sequencing dna Search Results


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    AceSeq DNA Sequencing Kit is a rapid,high performance DNA library prep solution for next generation sequencing on Illumina platforms AceSeq DNA Seq Kit prepares single paired end and multiplexed genomic
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    Thermo Fisher 3730xl dna analyzer
    3730xl Dna Analyzer, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 52281 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Biotechnology Information dna sequences
    (a) Haplotype network of the combined three cp <t>DNA</t> ( <t>psb</t> A‐t rn H, rpl 32‐ trn L, and trn L‐ trn F); (b) Haplotype network of the combined four DNA ( ITS 2, psb A‐ trn H, mat K, and rbc L) regions using median joining method. Blue dots stand for Tripterygium wilfordii , red dots stand for Tripterygium hypoglaucum , green dots stands for Tripterygium regelii , and black dots stand for Celastrus orbiculatus
    Dna Sequences, supplied by Biotechnology Information, used in various techniques. Bioz Stars score: 92/100, based on 2081 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher abi prism 377 dna sequencer
    (a) Haplotype network of the combined three cp <t>DNA</t> ( <t>psb</t> A‐t rn H, rpl 32‐ trn L, and trn L‐ trn F); (b) Haplotype network of the combined four DNA ( ITS 2, psb A‐ trn H, mat K, and rbc L) regions using median joining method. Blue dots stand for Tripterygium wilfordii , red dots stand for Tripterygium hypoglaucum , green dots stands for Tripterygium regelii , and black dots stand for Celastrus orbiculatus
    Abi Prism 377 Dna Sequencer, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 92/100, based on 3170 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Bioedit Company dna sequences
    Titration of the <t>DNA</t> donor and integrase concentrations for the strand transfer assay. (A) Purification of <t>HIV-O</t> recombinant integrase proteins from clades A (IN-O/A) and B (IN-O/B) and a divergent strain (IN-O/Div) and HIV-M recombinant integrase protein (IN-M/B). SDS-PAGE analysis showing that the integrase proteins resolved predominantly in a high-density band (35.6 kDa); the two bands at the bottom represent nonspecifically cleaved forms of the protein. (B) Calibration of the DNA donor mimicking LTR-O using various concentrations, ranging from 0 to 2,400 nM; (C) calibration of the different HIV-O integrases, IN-O/A (clade A), IN-O/B (clade B), and IN-O/Div (divergent strain), and the HIV-M integrase, IN-M/B (HIV-M subtype B), using various concentrations, ranging from 0 to 1,600 nM.
    Dna Sequences, supplied by Bioedit Company, used in various techniques. Bioz Stars score: 92/100, based on 1763 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher abi 3730 dna sequencer
    Titration of the <t>DNA</t> donor and integrase concentrations for the strand transfer assay. (A) Purification of <t>HIV-O</t> recombinant integrase proteins from clades A (IN-O/A) and B (IN-O/B) and a divergent strain (IN-O/Div) and HIV-M recombinant integrase protein (IN-M/B). SDS-PAGE analysis showing that the integrase proteins resolved predominantly in a high-density band (35.6 kDa); the two bands at the bottom represent nonspecifically cleaved forms of the protein. (B) Calibration of the DNA donor mimicking LTR-O using various concentrations, ranging from 0 to 2,400 nM; (C) calibration of the different HIV-O integrases, IN-O/A (clade A), IN-O/B (clade B), and IN-O/Div (divergent strain), and the HIV-M integrase, IN-M/B (HIV-M subtype B), using various concentrations, ranging from 0 to 1,600 nM.
    Abi 3730 Dna Sequencer, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 2282 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore dna sequence
    Titration of the <t>DNA</t> donor and integrase concentrations for the strand transfer assay. (A) Purification of <t>HIV-O</t> recombinant integrase proteins from clades A (IN-O/A) and B (IN-O/B) and a divergent strain (IN-O/Div) and HIV-M recombinant integrase protein (IN-M/B). SDS-PAGE analysis showing that the integrase proteins resolved predominantly in a high-density band (35.6 kDa); the two bands at the bottom represent nonspecifically cleaved forms of the protein. (B) Calibration of the DNA donor mimicking LTR-O using various concentrations, ranging from 0 to 2,400 nM; (C) calibration of the different HIV-O integrases, IN-O/A (clade A), IN-O/B (clade B), and IN-O/Div (divergent strain), and the HIV-M integrase, IN-M/B (HIV-M subtype B), using various concentrations, ranging from 0 to 1,600 nM.
    Dna Sequence, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 1652 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Genewiz dna sequence analysis
    The <t>bbb22</t> gene alone is not sufficient to maintain wild-type levels of spirochete loads in infected mouse tissues. <t>DNA</t> was isolated from ear, heart, and joint tissues of C3H/HeN mice inoculated with 1 × 10 4 bbb22-23 + or 22dist p -bbb22 + spirochetes.
    Dna Sequence Analysis, supplied by Genewiz, used in various techniques. Bioz Stars score: 92/100, based on 311 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Eurofins dna sequencing
    The <t>bbb22</t> gene alone is not sufficient to maintain wild-type levels of spirochete loads in infected mouse tissues. <t>DNA</t> was isolated from ear, heart, and joint tissues of C3H/HeN mice inoculated with 1 × 10 4 bbb22-23 + or 22dist p -bbb22 + spirochetes.
    Dna Sequencing, supplied by Eurofins, used in various techniques. Bioz Stars score: 92/100, based on 4234 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher automatic dna sequencer
    The <t>bbb22</t> gene alone is not sufficient to maintain wild-type levels of spirochete loads in infected mouse tissues. <t>DNA</t> was isolated from ear, heart, and joint tissues of C3H/HeN mice inoculated with 1 × 10 4 bbb22-23 + or 22dist p -bbb22 + spirochetes.
    Automatic Dna Sequencer, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 1235 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    GenScript dna sequences
    Aptamer Binding Selectively Reduces Mutant <t>Huntingtin’s</t> PRC2-Stimulating Activity (A) The affinities of Q78-huntingtin alone and huntingtin-aptamer complexes to PRC2 were compared by immunoprecipitation with an anti-EZH2 antibody and an anti-huntingtin antibody (HP-1), followed by immunoblotting for huntingtin and EZH2. The experiment was repeated three times. (B) Autoradiogram of bands of 3 H-methyl histone H3 produced by PRC2 in the absence and presence of mutant-huntingtin-preferring <t>DNA</t> aptamers. Bottom: the bar graph of the band densitometry results, showing that aptamer binding significantly reduced the ability of mutant huntingtin to enhance basal PRC2 activity. Mean band intensities were measured from three independent experiments. Asterisks indicate statistically significant differences compared with unbound Q78-huntingtin. Error bars represent SEM. *p
    Dna Sequences, supplied by GenScript, used in various techniques. Bioz Stars score: 92/100, based on 590 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher abi prism 3100 dna sequencer
    Aptamer Binding Selectively Reduces Mutant <t>Huntingtin’s</t> PRC2-Stimulating Activity (A) The affinities of Q78-huntingtin alone and huntingtin-aptamer complexes to PRC2 were compared by immunoprecipitation with an anti-EZH2 antibody and an anti-huntingtin antibody (HP-1), followed by immunoblotting for huntingtin and EZH2. The experiment was repeated three times. (B) Autoradiogram of bands of 3 H-methyl histone H3 produced by PRC2 in the absence and presence of mutant-huntingtin-preferring <t>DNA</t> aptamers. Bottom: the bar graph of the band densitometry results, showing that aptamer binding significantly reduced the ability of mutant huntingtin to enhance basal PRC2 activity. Mean band intensities were measured from three independent experiments. Asterisks indicate statistically significant differences compared with unbound Q78-huntingtin. Error bars represent SEM. *p
    Abi Prism 3100 Dna Sequencer, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 1382 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher abi 377 dna sequencer
    Aptamer Binding Selectively Reduces Mutant <t>Huntingtin’s</t> PRC2-Stimulating Activity (A) The affinities of Q78-huntingtin alone and huntingtin-aptamer complexes to PRC2 were compared by immunoprecipitation with an anti-EZH2 antibody and an anti-huntingtin antibody (HP-1), followed by immunoblotting for huntingtin and EZH2. The experiment was repeated three times. (B) Autoradiogram of bands of 3 H-methyl histone H3 produced by PRC2 in the absence and presence of mutant-huntingtin-preferring <t>DNA</t> aptamers. Bottom: the bar graph of the band densitometry results, showing that aptamer binding significantly reduced the ability of mutant huntingtin to enhance basal PRC2 activity. Mean band intensities were measured from three independent experiments. Asterisks indicate statistically significant differences compared with unbound Q78-huntingtin. Error bars represent SEM. *p
