Shelterin-Like Proteins and Yku Inhibit Nucleolytic Processing of Saccharomyces cerevisiae Telomeres
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{"title"=>"Shelterin-like proteins and Yku inhibit nucleolytic processing of Saccharomyces cerevisiae telomeres", "type"=>"journal", "authors"=>[{"first_name"=>"Diego", "last_name"=>"Bonetti", "scopus_author_id"=>"18436180400"}, {"first_name"=>"Michela", "last_name"=>"Clerici", "scopus_author_id"=>"8590394000"}, {"first_name"=>"Savani", "last_name"=>"Anbalagan", "scopus_author_id"=>"36239688600"}, {"first_name"=>"Marina", "last_name"=>"Martina", "scopus_author_id"=>"35079071800"}, {"first_name"=>"Giovanna", "last_name"=>"Lucchini", "scopus_author_id"=>"7005358339"}, {"first_name"=>"Maria Pia", "last_name"=>"Longhese", "scopus_author_id"=>"6701366616"}], "year"=>2010, "source"=>"PLoS Genetics", "identifiers"=>{"issn"=>"15537390", "scopus"=>"2-s2.0-77953245442", "pui"=>"358967346", "doi"=>"10.1371/journal.pgen.1000966", "isbn"=>"2008215148", "sgr"=>"77953245442", "pmid"=>"20523746"}, "id"=>"3ef99fe8-7c9b-31f9-9878-ebf115e63046", "abstract"=>"Eukaryotic cells distinguish their chromosome ends from accidental DNA double-strand breaks (DSBs) by packaging them into protective structures called telomeres that prevent DNA repair/recombination activities. Here we investigate the role of key telomeric proteins in protecting budding yeast telomeres from degradation. We show that the Saccharomyces cerevisiae shelterin-like proteins Rif1, Rif2, and Rap1 inhibit nucleolytic processing at both de novo and native telomeres during G1 and G2 cell cycle phases, with Rif2 and Rap1 showing the strongest effects. Also Yku prevents telomere resection in G1, independently of its role in non-homologous end joining. Yku and the shelterin-like proteins have additive effects in inhibiting DNA degradation at G1 de novo telomeres, where Yku plays the major role in preventing initiation, whereas Rif1, Rif2, and Rap1 act primarily by limiting extensive resection. In fact, exonucleolytic degradation of a de novo telomere is more efficient in yku70Delta than in rif2Delta G1 cells, but generation of ssDNA in Yku-lacking cells is limited to DNA regions close to the telomere tip. This limited processing is due to the inhibitory action of Rap1, Rif1, and Rif2, as their inactivation allows extensive telomere resection not only in wild-type but also in yku70Delta G1 cells. Finally, Rap1 and Rif2 prevent telomere degradation by inhibiting MRX access to telomeres, which are also protected from the Exo1 nuclease by Yku. Thus, chromosome end degradation is controlled by telomeric proteins that specifically inhibit the action of different nucleases.", "link"=>"http://www.mendeley.com/research/shelterinlike-proteins-yku-inhibit-nucleolytic-processing-saccharomyces-cerevisiae-telomeres", "reader_count"=>55, "reader_count_by_academic_status"=>{"Researcher"=>22, "Student > Doctoral Student"=>3, "Student > Ph. D. Student"=>19, "Student > Postgraduate"=>3, "Student > Master"=>3, "Student > Bachelor"=>2, "Professor"=>3}, "reader_count_by_user_role"=>{"Researcher"=>22, "Student > Doctoral Student"=>3, "Student > Ph. D. Student"=>19, "Student > Postgraduate"=>3, "Student > Master"=>3, "Student > Bachelor"=>2, "Professor"=>3}, "reader_count_by_subject_area"=>{"Unspecified"=>1, "Biochemistry, Genetics and Molecular Biology"=>10, "Agricultural and Biological Sciences"=>44}, "reader_count_by_subdiscipline"=>{"Agricultural and Biological Sciences"=>{"Agricultural and Biological Sciences"=>44}, "Biochemistry, Genetics and Molecular Biology"=>{"Biochemistry, Genetics and Molecular Biology"=>10}, "Unspecified"=>{"Unspecified"=>1}}, "reader_count_by_country"=>{"Canada"=>1, "United States"=>4, "France"=>1, "Germany"=>1}, "group_count"=>1}

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Figshare

