNA genes (nt 13 636–1006). ( A ) The first two panels on the left show the RIs of HEK293 cells and the interpretation based primarily on earlier 2DNAGE analysis of mtDNA RIs ( 7 , 9 ). Abbreviations: 1n, 3.9 kb non-replicating HincII fragment. (b) bubble arcs. MtDNA bubble arcs are usually very sensitive to RNase H due to the presence of patches of RNA–DNA especially on the lagging strand; these therefore also fall in the category of RITOLS as do various other RIs. Here, y and y′ indicate ascending and descending parts of the y arc and (dy) indicates double-Y structures. These will eventually form resolution intermediates resembling Holliday junctions (HJL-Holliday junction like molecules). The two panels on the right show a comparison of RIs of non-induced and fully induced cells expressing Twinkle wt. The only notable difference in this case is a reproducible reduction in one of the HJL RIs as indicated. ( B ) A normal pattern of RIs similar to non-expressing cells was observed in cells expressing Twinky or the ▵Linker variant. ( C ) K421A, G575D and ▵70–343 show similar patterns of replication stalling, with increased bubble (b) and descending Y-arc (y′) intensities, a sharpening and lengthening of the bubble arc and loss of RITOLS (ovals). The right-most panel shows the same exposure 2D gel pattern of the non-induced ▵70–343 line showing the typical HincII fragment pattern, including abundant RITOLS. ( D ) A limited S1 digestion illustrates that stalled RIs observed in panel C are S1 insensitive (right two panels). The effectiveness of the S1 treatment is illustrated by the left two panels, showing the effect on Twinkle non-induced cells. Similar to the S1 treatment RNase H treatment shows that the stalling RIs observed with Twinkle mutants are largely insensitive to this enzyme (Supplementary Figure 2), showing that the observed stalled RIs in panel C are essentially dsDNA. Although the intensities in subfigures A–D cannot be directly compared due to differences in exposure time, each panel contains appropriate controls of similar exposure. For example, the exposures of the left two panels in C have been chosen to be similar in comparison to the right-most panels to properly illustrate the severity of the stalling phenotype.

    Article Snippet: Purified mtDNA was digested with HincII and where mentioned further treated with RNase H or S1 nuclease (Fermentas, Hanover, MD, USA) with the indicated amounts and time.

    Techniques: Purification, Expressing, Variant Assay, Two-Dimensional Gel Electrophoresis