    Abi 377 Dna Sequencer, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 1398 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher 377 dna sequencer
    Aptamer Binding Selectively Reduces Mutant <t>Huntingtin’s</t> PRC2-Stimulating Activity (A) The affinities of Q78-huntingtin alone and huntingtin-aptamer complexes to PRC2 were compared by immunoprecipitation with an anti-EZH2 antibody and an anti-huntingtin antibody (HP-1), followed by immunoblotting for huntingtin and EZH2. The experiment was repeated three times. (B) Autoradiogram of bands of 3 H-methyl histone H3 produced by PRC2 in the absence and presence of mutant-huntingtin-preferring <t>DNA</t> aptamers. Bottom: the bar graph of the band densitometry results, showing that aptamer binding significantly reduced the ability of mutant huntingtin to enhance basal PRC2 activity. Mean band intensities were measured from three independent experiments. Asterisks indicate statistically significant differences compared with unbound Q78-huntingtin. Error bars represent SEM. *p
    377 Dna Sequencer, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 91/100, based on 749 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Macrogen dna sequence analysis
    Site directed mutagenesis of the KRRIH region of Msm <t>UdgX</t> and its activity assay. ( A ( i ) and B ( i )) Line diagrams of Msm UdgX and its mutants highlighting the regions targeted by site directed mutagenesis. Motif A, motif B and the KRRIH loop region are shown in red, blue and yellow boxes, respectively. ( A ( ii )) Uracil excision/binding assays with Msm UdgX (WT) and its R107S, H109S and SSAS mutant derivatives. ∼200 ng of each protein was reacted with 5′ 32 P-labeled SSU9, the reaction mix was resolved on 8 M urea-PAGE (15%) and analyzed by phosphor imaging. The presence of uracil excision activity is seen by the fast migrating product band, ‘P’ whereas the <t>uracil-DNA</t> binding activity by the slow migrating complex, ‘C’. ( B ( ii )) Uracil excision/binding assays with Msm UdgX (WT) and its F4, F1, LD and F4 LD mutant derivatives (∼200 ng each) using SSU9. ( C ) Uracil release assay of Msm UdgX H109S (∼200 ng) using 3 H-uracil containing DNA (21 000 c.p.m) as substrate. Released uracil was separated by TLC (CEL 300 PEI/UV254) and seen as a fast migrating species upon phosphor imaging (the radioactive spot comigrated with the cold uracil spot detected by UV shadowing). ( D ) Uracil excision/binding assay with Msm UdgX and its H109S, H109G, H109Q, H109A and C27S derivatives. ∼200 ng of each protein was used in a standard UDG assay with SSU9 substrate.
    Dna Sequence Analysis, supplied by Macrogen, used in various techniques. Bioz Stars score: 92/100, based on 193 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher dna sequencing kit
    Site directed mutagenesis of the KRRIH region of Msm <t>UdgX</t> and its activity assay. ( A ( i ) and B ( i )) Line diagrams of Msm UdgX and its mutants highlighting the regions targeted by site directed mutagenesis. Motif A, motif B and the KRRIH loop region are shown in red, blue and yellow boxes, respectively. ( A ( ii )) Uracil excision/binding assays with Msm UdgX (WT) and its R107S, H109S and SSAS mutant derivatives. ∼200 ng of each protein was reacted with 5′ 32 P-labeled SSU9, the reaction mix was resolved on 8 M urea-PAGE (15%) and analyzed by phosphor imaging. The presence of uracil excision activity is seen by the fast migrating product band, ‘P’ whereas the <t>uracil-DNA</t> binding activity by the slow migrating complex, ‘C’. ( B ( ii )) Uracil excision/binding assays with Msm UdgX (WT) and its F4, F1, LD and F4 LD mutant derivatives (∼200 ng each) using SSU9. ( C ) Uracil release assay of Msm UdgX H109S (∼200 ng) using 3 H-uracil containing DNA (21 000 c.p.m) as substrate. Released uracil was separated by TLC (CEL 300 PEI/UV254) and seen as a fast migrating species upon phosphor imaging (the radioactive spot comigrated with the cold uracil spot detected by UV shadowing). ( D ) Uracil excision/binding assay with Msm UdgX and its H109S, H109G, H109Q, H109A and C27S derivatives. ∼200 ng of each protein was used in a standard UDG assay with SSU9 substrate.
    Dna Sequencing Kit, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 1083 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher abi prism 3730 dna sequencer
    Site directed mutagenesis of the KRRIH region of Msm <t>UdgX</t> and its activity assay. ( A ( i ) and B ( i )) Line diagrams of Msm UdgX and its mutants highlighting the regions targeted by site directed mutagenesis. Motif A, motif B and the KRRIH loop region are shown in red, blue and yellow boxes, respectively. ( A ( ii )) Uracil excision/binding assays with Msm UdgX (WT) and its R107S, H109S and SSAS mutant derivatives. ∼200 ng of each protein was reacted with 5′ 32 P-labeled SSU9, the reaction mix was resolved on 8 M urea-PAGE (15%) and analyzed by phosphor imaging. The presence of uracil excision activity is seen by the fast migrating product band, ‘P’ whereas the <t>uracil-DNA</t> binding activity by the slow migrating complex, ‘C’. ( B ( ii )) Uracil excision/binding assays with Msm UdgX (WT) and its F4, F1, LD and F4 LD mutant derivatives (∼200 ng each) using SSU9. ( C ) Uracil release assay of Msm UdgX H109S (∼200 ng) using 3 H-uracil containing DNA (21 000 c.p.m) as substrate. Released uracil was separated by TLC (CEL 300 PEI/UV254) and seen as a fast migrating species upon phosphor imaging (the radioactive spot comigrated with the cold uracil spot detected by UV shadowing). ( D ) Uracil excision/binding assay with Msm UdgX and its H109S, H109G, H109Q, H109A and C27S derivatives. ∼200 ng of each protein was used in a standard UDG assay with SSU9 substrate.
    Abi Prism 3730 Dna Sequencer, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 91/100, based on 793 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Eurofins dna sequences
    Telomere <t>DNA</t> G-quadruplex unfolding by arginine to alanine mutants of <t>UP1+RGG</t> monitored using CD spectroscopy. ( A, B ) Unfolding of K + form of Tel22 G-quadruplex DNA by TriRGG (A) and AllRGG (B) mutants. The G-quadruplex DNA was titrated with increasing molar excess of proteins. The black arrows in the spectra indicate the gradual decrease in ellipticity at 295 nm with increasing protein concentration. ( C ) The normalized ellipticity at 295 nm of Tel22 at the final titration step (at 1:6 molar ratio of DNA to protein) for UP1, AllRGG, TriRGG and UP1+RGG showing the relative foldedness of the G-quadruplex structure.
    Dna Sequences, supplied by Eurofins, used in various techniques. Bioz Stars score: 92/100, based on 527 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher abi 3730 automated dna sequencer
    Telomere <t>DNA</t> G-quadruplex unfolding by arginine to alanine mutants of <t>UP1+RGG</t> monitored using CD spectroscopy. ( A, B ) Unfolding of K + form of Tel22 G-quadruplex DNA by TriRGG (A) and AllRGG (B) mutants. The G-quadruplex DNA was titrated with increasing molar excess of proteins. The black arrows in the spectra indicate the gradual decrease in ellipticity at 295 nm with increasing protein concentration. ( C ) The normalized ellipticity at 295 nm of Tel22 at the final titration step (at 1:6 molar ratio of DNA to protein) for UP1, AllRGG, TriRGG and UP1+RGG showing the relative foldedness of the G-quadruplex structure.
    Abi 3730 Automated Dna Sequencer, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 95/100, based on 594 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    The Kit is designed for rapid and efficient purification of circulating cell free DNA cfDNA The system utilizes silicacoated paramagnetic particles to purify cfDNA from less than 0 3 mL
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    Sigma offers glass plates Mylar combs and spacers designed specifically to meet the rigid specifications of the ABI TM 373 and 377 DNA sequencing units Notched glass places are beveled
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    Sequencing module PCR cleanup and DNA sequencing for up to 9 fish samples prepaid service that can be claimed with an authorized sequencing vendor sample shipping included 2 modules required
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    Image Search Results