  • {"files"=>["https://ndownloader.figshare.com/files/847530"], "description"=>"<p>(A–F) HO expression was induced at time zero by galactose addition to α-factor-arrested cells with the indicated genotypes that were then kept arrested in G1. (A) RsaI- and EcoRV-digested genomic DNA was hybridized with probe B as in <a href=\"http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1000966#pgen-1000966-g001\" target=\"_blank\">Figure 1E</a>. (B) Densitometric analysis. Plotted values are the mean value ±SD from three independent experiments as in (A). (C) RsaI-digested genomic DNA was hybridized with probe A as in <a href=\"http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1000966#pgen-1000966-g003\" target=\"_blank\">Figure 3E</a>. (D) Densitometric analysis. Plotted values are the mean value ±SD from three independent experiments as in (C). (E) RsaI- and EcoRV-digested genomic DNA was hybridized with probe B as in <a href=\"http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1000966#pgen-1000966-g001\" target=\"_blank\">Figure 1E</a>. (F) Densitometric analysis. Plotted values are the mean value ±SD from three independent experiments as in (E). (G,H) HO expression was induced at time zero by galactose addition to nocodazole-arrested cells with the indicated genotypes that were then kept arrested in G2. (G) RsaI- and EcoRV-digested genomic DNA was hybridized with probe B as in <a href=\"http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1000966#pgen-1000966-g001\" target=\"_blank\">Figure 1E</a>. (H) Densitometric analysis. Plotted values are the mean value ±SD from three independent experiments as in (G).</p>", "links"=>[], "tags"=>["rap1", "inactivation", "enhances", "resection", "de", "novo", "telomere"], "article_id"=>517978, "categories"=>["Genetics", "Molecular Biology"], "users"=>["Diego Bonetti", "Michela Clerici", "Savani Anbalagan", "Marina Martina", "Giovanna Lucchini", "Maria Pia Longhese"], "doi"=>"https://dx.doi.org/10.1371/journal.pgen.1000966.g004", "stats"=>{"downloads"=>0, "page_views"=>0, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Rif2_and_Rap1_inactivation_enhances_resection_at_a_de_novo_telomere_in_yku70_916_cells_/517978", "title"=>"Rif2 and Rap1 inactivation enhances resection at a de novo telomere in <i>yku70Δ</i> cells.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2010-05-27 02:12:58"}
  • {"files"=>["https://ndownloader.figshare.com/files/848149"], "description"=>"<p>In G1, Yku protects telomeres from Exo1, while Rap1, Rif1 and Rif2 mainly act by preventing MRX access. As MRX action is still inhibited by Rap1, Rif1 and Rif2 in <i>yku70Δ</i> G1 cells, Yku might protect G1 telomeres also from MRX. In G2, only Rap1 and Rif2 still exert their inhibitory effects on telomere processing. Telomere resection can take place in G2 because Yku does not exert its inhibitory effect and Cdk1 activity potentiates nuclease actions.</p>", "links"=>[], "tags"=>["limiting", "dna", "degradation"], "article_id"=>518606, "categories"=>["Genetics", "Molecular Biology"], "users"=>["Diego Bonetti", "Michela Clerici", "Savani Anbalagan", "Marina Martina", "Giovanna Lucchini", "Maria Pia Longhese"], "doi"=>"https://dx.doi.org/10.1371/journal.pgen.1000966.g008", "stats"=>{"downloads"=>1, "page_views"=>3, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_A_working_model_for_limiting_DNA_degradation_at_telomeres_/518606", "title"=>"A working model for limiting DNA degradation at telomeres.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2010-05-27 02:23:26"}