    (a) Haplotype network of the combined three cp DNA ( psb A‐t rn H, rpl 32‐ trn L, and trn L‐ trn F); (b) Haplotype network of the combined four DNA ( ITS 2, psb A‐ trn H, mat K, and rbc L) regions using median joining method. Blue dots stand for Tripterygium wilfordii , red dots stand for Tripterygium hypoglaucum , green dots stands for Tripterygium regelii , and black dots stand for Celastrus orbiculatus

    Journal: Ecology and Evolution

    Article Title: Phylogeographic and phylogenetic analysis for Tripterygium species delimitation, et al. Phylogeographic and phylogenetic analysis for Tripterygium species delimitation

    doi: 10.1002/ece3.3344

    Figure Lengend Snippet: (a) Haplotype network of the combined three cp DNA ( psb A‐t rn H, rpl 32‐ trn L, and trn L‐ trn F); (b) Haplotype network of the combined four DNA ( ITS 2, psb A‐ trn H, mat K, and rbc L) regions using median joining method. Blue dots stand for Tripterygium wilfordii , red dots stand for Tripterygium hypoglaucum , green dots stands for Tripterygium regelii , and black dots stand for Celastrus orbiculatus

    Article Snippet: The other four DNA sequences (i.e., ITS2, psb A‐trn H, mat K, and rbc L) were downloaded from the National Center for Biotechnology Information (NCBI) nucleotide database ( https://www.ncbi.nlm.nih.gov/nuccore ) and combined as the second matrix, which was used in the phylogenetic analysis, including Bayesian inference (BI), maximum likelihood (ML), and haplotype network construction.

    Techniques:

    BEAST ‐derived chronogram of haplotypes based on the combined cp DNA ( psb A‐t rn H, rpl 32‐ trn L, and trn L‐ trn F) sequences. Blue bars on each node show 95% highest posterior density ( HPD ) confidence intervals for divergence estimates. The number on each branch indicates the posterior probability ( PP ). The node age (Myr) of the major lineages was shown near the blue bars

    Journal: Ecology and Evolution

    Article Title: Phylogeographic and phylogenetic analysis for Tripterygium species delimitation, et al. Phylogeographic and phylogenetic analysis for Tripterygium species delimitation

    doi: 10.1002/ece3.3344

    Figure Lengend Snippet: BEAST ‐derived chronogram of haplotypes based on the combined cp DNA ( psb A‐t rn H, rpl 32‐ trn L, and trn L‐ trn F) sequences. Blue bars on each node show 95% highest posterior density ( HPD ) confidence intervals for divergence estimates. The number on each branch indicates the posterior probability ( PP ). The node age (Myr) of the major lineages was shown near the blue bars

    Article Snippet: The other four DNA sequences (i.e., ITS2, psb A‐trn H, mat K, and rbc L) were downloaded from the National Center for Biotechnology Information (NCBI) nucleotide database ( https://www.ncbi.nlm.nih.gov/nuccore ) and combined as the second matrix, which was used in the phylogenetic analysis, including Bayesian inference (BI), maximum likelihood (ML), and haplotype network construction.

    Techniques: Derivative Assay

    Phylogenetic trees of haplotypes based on the combined three plastid DNA regions (a) ( psb A‐t rn H, rpl 32‐ trn L, and trn L‐ trn F) and the combined four DNA regions (b) ( ITS 2, psb A‐ trn H, mat K, and rbc L) using Bayesian inference method. The number on each branch indicates the posterior probability ( PP ). Model selection: (a) GTR +I+G; (b) GTR +I. The average standard deviation of the split frequencies, (a) 0.006417; (b) 0.002667

    Journal: Ecology and Evolution

    Article Title: Phylogeographic and phylogenetic analysis for Tripterygium species delimitation, et al. Phylogeographic and phylogenetic analysis for Tripterygium species delimitation

    doi: 10.1002/ece3.3344

    Figure Lengend Snippet: Phylogenetic trees of haplotypes based on the combined three plastid DNA regions (a) ( psb A‐t rn H, rpl 32‐ trn L, and trn L‐ trn F) and the combined four DNA regions (b) ( ITS 2, psb A‐ trn H, mat K, and rbc L) using Bayesian inference method. The number on each branch indicates the posterior probability ( PP ). Model selection: (a) GTR +I+G; (b) GTR +I. The average standard deviation of the split frequencies, (a) 0.006417; (b) 0.002667

    Article Snippet: The other four DNA sequences (i.e., ITS2, psb A‐trn H, mat K, and rbc L) were downloaded from the National Center for Biotechnology Information (NCBI) nucleotide database ( https://www.ncbi.nlm.nih.gov/nuccore ) and combined as the second matrix, which was used in the phylogenetic analysis, including Bayesian inference (BI), maximum likelihood (ML), and haplotype network construction.