  • {"files"=>["https://ndownloader.figshare.com/files/847038"], "description"=>"<p>(A) The HO-induced telomere system. Galactose-induced HO endonuclease generates a single DSB at an HO cleavage site (HOcs) adjacent to an 81-bp TG repeat sequence (TG tracts) that is inserted at the <i>ADH4</i> locus on chromosome VII. RsaI- and EcoRV-digested genomic DNA was hybridized with two single-stranded riboprobes, which anneal to either the 5′ C-strand (probe A) or the 3′ G-strand (probe B) to a site located 212 bp from the HO cutting site. Both probes reveal an uncut 390 nt DNA fragment (uncut), which is converted by HO cleavage into a 166 nt fragment (cut) that can be detected by both probe A (5′ C-strand) and probe B (3′ G-strand). Degradation of the 5′ C-strand leads to disappearance of the probe A signal as resection proceeds beyond the hybridization region. Furthermore, it eliminates the cutting sites for the EcoRV (E) and RsaI (R) restriction enzymes, thus converting the 3′ cut G-strand into longer r1 (304 nt) and r2 (346 nt) DNA fragments detected by probe B. Both probes also detects a 138 nt fragment from the <i>ade2-101</i> locus on Chr. XV (INT), which serves as internal loading control. (B–F) HO expression was induced at time zero by galactose addition to α-factor-arrested wild type (YLL2599) and otherwise isogenic <i>rif2Δ</i>, <i>rap1ΔC</i> and <i>rif1Δ</i> cell cultures that were then kept arrested in G1. (B) FACS analysis of DNA content. (C) RsaI- and EcoRV-digested genomic DNA was hybridized with probe A. Degradation of the 5′ C-strand leads to the disappearance of the 166 nt signal (cut C-strand) generated by this probe. (D) Densitometric analysis. Plotted values are the mean value ±SD from three independent experiments as in (C). (E) The same RsaI- and EcoRV-digested genomic DNA analyzed in (C) was hybridized with probe B. Degradation of the 5′ C-strand leads to the conversion of the 3′ cut G-strand 166 nt fragment into the slower migrating r1 DNA fragment described in (A). (F) Densitometric analysis. Plotted values are the mean value ±SD from three independent experiments as in (E).</p>", "links"=>[], "tags"=>["rif2", "inhibit", "resection", "de", "novo", "telomere"], "article_id"=>517488, "categories"=>["Genetics", "Molecular Biology"], "users"=>["Diego Bonetti", "Michela Clerici", "Savani Anbalagan", "Marina Martina", "Giovanna Lucchini", "Maria Pia Longhese"], "doi"=>"https://dx.doi.org/10.1371/journal.pgen.1000966.g001", "stats"=>{"downloads"=>1, "page_views"=>13, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Rap1_Rif1_and_Rif2_inhibit_resection_at_a_de_novo_telomere_in_G1_/517488", "title"=>"Rap1, Rif1, and Rif2 inhibit resection at a de novo telomere in G1.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2010-05-27 02:04:48"}
  • {"files"=>["https://ndownloader.figshare.com/files/847702"], "description"=>"<p>(A,B) G1-arrested (G1) wild type (YLL2599) and otherwise isogenic <i>yku70Δ</i> cell cultures were incubated at either 23°C or 37°C for 4 hours in the presence of α-factor. (A) Genomic DNA was digested with XhoI and single-stranded telomere overhangs were visualized by in-gel hybridization (native gel) using an end-labeled C-rich oligonucleotide <a href=\"http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1000966#pgen.1000966-Dionne1\" target=\"_blank\">[31]</a>. The same DNA samples were separated on a 0.8% agarose gel, denatured and hybridized with the end-labeled C-rich oligonucleotide for loading and telomere length control (denatured gel). (B) FACS analysis of DNA content. (C) Genomic DNA prepared from wild type (YLL2599) and otherwise isogenic <i>rif2Δ</i> and <i>rap1ΔC</i> cell cultures, exponentially growing at 25°C, was digested with XhoI and the single-stranded telomere overhangs were visualized by in-gel hybridization as in (A). (D) Wild type (YLL2599) and otherwise isogenic <i>yku70Δ</i> cell cultures exponentially growing (cyc) at 23°C were incubated at 37°C for the indicated time points. G1-arrested wild type and <i>yku70Δ</i> cells (G1) were incubated at either 23°C or 37°C for 4 hours. Rad53 was visualized at the indicated times by western analysis with anti-Rad53 antibodies. (E) α-factor arrested wild type (YLL2599) and otherwise isogenic <i>rif2Δ</i> and <i>rap1ΔC</i> cell cultures were released into the cell cycle at 25°C. Rad53 was visualized as in (D). (F) α-factor-arrested <i>cdc13-1</i> and <i>cdc13-1 rif2Δ</i> cells were released into the cell cycle at 28°C. Rad53 was visualized as in (D).