    Techniques: Selection, Standard Deviation

    Titration of the DNA donor and integrase concentrations for the strand transfer assay. (A) Purification of HIV-O recombinant integrase proteins from clades A (IN-O/A) and B (IN-O/B) and a divergent strain (IN-O/Div) and HIV-M recombinant integrase protein (IN-M/B). SDS-PAGE analysis showing that the integrase proteins resolved predominantly in a high-density band (35.6 kDa); the two bands at the bottom represent nonspecifically cleaved forms of the protein. (B) Calibration of the DNA donor mimicking LTR-O using various concentrations, ranging from 0 to 2,400 nM; (C) calibration of the different HIV-O integrases, IN-O/A (clade A), IN-O/B (clade B), and IN-O/Div (divergent strain), and the HIV-M integrase, IN-M/B (HIV-M subtype B), using various concentrations, ranging from 0 to 1,600 nM.

    Journal: Antimicrobial Agents and Chemotherapy

    Article Title: HIV-1 Group O Integrase Displays Lower Enzymatic Efficiency and Higher Susceptibility to Raltegravir than HIV-1 Group M Subtype B Integrase

    doi: 10.1128/AAC.03819-14

    Figure Lengend Snippet: Titration of the DNA donor and integrase concentrations for the strand transfer assay. (A) Purification of HIV-O recombinant integrase proteins from clades A (IN-O/A) and B (IN-O/B) and a divergent strain (IN-O/Div) and HIV-M recombinant integrase protein (IN-M/B). SDS-PAGE analysis showing that the integrase proteins resolved predominantly in a high-density band (35.6 kDa); the two bands at the bottom represent nonspecifically cleaved forms of the protein. (B) Calibration of the DNA donor mimicking LTR-O using various concentrations, ranging from 0 to 2,400 nM; (C) calibration of the different HIV-O integrases, IN-O/A (clade A), IN-O/B (clade B), and IN-O/Div (divergent strain), and the HIV-M integrase, IN-M/B (HIV-M subtype B), using various concentrations, ranging from 0 to 1,600 nM.

    Article Snippet: DNA sequences were compared with the HIV-1 group M reference strain HxB2 ( ) using BioEdit software for alignment (BioEdit sequence alignment editor v7.1.3, copyright 1997-2011, Tom Hall).

    Techniques: Titration, Purification, Recombinant, SDS Page

    Enzymatic activity of the different HIV-O integrases from clade A (IN-O/A), clade B (IN-O/B), and divergent strain (IN-O/Div) in comparison with HIV-M integrase (IN-M/B). (A) Strand transfer activity: these data were generated using various concentrations of DNA target (0 to 120 nM); (B) 3′ processing activity: the data were generated using various concentrations of LTR-O (0- to 40 nM).

    Journal: Antimicrobial Agents and Chemotherapy

    Article Title: HIV-1 Group O Integrase Displays Lower Enzymatic Efficiency and Higher Susceptibility to Raltegravir than HIV-1 Group M Subtype B Integrase

    doi: 10.1128/AAC.03819-14

    Figure Lengend Snippet: Enzymatic activity of the different HIV-O integrases from clade A (IN-O/A), clade B (IN-O/B), and divergent strain (IN-O/Div) in comparison with HIV-M integrase (IN-M/B). (A) Strand transfer activity: these data were generated using various concentrations of DNA target (0 to 120 nM); (B) 3′ processing activity: the data were generated using various concentrations of LTR-O (0- to 40 nM).

    Article Snippet: DNA sequences were compared with the HIV-1 group M reference strain HxB2 ( ) using BioEdit software for alignment (BioEdit sequence alignment editor v7.1.3, copyright 1997-2011, Tom Hall).

    Techniques: Activity Assay, Generated

    The bbb22 gene alone is not sufficient to maintain wild-type levels of spirochete loads in infected mouse tissues. DNA was isolated from ear, heart, and joint tissues of C3H/HeN mice inoculated with 1 × 10 4 bbb22-23 + or 22dist p -bbb22 + spirochetes.

    Journal: Infection and Immunity

    Article Title: Molecular Dissection of a Borrelia burgdorferiIn Vivo Essential Purine Transport System

    doi: 10.1128/IAI.02859-14

    Figure Lengend Snippet: The bbb22 gene alone is not sufficient to maintain wild-type levels of spirochete loads in infected mouse tissues. DNA was isolated from ear, heart, and joint tissues of C3H/HeN mice inoculated with 1 × 10 4 bbb22-23 + or 22dist p -bbb22 + spirochetes.

    Article Snippet: Plasmids pBSV2G 22dist p - bbb22 + and pBSV2G 22prox p - bbb22 + were confirmed by restriction digestion and DNA sequence analysis (Genewiz).

    Techniques: Infection, Isolation, Mouse Assay

    Aptamer Binding Selectively Reduces Mutant Huntingtin’s PRC2-Stimulating Activity (A) The affinities of Q78-huntingtin alone and huntingtin-aptamer complexes to PRC2 were compared by immunoprecipitation with an anti-EZH2 antibody and an anti-huntingtin antibody (HP-1), followed by immunoblotting for huntingtin and EZH2. The experiment was repeated three times. (B) Autoradiogram of bands of 3 H-methyl histone H3 produced by PRC2 in the absence and presence of mutant-huntingtin-preferring DNA aptamers. Bottom: the bar graph of the band densitometry results, showing that aptamer binding significantly reduced the ability of mutant huntingtin to enhance basal PRC2 activity. Mean band intensities were measured from three independent experiments. Asterisks indicate statistically significant differences compared with unbound Q78-huntingtin. Error bars represent SEM. *p

    Journal: Molecular Therapy. Nucleic Acids

    Article Title: Novel DNA Aptamers that Bind to Mutant Huntingtin and Modify Its Activity

    doi: 10.1016/j.omtn.2018.03.008

    Figure Lengend Snippet: Aptamer Binding Selectively Reduces Mutant Huntingtin’s PRC2-Stimulating Activity (A) The affinities of Q78-huntingtin alone and huntingtin-aptamer complexes to PRC2 were compared by immunoprecipitation with an anti-EZH2 antibody and an anti-huntingtin antibody (HP-1), followed by immunoblotting for huntingtin and EZH2. The experiment was repeated three times. (B) Autoradiogram of bands of 3 H-methyl histone H3 produced by PRC2 in the absence and presence of mutant-huntingtin-preferring DNA aptamers. Bottom: the bar graph of the band densitometry results, showing that aptamer binding significantly reduced the ability of mutant huntingtin to enhance basal PRC2 activity. Mean band intensities were measured from three independent experiments. Asterisks indicate statistically significant differences compared with unbound Q78-huntingtin. Error bars represent SEM. *p

    Article Snippet: Three lysine aspartic acid (KD) mutant huntingtin constructs (K2449D, K2932D/K2934D, K2449D/K2932D/K2934D) were generated by submission of the DNA sequences to Genscript (Piscataway, NJ, USA), which provided synthesized DNA in the pFastBac1 vector using SalI/SacII restriction digest and standard molecular biology techniques.

    Techniques: Binding Assay, Mutagenesis, Activity Assay, Immunoprecipitation, Produced

    Screening of DNA Aptamers that Can Preferentially Bind to Q78-Huntingtin (A) A schematic illustrating DNA aptamer screening to identify aptamers that bind preferentially to Q23- or Q78-purified human recombinant huntingtin. (B) Bar graphs presenting ELISA data show four representative aptamers that bind significantly more to the purified Q78-huntingtin (Q78) compared with Q23-huntingtin (Q23). Mean optical densities at 405 nm were measured by ELISA plate reader from three independent experiments. *p

    Journal: Molecular Therapy. Nucleic Acids

    Article Title: Novel DNA Aptamers that Bind to Mutant Huntingtin and Modify Its Activity

    doi: 10.1016/j.omtn.2018.03.008

    Figure Lengend Snippet: Screening of DNA Aptamers that Can Preferentially Bind to Q78-Huntingtin (A) A schematic illustrating DNA aptamer screening to identify aptamers that bind preferentially to Q23- or Q78-purified human recombinant huntingtin. (B) Bar graphs presenting ELISA data show four representative aptamers that bind significantly more to the purified Q78-huntingtin (Q78) compared with Q23-huntingtin (Q23). Mean optical densities at 405 nm were measured by ELISA plate reader from three independent experiments. *p

    Article Snippet: Three lysine aspartic acid (KD) mutant huntingtin constructs (K2449D, K2932D/K2934D, K2449D/K2932D/K2934D) were generated by submission of the DNA sequences to Genscript (Piscataway, NJ, USA), which provided synthesized DNA in the pFastBac1 vector using SalI/SacII restriction digest and standard molecular biology techniques.

    Techniques: Purification, Recombinant, Enzyme-linked Immunosorbent Assay

    CTD-II Domain near K2932/K2934 in Mutant Huntingtin Is a Potential Binding Site of the MS3 Aptamer (A) A schematic view of huntingtin with five domains delineated as previously described. 8 Caspase-6 cleavage site is indicated by the red arrow. The polyglutamine tract is denoted by the green bar. (B) The biotinylated DNA aptamer MS1 (Biotin, first blot) was detected in uncleaved Q78-huntingtin, and the larger fragment from caspase-6 cleavage of Q78-huntingtin with an anti-biotin antibody. The representative blot showed that most of the aptamer appeared to be bound to the C-terminal fragment and un-cleaved huntingtin (arrows), but not the amino-terminal fragment (arrowhead), which was confirmed by re-probing the blot with a huntingtin C-terminal antibody (HF1, second blot) and an amino-terminal antibody (mAb2166, third blot). The experiment was repeated three times. (C) The same experiment was performed for the remaining biotinylated aptamers (MS2, MS3, MS4) and MS1. The representative immunoblot demonstrated that all aptamers preferentially bound to the C terminus of Q78-huntingtin. The experiment was repeated three times. (D and E) SLM analysis was performed with the huntingtin-aptamer complex and huntingtin alone. The methylated lysine residues were detected by LC/MS/MS and assigned a score based on the equation described in the Materials and Methods . The bar graphs showing the score of Q23- (D) and Q78- (E) huntingtin revealed that K2932/K2934 in CTD-II was the most probable binding site of MS3 on Q78-huntingtin. (F) Representative immunoblot showing purified Q23-, Q78-, Q78 K2449D-, Q78 K2932D/K2934D-, and Q78 K2449D/K2932D/K2934D-huntingtin bound with biotinylated MS3 or GCdx aptamers by probing with an anti-biotin antibody (Biotin) and an anti-huntingtin antibody (mAb2166). The location of full-length huntingtin is indicated by the arrow in both immunoblots. The asterisk marked a blurry band detected by anti-biotin antibody, but not by mAb2166, implying that a protein (or more than one) from insect cells, showing affinities to MS3 aptamer, was particularly enriched in Q78 K2449D-purified protein and less in Q78 K2932D/K2934D-purified proteins. The experiment was repeated three times.

    Journal: Molecular Therapy. Nucleic Acids

    Article Title: Novel DNA Aptamers that Bind to Mutant Huntingtin and Modify Its Activity

    doi: 10.1016/j.omtn.2018.03.008

    Figure Lengend Snippet: CTD-II Domain near K2932/K2934 in Mutant Huntingtin Is a Potential Binding Site of the MS3 Aptamer (A) A schematic view of huntingtin with five domains delineated as previously described. 8 Caspase-6 cleavage site is indicated by the red arrow. The polyglutamine tract is denoted by the green bar. (B) The biotinylated DNA aptamer MS1 (Biotin, first blot) was detected in uncleaved Q78-huntingtin, and the larger fragment from caspase-6 cleavage of Q78-huntingtin with an anti-biotin antibody. The representative blot showed that most of the aptamer appeared to be bound to the C-terminal fragment and un-cleaved huntingtin (arrows), but not the amino-terminal fragment (arrowhead), which was confirmed by re-probing the blot with a huntingtin C-terminal antibody (HF1, second blot) and an amino-terminal antibody (mAb2166, third blot). The experiment was repeated three times. (C) The same experiment was performed for the remaining biotinylated aptamers (MS2, MS3, MS4) and MS1. The representative immunoblot demonstrated that all aptamers preferentially bound to the C terminus of Q78-huntingtin. The experiment was repeated three times. (D and E) SLM analysis was performed with the huntingtin-aptamer complex and huntingtin alone. The methylated lysine residues were detected by LC/MS/MS and assigned a score based on the equation described in the Materials and Methods . The bar graphs showing the score of Q23- (D) and Q78- (E) huntingtin revealed that K2932/K2934 in CTD-II was the most probable binding site of MS3 on Q78-huntingtin. (F) Representative immunoblot showing purified Q23-, Q78-, Q78 K2449D-, Q78 K2932D/K2934D-, and Q78 K2449D/K2932D/K2934D-huntingtin bound with biotinylated MS3 or GCdx aptamers by probing with an anti-biotin antibody (Biotin) and an anti-huntingtin antibody (mAb2166). The location of full-length huntingtin is indicated by the arrow in both immunoblots. The asterisk marked a blurry band detected by anti-biotin antibody, but not by mAb2166, implying that a protein (or more than one) from insect cells, showing affinities to MS3 aptamer, was particularly enriched in Q78 K2449D-purified protein and less in Q78 K2932D/K2934D-purified proteins. The experiment was repeated three times.

    Article Snippet: Three lysine aspartic acid (KD) mutant huntingtin constructs (K2449D, K2932D/K2934D, K2449D/K2932D/K2934D) were generated by submission of the DNA sequences to Genscript (Piscataway, NJ, USA), which provided synthesized DNA in the pFastBac1 vector using SalI/SacII restriction digest and standard molecular biology techniques.