</p>", "links"=>[], "tags"=>["single-stranded", "overhangs"], "article_id"=>518158, "categories"=>["Genetics", "Molecular Biology"], "users"=>["Diego Bonetti", "Michela Clerici", "Savani Anbalagan", "Marina Martina", "Giovanna Lucchini", "Maria Pia Longhese"], "doi"=>"https://dx.doi.org/10.1371/journal.pgen.1000966.g005", "stats"=>{"downloads"=>0, "page_views"=>1, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Analysis_of_single_stranded_overhangs_at_native_telomeres_/518158", "title"=>"Analysis of single-stranded overhangs at native telomeres.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2010-05-27 02:15:58"}
  • {"files"=>["https://ndownloader.figshare.com/files/847172"], "description"=>"<p>HO expression was induced at time zero by galactose addition to nocodazole-arrested wild type (YLL2599) and otherwise isogenic <i>rif2Δ</i>, <i>rap1ΔC</i> and <i>rif1Δ</i> cell cultures that were then kept arrested in G2. (A) FACS analysis of DNA content. (B) RsaI- and EcoRV-digested genomic DNA was hybridized with probe B as in <a href=\"http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1000966#pgen-1000966-g001\" target=\"_blank\">Figure 1E</a>. (C) Densitometric analysis. Plotted values are the mean value ±SD from three independent experiments as in (B).</p>", "links"=>[], "tags"=>["rif2", "inhibit", "resection", "de", "novo", "telomere"], "article_id"=>517623, "categories"=>["Genetics", "Molecular Biology"], "users"=>["Diego Bonetti", "Michela Clerici", "Savani Anbalagan", "Marina Martina", "Giovanna Lucchini", "Maria Pia Longhese"], "doi"=>"https://dx.doi.org/10.1371/journal.pgen.1000966.g002", "stats"=>{"downloads"=>1, "page_views"=>8, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Rap1_Rif1_and_Rif2_inhibit_resection_at_a_de_novo_telomere_in_G2_/517623", "title"=>"Rap1, Rif1, and Rif2 inhibit resection at a de novo telomere in G2.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2010-05-27 02:07:03"}
  • {"files"=>["https://ndownloader.figshare.com/files/421627"], "description"=>"<div><p>Eukaryotic cells distinguish their chromosome ends from accidental DNA double-strand breaks (DSBs) by packaging them into protective structures called telomeres that prevent DNA repair/recombination activities. Here we investigate the role of key telomeric proteins in protecting budding yeast telomeres from degradation. We show that the <em>Saccharomyces cerevisiae</em> shelterin-like proteins Rif1, Rif2, and Rap1 inhibit nucleolytic processing at both de novo and native telomeres during G1 and G2 cell cycle phases, with Rif2 and Rap1 showing the strongest effects. Also Yku prevents telomere resection in G1, independently of its role in non-homologous end joining. Yku and the shelterin-like proteins have additive effects in inhibiting DNA degradation at G1 de novo telomeres, where Yku plays the major role in preventing initiation, whereas Rif1, Rif2, and Rap1 act primarily by limiting extensive resection. In fact, exonucleolytic degradation of a de novo telomere is more efficient in <em>yku70Δ</em> than in <em>rif2Δ</em> G1 cells, but generation of ssDNA in Yku-lacking cells is limited to DNA regions close to the telomere tip. This limited processing is due to the inhibitory action of Rap1, Rif1, and Rif2, as their inactivation allows extensive telomere resection not only in wild-type but also in <em>yku70Δ</em> G1 cells. Finally, Rap1 and Rif2 prevent telomere degradation by inhibiting MRX access to telomeres, which are also protected from the Exo1 nuclease by Yku. Thus, chromosome end degradation is controlled by telomeric proteins that specifically inhibit the action of different nucleases.</p></div>", "links"=>[], "tags"=>["shelterin-like", "proteins", "yku", "inhibit", "nucleolytic", "telomeres"], "article_id"=>143269, "categories"=>["Genetics", "Molecular Biology"], "users"=>["Diego Bonetti", "Michela Clerici", "Savani Anbalagan", "Marina Martina", "Giovanna Lucchini", "Maria Pia Longhese"], "doi"=>"https://dx.doi.org/10.1371/journal.pgen.1000966", "stats"=>{"downloads"=>14, "page_views"=>11, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/Shelterin_Like_Proteins_and_Yku_Inhibit_Nucleolytic_Processing_of_Saccharomyces_cerevisiae_Telomeres/143269", "title"=>"Shelterin-Like Proteins and Yku Inhibit Nucleolytic Processing of <em>Saccharomyces cerevisiae</em> Telomeres", "pos_in_sequence"=>0, "defined_type"=>3, "published_date"=>"2010-05-27 00:54:29"}