    Techniques: Mutagenesis, Binding Assay, Methylation, Liquid Chromatography with Mass Spectroscopy, Mass Spectrometry, Purification, Western Blot

    Site directed mutagenesis of the KRRIH region of Msm UdgX and its activity assay. ( A ( i ) and B ( i )) Line diagrams of Msm UdgX and its mutants highlighting the regions targeted by site directed mutagenesis. Motif A, motif B and the KRRIH loop region are shown in red, blue and yellow boxes, respectively. ( A ( ii )) Uracil excision/binding assays with Msm UdgX (WT) and its R107S, H109S and SSAS mutant derivatives. ∼200 ng of each protein was reacted with 5′ 32 P-labeled SSU9, the reaction mix was resolved on 8 M urea-PAGE (15%) and analyzed by phosphor imaging. The presence of uracil excision activity is seen by the fast migrating product band, ‘P’ whereas the uracil-DNA binding activity by the slow migrating complex, ‘C’. ( B ( ii )) Uracil excision/binding assays with Msm UdgX (WT) and its F4, F1, LD and F4 LD mutant derivatives (∼200 ng each) using SSU9. ( C ) Uracil release assay of Msm UdgX H109S (∼200 ng) using 3 H-uracil containing DNA (21 000 c.p.m) as substrate. Released uracil was separated by TLC (CEL 300 PEI/UV254) and seen as a fast migrating species upon phosphor imaging (the radioactive spot comigrated with the cold uracil spot detected by UV shadowing). ( D ) Uracil excision/binding assay with Msm UdgX and its H109S, H109G, H109Q, H109A and C27S derivatives. ∼200 ng of each protein was used in a standard UDG assay with SSU9 substrate.

    Journal: Nucleic Acids Research

    Article Title: A unique uracil-DNA binding protein of the uracil DNA glycosylase superfamily

    doi: 10.1093/nar/gkv854

    Figure Lengend Snippet: Site directed mutagenesis of the KRRIH region of Msm UdgX and its activity assay. ( A ( i ) and B ( i )) Line diagrams of Msm UdgX and its mutants highlighting the regions targeted by site directed mutagenesis. Motif A, motif B and the KRRIH loop region are shown in red, blue and yellow boxes, respectively. ( A ( ii )) Uracil excision/binding assays with Msm UdgX (WT) and its R107S, H109S and SSAS mutant derivatives. ∼200 ng of each protein was reacted with 5′ 32 P-labeled SSU9, the reaction mix was resolved on 8 M urea-PAGE (15%) and analyzed by phosphor imaging. The presence of uracil excision activity is seen by the fast migrating product band, ‘P’ whereas the uracil-DNA binding activity by the slow migrating complex, ‘C’. ( B ( ii )) Uracil excision/binding assays with Msm UdgX (WT) and its F4, F1, LD and F4 LD mutant derivatives (∼200 ng each) using SSU9. ( C ) Uracil release assay of Msm UdgX H109S (∼200 ng) using 3 H-uracil containing DNA (21 000 c.p.m) as substrate. Released uracil was separated by TLC (CEL 300 PEI/UV254) and seen as a fast migrating species upon phosphor imaging (the radioactive spot comigrated with the cold uracil spot detected by UV shadowing). ( D ) Uracil excision/binding assay with Msm UdgX and its H109S, H109G, H109Q, H109A and C27S derivatives. ∼200 ng of each protein was used in a standard UDG assay with SSU9 substrate.

    Article Snippet: The PCR product (662 bp) was blunt end ligated into pJET1.2 (MBI) vector to generate pJETMsm UdgX and confirmed by restriction digestion and DNA sequence analysis (Macrogen, S. Korea).

    Techniques: Mutagenesis, Activity Assay, Binding Assay, Labeling, Polyacrylamide Gel Electrophoresis, Imaging, Release Assay, Thin Layer Chromatography

    Activity assay of Msm UdgX. ( A ) Assay of Msm UdgX (100 ng) on 5′ 32 P-end labeled SSU9:G and SSU9 substrate, ‘S’ (∼10,000 c.p.m). Family 1 UDG (Ung) was used as control, Ugi was added as indicated. Reactions were resolved on 8 M Urea PAGE (15%) and analyzed by phosphor imaging. The presence of uracil excision activity can be seen as a fast migrating species forming a product ‘P’ whereas uracil binding activity form a slow migrating complex ‘C’ [panel (i) on left shows a diagrammatic sketch whereas the panel on right (ii) shows the results of the experiment]. ( B ) Proteinase K (1.2 U) digestion of the complex, ‘C’ observed in A, and re-resolved on 8 M Urea PAGE (15%) and analyzed by phosphor imaging. ( C ) Uracil release assay using 3 H-uracil containing DNA substrate. Msm UdgX (∼1 μg) or Ung (100 ng) was allowed to react with 3 H-uracil DNA substrate (∼21 000 c.p.m.), the samples were mixed with unlabeled uracil and spotted on the CEL 300 PEI/UV254, resolved using a mobile phase of 0.2% formic acid (v/v) containing 0.55 M LiCl at 4°C and analyzed by phosphor imaging. Released uracil is seen as a fast migrating species. ( D ) Binding of Msm UdgX (∼1 μg) with unlabeled uracil containing DNA oligomers (50 pmol) of different sizes resolved on 15% SDS PAGE and visualized by Coomassie blue staining. ( E ) Activity assay of Msm UdgX (4 pmol) in the presence of Msm Ung (4 pmol) using 5′ 32 P-labeled SSU9:G. Reactions were resolved on 8 M urea-PAGE (15%) and analyzed by phosphor imaging. The 1 st and 2 nd represents the order of addition, and ‘+’ in both Msm Ung and Msm UdgX indicates that both the protein are added simultaneously.

    Journal: Nucleic Acids Research

    Article Title: A unique uracil-DNA binding protein of the uracil DNA glycosylase superfamily

    doi: 10.1093/nar/gkv854

    Figure Lengend Snippet: Activity assay of Msm UdgX. ( A ) Assay of Msm UdgX (100 ng) on 5′ 32 P-end labeled SSU9:G and SSU9 substrate, ‘S’ (∼10,000 c.p.m). Family 1 UDG (Ung) was used as control, Ugi was added as indicated. Reactions were resolved on 8 M Urea PAGE (15%) and analyzed by phosphor imaging. The presence of uracil excision activity can be seen as a fast migrating species forming a product ‘P’ whereas uracil binding activity form a slow migrating complex ‘C’ [panel (i) on left shows a diagrammatic sketch whereas the panel on right (ii) shows the results of the experiment]. ( B ) Proteinase K (1.2 U) digestion of the complex, ‘C’ observed in A, and re-resolved on 8 M Urea PAGE (15%) and analyzed by phosphor imaging. ( C ) Uracil release assay using 3 H-uracil containing DNA substrate. Msm UdgX (∼1 μg) or Ung (100 ng) was allowed to react with 3 H-uracil DNA substrate (∼21 000 c.p.m.), the samples were mixed with unlabeled uracil and spotted on the CEL 300 PEI/UV254, resolved using a mobile phase of 0.2% formic acid (v/v) containing 0.55 M LiCl at 4°C and analyzed by phosphor imaging. Released uracil is seen as a fast migrating species. ( D ) Binding of Msm UdgX (∼1 μg) with unlabeled uracil containing DNA oligomers (50 pmol) of different sizes resolved on 15% SDS PAGE and visualized by Coomassie blue staining. ( E ) Activity assay of Msm UdgX (4 pmol) in the presence of Msm Ung (4 pmol) using 5′ 32 P-labeled SSU9:G. Reactions were resolved on 8 M urea-PAGE (15%) and analyzed by phosphor imaging. The 1 st and 2 nd represents the order of addition, and ‘+’ in both Msm Ung and Msm UdgX indicates that both the protein are added simultaneously.

    Article Snippet: The PCR product (662 bp) was blunt end ligated into pJET1.2 (MBI) vector to generate pJETMsm UdgX and confirmed by restriction digestion and DNA sequence analysis (Macrogen, S. Korea).

    Techniques: Activity Assay, Labeling, Polyacrylamide Gel Electrophoresis, Imaging, Binding Assay, Release Assay, SDS Page, Staining

    Proposed pathway for repair of uracil- Msm UdgX complexes. Binding of Msm UdgX to uracil would stall replication which could in turn lead to generation of single stranded gaps or double stranded breaks. Activation of RecA protein upon binding to ss DNA would induce the SOS response through cleavage of LexA. RecBCD proteins would also repair the complex through homologous recombination repair pathway.

    Journal: Nucleic Acids Research

    Article Title: A unique uracil-DNA binding protein of the uracil DNA glycosylase superfamily

    doi: 10.1093/nar/gkv854

    Figure Lengend Snippet: Proposed pathway for repair of uracil- Msm UdgX complexes. Binding of Msm UdgX to uracil would stall replication which could in turn lead to generation of single stranded gaps or double stranded breaks. Activation of RecA protein upon binding to ss DNA would induce the SOS response through cleavage of LexA. RecBCD proteins would also repair the complex through homologous recombination repair pathway.

    Article Snippet: The PCR product (662 bp) was blunt end ligated into pJET1.2 (MBI) vector to generate pJETMsm UdgX and confirmed by restriction digestion and DNA sequence analysis (Macrogen, S. Korea).

    Techniques: Binding Assay, Activation Assay, Homologous Recombination

    Telomere DNA G-quadruplex unfolding by arginine to alanine mutants of UP1+RGG monitored using CD spectroscopy. ( A, B ) Unfolding of K + form of Tel22 G-quadruplex DNA by TriRGG (A) and AllRGG (B) mutants. The G-quadruplex DNA was titrated with increasing molar excess of proteins. The black arrows in the spectra indicate the gradual decrease in ellipticity at 295 nm with increasing protein concentration. ( C ) The normalized ellipticity at 295 nm of Tel22 at the final titration step (at 1:6 molar ratio of DNA to protein) for UP1, AllRGG, TriRGG and UP1+RGG showing the relative foldedness of the G-quadruplex structure.

    Journal: Nucleic Acids Research

    Article Title: RGG-box in hnRNPA1 specifically recognizes the telomere G-quadruplex DNA and enhances the G-quadruplex unfolding ability of UP1 domain

    doi: 10.1093/nar/gky854

    Figure Lengend Snippet: Telomere DNA G-quadruplex unfolding by arginine to alanine mutants of UP1+RGG monitored using CD spectroscopy. ( A, B ) Unfolding of K + form of Tel22 G-quadruplex DNA by TriRGG (A) and AllRGG (B) mutants. The G-quadruplex DNA was titrated with increasing molar excess of proteins. The black arrows in the spectra indicate the gradual decrease in ellipticity at 295 nm with increasing protein concentration. ( C ) The normalized ellipticity at 295 nm of Tel22 at the final titration step (at 1:6 molar ratio of DNA to protein) for UP1, AllRGG, TriRGG and UP1+RGG showing the relative foldedness of the G-quadruplex structure.

    Article Snippet: DNA preparation for CD, NMR and fluorescence kinetics experiments The DNA sequences (Table ) for binding studies with UP1, UP1+RGG and the RGG-box were ordered from Eurofins except the abasicloops-Tel22 DNA that was ordered from Dharmacon.

    Techniques: Spectroscopy, Protein Concentration, Titration

    Interaction of RGG-box with the single stranded and G-quadruplex DNA monitored through NMR spectroscopy. ( A ) 2D 15 N– 1 H HSQC spectrum of the free RGG-box (black) and in complex with Tel22ss at 1:6 protein to DNA molar ratio (red). No significant chemical shift perturbations were observed for this interaction. Single stranded Tel22ss is shown as a cartoon. ( B ) 2D 15 N– 1 H HSQC spectrum of the RGG-box (black) and in complex with Tel22 at 1:6 protein to DNA molar ratio (red). Specific chemical shift perturbations were observed for several residues (marked with green arrows). A representative cartoon of monomeric G-quadruplex form of Tel22 is shown (only one conformation in K + ion is shown). ( C ) A subset of residues of RGG-box that show specific chemical shift perturbation upon addition of Tel22 is shown. The RGG-box and Tel22 complex was in fast exchange (weak binding) as we observed continuous movement of resonance peaks upon addition of increasing amount of the Tel22 DNA. Three steps of titration at different protein to DNA ratios are shown: black at 1:0, green at 1: 1, and red at 1:6. ( D ) The titration curves showing chemical shift change plotted as a function of increasing DNA:protein ratio for 14 interacting residues of the RGG-box is shown.

    Journal: Nucleic Acids Research

    Article Title: RGG-box in hnRNPA1 specifically recognizes the telomere G-quadruplex DNA and enhances the G-quadruplex unfolding ability of UP1 domain

    doi: 10.1093/nar/gky854

    Figure Lengend Snippet: Interaction of RGG-box with the single stranded and G-quadruplex DNA monitored through NMR spectroscopy. ( A ) 2D 15 N– 1 H HSQC spectrum of the free RGG-box (black) and in complex with Tel22ss at 1:6 protein to DNA molar ratio (red). No significant chemical shift perturbations were observed for this interaction. Single stranded Tel22ss is shown as a cartoon. ( B ) 2D 15 N– 1 H HSQC spectrum of the RGG-box (black) and in complex with Tel22 at 1:6 protein to DNA molar ratio (red). Specific chemical shift perturbations were observed for several residues (marked with green arrows). A representative cartoon of monomeric G-quadruplex form of Tel22 is shown (only one conformation in K + ion is shown). ( C ) A subset of residues of RGG-box that show specific chemical shift perturbation upon addition of Tel22 is shown. The RGG-box and Tel22 complex was in fast exchange (weak binding) as we observed continuous movement of resonance peaks upon addition of increasing amount of the Tel22 DNA. Three steps of titration at different protein to DNA ratios are shown: black at 1:0, green at 1: 1, and red at 1:6. ( D ) The titration curves showing chemical shift change plotted as a function of increasing DNA:protein ratio for 14 interacting residues of the RGG-box is shown.

    Article Snippet: DNA preparation for CD, NMR and fluorescence kinetics experiments The DNA sequences (Table ) for binding studies with UP1, UP1+RGG and the RGG-box were ordered from Eurofins except the abasicloops-Tel22 DNA that was ordered from Dharmacon.