  • {"files"=>["https://ndownloader.figshare.com/files/847310"], "description"=>"<p>(A–C) HO expression was induced at time zero by galactose addition to nocodazole-arrested wild type (YLL2599) and otherwise isogenic <i>yku70Δ</i> cell cultures that were then kept arrested in G2. (A) FACS analysis of DNA content. (B) RsaI- and EcoRV-digested genomic DNA was hybridized with probe A as described in <a href=\"http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1000966#pgen-1000966-g001\" target=\"_blank\">Figure 1C</a>. (C) Densitometric analysis. Plotted values are the mean value ±SD from three independent experiments as in (B). (D–F) HO expression was induced at time zero by galactose addition to α-factor-arrested wild type (YLL2599) and otherwise isogenic <i>yku70Δ</i> and <i>dnl4Δ</i> cell cultures that were then kept arrested in G1. (D) FACS analysis of DNA content. (E) RsaI-digested genomic DNA was hybridized with the single-stranded riboprobe A described in <a href=\"http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1000966#pgen-1000966-g001\" target=\"_blank\">Figure 1A</a>, which anneals to the 5′ C-strand and reveals an uncut 460 nt DNA fragment (uncut). After HO cleavage, this fragment is converted into a 304 nt fragment (cut) detected by the same probe (cut C-strand). (F) Densitometric analysis. Plotted values are the mean value ±SD from three independent experiments as in (E). (G) The system used to generate an HO-induced DSB. Hybridization of EcoRV-digested genomic DNA with a probe that anneals to the 5′ strand to a site located 215 nt from the HO cutting site reveals a 430 nt HO-cut 5′-strand fragment. Loss of the 5′ strand beyond the hybridization region leads to disappearance of the signal generated by the probe. (H–L) HO expression was induced at time zero by galactose addition to α-factor-arrested wild type (YLL2600) and otherwise isogenic <i>yku70Δ</i> and <i>dnl4Δ</i> cells, all carrying the system in (G). Cells were then kept arrested in G1. (H) FACS analysis of DNA content. (I) EcoRV-digested genomic DNA was hybridized with the probe indicated in (G). The INT band, corresponding to a chromosome IV sequence, serves as internal loading control. (L) Densitometric analysis. Plotted values are the mean value ±SD from three independent experiments as in (I).</p>", "links"=>[], "tags"=>["inhibits", "resection", "de", "novo", "telomere"], "article_id"=>517764, "categories"=>["Genetics", "Molecular Biology"], "users"=>["Diego Bonetti", "Michela Clerici", "Savani Anbalagan", "Marina Martina", "Giovanna Lucchini", "Maria Pia Longhese"], "doi"=>"https://dx.doi.org/10.1371/journal.pgen.1000966.g003", "stats"=>{"downloads"=>3, "page_views"=>5, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Yku_inhibits_resection_at_a_de_novo_telomere_specifically_in_G1_/517764", "title"=>"Yku inhibits resection at a de novo telomere specifically in G1.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2010-05-27 02:09:24"}
  • {"files"=>["https://ndownloader.figshare.com/files/847853"], "description"=>"<p>(A) G1-arrested cells were incubated at 37°C for 4 hours in the presence of α-factor. Genomic DNA was analyzed as in <a href=\"http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1000966#pgen-1000966-g005\" target=\"_blank\">Figure 5A</a>. (B) Exponentially growing cells were incubated at 37°C for 4 hours. Genomic DNA was analyzed as in <a href=\"http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1000966#pgen-1000966-g005\" target=\"_blank\">Figure 5A</a>. (C,D) Genomic DNA prepared from exponentially growing cells at 25°C was analyzed as in <a href=\"http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1000966#pgen-1000966-g005\" target=\"_blank\">Figure 5A</a>.</p>", "links"=>[], "tags"=>["requirements", "ssdna"], "article_id"=>518309, "categories"=>["Genetics", "Molecular Biology"], "users"=>["Diego Bonetti", "Michela Clerici", "Savani Anbalagan", "Marina Martina", "Giovanna Lucchini", "Maria Pia Longhese"], "doi"=>"https://dx.doi.org/10.1371/journal.pgen.1000966.g006", "stats"=>{"downloads"=>0, "page_views"=>0, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Nuclease_requirements_for_ssDNA_generation_at_native_telomeres_/518309", "title"=>"Nuclease requirements for ssDNA generation at native telomeres.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2010-05-27 02:18:29"}
  • {"files"=>["https://ndownloader.figshare.com/files/847995"], "description"=>"<p>(A–C) HO expression was induced at time zero by galactose addition to α-factor-arrested <i>yku70Δ</i>, <i>yku70Δ exo1Δ</i> and <i>yku70Δ mre11Δ</i> cell cultures that were then kept arrested in G1. (A) FACS analysis of DNA content. (B) RsaI-digested genomic DNA was hybridized with probe A as in <a href=\"http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1000966#pgen-1000966-g003\" target=\"_blank\">Figure 3E</a>. (C) Densitometric analysis. Plotted values are the mean value ±SD from three independent experiments as in (B). (D–F) HO expression was induced at time zero by galactose addition to α-factor-arrested <i>rif2Δ</i>, <i>rif2Δ exo1Δ</i> and <i>rif2Δ mre11Δ</i> cell cultures that were then kept arrested in G1. (D) FACS analysis of DNA content. (E) RsaI- and EcoRV-digested genomic DNA was hybridized with probe A as described in <a href=\"http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1000966#pgen-1000966-g001\" target=\"_blank\">Figure 1C</a>. (F) Densitometric analysis. Plotted values are the mean value ±SD from three independent experiments as in (E). (G) HO expression was induced at time zero by galactose addition to α-factor-arrested wild type, <i>rif2Δ</i> and <i>rap1ΔC</i> cells, all expressing a fully functional <i>MRE11-MYC</i> tagged allele. Cells were then kept arrested in G1 and chromatin samples taken at different times after HO induction were immunoprecipitated with anti-Myc antibody. Coimmunoprecipitated DNA was analyzed by quantitative real-time PCR (qPCR) using primer pairs located at the nontelomeric <i>ARO1</i> fragment of chromosome IV (CON) and 640 bp proximal to the HO site (TEL), respectively. Data are expressed as relative fold enrichment of TEL over CON signal after normalization to input signals for each primer set. The data presented are the mean of those obtained in three independent experiments. Error bars indicate s. d.</p>", "links"=>[], "tags"=>["requirements", "ssdna", "de", "novo"], "article_id"=>518450, "categories"=>["Genetics", "Molecular Biology"], "users"=>["Diego Bonetti", "Michela Clerici", "Savani Anbalagan", "Marina Martina", "Giovanna Lucchini", "Maria Pia Longhese"], "doi"=>"https://dx.doi.org/10.1371/journal.pgen.1000966.g007", "stats"=>{"downloads"=>1, "page_views"=>5, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Nuclease_requirements_for_ssDNA_generation_at_a_de_novo_telomere_/518450", "title"=>"Nuclease requirements for ssDNA generation at a de novo telomere.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2010-05-27 02:20:50"}

PMC Usage Stats | Further Information

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  • {"unique-ip"=>"4", "full-text"=>"4", "pdf"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"1", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2018", "month"=>"12"}
  • {"unique-ip"=>"3", "full-text"=>"2", "pdf"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"1", "cited-by"=>"0", "year"=>"2018", "month"=>"11"}
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Relative Metric

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