    Techniques: Nuclear Magnetic Resonance, Spectroscopy, Binding Assay, Titration

    Telomere DNA G-quadruplex unfolding by UP1+RGG and UP1 monitored using CD spectroscopy. ( A, B ) Unfolding of K + and Na + forms of Tel22 G-quadruplex DNA by UP1. The G-quadruplex DNAs were titrated with increasing molar excess of proteins. The black arrow in the spectra indicates the gradual decrease in ellipticity at 295 nm with increasing protein concentration. ( C, D ) Unfolding of K + and Na + forms of Tel22 G-quadruplex DNA by UP1+RGG. The G-quadruplex DNAs were titrated with increasing molar excess of proteins. The black arrow in the spectra indicates the gradual decrease in ellipticity at 295 nm with increasing protein concentration. ( E, F ) The ellipticity at 295 nm was normalized and plotted to show the relative foldedness of both K + and Na + forms of quadruplexes upon UP1 or UP1+RGG addition at each step of titration.

    Journal: Nucleic Acids Research

    Article Title: RGG-box in hnRNPA1 specifically recognizes the telomere G-quadruplex DNA and enhances the G-quadruplex unfolding ability of UP1 domain

    doi: 10.1093/nar/gky854

    Figure Lengend Snippet: Telomere DNA G-quadruplex unfolding by UP1+RGG and UP1 monitored using CD spectroscopy. ( A, B ) Unfolding of K + and Na + forms of Tel22 G-quadruplex DNA by UP1. The G-quadruplex DNAs were titrated with increasing molar excess of proteins. The black arrow in the spectra indicates the gradual decrease in ellipticity at 295 nm with increasing protein concentration. ( C, D ) Unfolding of K + and Na + forms of Tel22 G-quadruplex DNA by UP1+RGG. The G-quadruplex DNAs were titrated with increasing molar excess of proteins. The black arrow in the spectra indicates the gradual decrease in ellipticity at 295 nm with increasing protein concentration. ( E, F ) The ellipticity at 295 nm was normalized and plotted to show the relative foldedness of both K + and Na + forms of quadruplexes upon UP1 or UP1+RGG addition at each step of titration.

    Article Snippet: DNA preparation for CD, NMR and fluorescence kinetics experiments The DNA sequences (Table ) for binding studies with UP1, UP1+RGG and the RGG-box were ordered from Eurofins except the abasicloops-Tel22 DNA that was ordered from Dharmacon.

    Techniques: Spectroscopy, Protein Concentration, Titration

    Telomere DNA G-quadruplex unfolding by UP1+RGG and UP1 monitored using NMR and fluorescence spectroscopy. ( A ) 1D 1 H NMR spectra of Na + form of Tel22 showing gradual loss of imino proton peaks upon titration with increasing concentrations of UP1 (blue) and UP1+RGG (red). ( B ) Unfolding of the 5′-FAM and 3′-TAMRA labeled K + form of Tel22 DNA G-quadruplex (5′FAM-Tel22-TAMRA3′) by UP1 (blue) and UP1+RGG (red) monitored by observing the emission of FAM at 516 nM. 5′FAM-Tel22-TAMRA3′ DNA was mixed with 4 molar equivalents of UP1 or UP1+RGG and the emission spectrum was recorded over a time period. ( C ) Proposed model for RGG-box assisted recognition and unfolding of telomere DNA G-quadruplex unfolding by UP1+RGG.

    Journal: Nucleic Acids Research

    Article Title: RGG-box in hnRNPA1 specifically recognizes the telomere G-quadruplex DNA and enhances the G-quadruplex unfolding ability of UP1 domain

    doi: 10.1093/nar/gky854

    Figure Lengend Snippet: Telomere DNA G-quadruplex unfolding by UP1+RGG and UP1 monitored using NMR and fluorescence spectroscopy. ( A ) 1D 1 H NMR spectra of Na + form of Tel22 showing gradual loss of imino proton peaks upon titration with increasing concentrations of UP1 (blue) and UP1+RGG (red). ( B ) Unfolding of the 5′-FAM and 3′-TAMRA labeled K + form of Tel22 DNA G-quadruplex (5′FAM-Tel22-TAMRA3′) by UP1 (blue) and UP1+RGG (red) monitored by observing the emission of FAM at 516 nM. 5′FAM-Tel22-TAMRA3′ DNA was mixed with 4 molar equivalents of UP1 or UP1+RGG and the emission spectrum was recorded over a time period. ( C ) Proposed model for RGG-box assisted recognition and unfolding of telomere DNA G-quadruplex unfolding by UP1+RGG.

    Article Snippet: DNA preparation for CD, NMR and fluorescence kinetics experiments The DNA sequences (Table ) for binding studies with UP1, UP1+RGG and the RGG-box were ordered from Eurofins except the abasicloops-Tel22 DNA that was ordered from Dharmacon.

    Techniques: Nuclear Magnetic Resonance, Fluorescence, Spectroscopy, Titration, Labeling

    Interaction of UP1+RGG and UP1 with the single stranded and G-quadruplex DNA monitored through ITC. Raw and fitted isotherms are shown and the equilibrium K d s obtained upon fitting of the raw data is mentioned in each panel. ( A, B ) Interaction of UP1 with the single stranded Tel22ss DNA in the presence of 100 mM NaCl and 100 mM KCl respectively. ( C, D ) Interaction of UP1+RGG with the single stranded Tel22ss DNA in the presence of 100 mM NaCl and 100 mM KCl respectively. ( E, F ) Interaction of UP1 with the Na + and K + forms of Tel22 G-quadruplex DNA respectively. ( G, H ) Interaction of UP1+RGG with the Na + and K + forms of Tel22 G-quadruplex DNA respectively.

    Journal: Nucleic Acids Research

    Article Title: RGG-box in hnRNPA1 specifically recognizes the telomere G-quadruplex DNA and enhances the G-quadruplex unfolding ability of UP1 domain

    doi: 10.1093/nar/gky854

    Figure Lengend Snippet: Interaction of UP1+RGG and UP1 with the single stranded and G-quadruplex DNA monitored through ITC. Raw and fitted isotherms are shown and the equilibrium K d s obtained upon fitting of the raw data is mentioned in each panel. ( A, B ) Interaction of UP1 with the single stranded Tel22ss DNA in the presence of 100 mM NaCl and 100 mM KCl respectively. ( C, D ) Interaction of UP1+RGG with the single stranded Tel22ss DNA in the presence of 100 mM NaCl and 100 mM KCl respectively. ( E, F ) Interaction of UP1 with the Na + and K + forms of Tel22 G-quadruplex DNA respectively. ( G, H ) Interaction of UP1+RGG with the Na + and K + forms of Tel22 G-quadruplex DNA respectively.

    Article Snippet: DNA preparation for CD, NMR and fluorescence kinetics experiments The DNA sequences (Table ) for binding studies with UP1, UP1+RGG and the RGG-box were ordered from Eurofins except the abasicloops-Tel22 DNA that was ordered from Dharmacon.

    Techniques: