{"id":18035,"date":"2021-05-14T14:29:07","date_gmt":"2021-05-14T18:29:07","guid":{"rendered":"https:\/\/www.med.unc.edu\/biochem\/?p=18035"},"modified":"2021-05-18T14:48:15","modified_gmt":"2021-05-18T18:48:15","slug":"gpgupta-ramsden-new-pub-nucleic-acids-research-ramsden-gpgupta-labs-publish-new-dna-repair-research-in-nucleic-acids-research","status":"publish","type":"post","link":"https:\/\/www.med.unc.edu\/biochem\/news\/gpgupta-ramsden-new-pub-nucleic-acids-research-ramsden-gpgupta-labs-publish-new-dna-repair-research-in-nucleic-acids-research\/","title":{"rendered":"G.P.Gupta &amp; Ramsden labs publish new dna repair research in Nucleic Acids Research"},"content":{"rendered":"<p><img loading=\"lazy\" decoding=\"async\" class=\"size-medium wp-image-18040 alignright\" src=\"https:\/\/www.med.unc.edu\/biochem\/wp-content\/uploads\/sites\/795\/2021\/05\/gupta-ramsden-300x142.png\" alt=\"G Gupta and D Ramsden\" width=\"300\" height=\"142\" srcset=\"https:\/\/www.med.unc.edu\/biochem\/wp-content\/uploads\/sites\/795\/2021\/05\/gupta-ramsden-300x142.png 300w, https:\/\/www.med.unc.edu\/biochem\/wp-content\/uploads\/sites\/795\/2021\/05\/gupta-ramsden.png 490w\" sizes=\"auto, (max-width: 300px) 100vw, 300px\" \/>Congrats on the new publication: G.P.Gupta and Ramsden labs! &#8220;Marker-free quantification of repair pathway utilization at Cas9-induced double-strand breaks&#8221; Nucleic Acids Research. This was a collaborative effort with\u00a0UNC Biochemistry and Biophysics, UNC Lineberger Comprehensive Cancer Center, UNC Department of Radiation Oncology, The Curriculum in Genetics and Molecular Biology, and UNC Pharmacy.<\/p>\n<p>Interested in DSB repair pathway choice? This paper is on NGS and dPCR methods to quantify DSB repair pathway use, without needing fluorescent reporters, in essentially any cell type.<\/p>\n<h2 id=\"247050768\" class=\"abstract-title js-splitscreen-abstract-title\">Abstract<\/h2>\n<section class=\"abstract\">\n<p class=\"chapter-para\">Genome integrity and genome engineering require efficient repair of DNA double-strand breaks (DSBs) by non-homologous end joining (NHEJ), homologous recombination (HR), or alternative end-joining pathways. Here we describe two complementary methods for marker-free quantification of DSB repair pathway utilization at Cas9-targeted chromosomal DSBs in mammalian cells. The first assay features the analysis of amplicon next-generation sequencing data using ScarMapper, an iterative break-associated alignment algorithm to classify individual repair products based on deletion size, microhomology usage, and insertions. The second assay uses repair pathway-specific droplet digital PCR assays (\u2018PathSig-dPCR\u2019) for absolute quantification of signature DSB repair outcomes. We show that ScarMapper and PathSig-dPCR enable comprehensive assessment of repair pathway utilization in different cell models, after a variety of experimental perturbations. We use these assays to measure the differential impact of DNA end resection on NHEJ, HR and polymerase theta-mediated end joining (TMEJ) repair. These approaches are adaptable to any cellular model system and genomic locus where Cas9-mediated targeting is feasible. Thus, ScarMapper and PathSig-dPCR allow for systematic fate mapping of a targeted DSB with facile and accurate quantification of DSB repair pathway choice at endogenous chromosomal loci.<\/p>\n<\/section>\n<p>Wanjuan Feng, Dennis A Simpson, Jang-Eun Cho, Juan Carvajal-Garcia, Chelsea\u00a0M Smith, Kathryn M Headley, Nate Hathaway, Dale A Ramsden, Gaorav\u00a0P Gupta, Marker-free quantification of repair pathway utilization at Cas9-induced double-strand breaks,\u00a0<em>Nucleic Acids Research<\/em>, 2021;, gkab299,\u00a0<a href=\"https:\/\/doi.org\/10.1093\/nar\/gkab299\">https:\/\/doi.org\/10.1093\/nar\/gkab299<\/a><\/p>\n<p>Congrats to all authors in UNC Biochemistry and Biophysics, UNC Lineberger Comprehensive Cancer Center, UNC Department of Radiation Oncology, The Curriculum in Genetics and Molecular Biology, and UNC Pharmacy.<\/p>\n","protected":false},"excerpt":{"rendered":"<p>Congrats on the new publication: G.P.Gupta and Ramsden labs! &#8220;Marker-free quantification of repair pathway utilization at Cas9-induced double-strand breaks&#8221; Nucleic Acids Research. This was a collaborative effort with\u00a0UNC Biochemistry and Biophysics, UNC Lineberger Comprehensive Cancer Center, UNC Department of Radiation Oncology, The Curriculum in Genetics and Molecular Biology, and UNC Pharmacy. Interested in DSB repair &hellip; <a href=\"https:\/\/www.med.unc.edu\/biochem\/news\/gpgupta-ramsden-new-pub-nucleic-acids-research-ramsden-gpgupta-labs-publish-new-dna-repair-research-in-nucleic-acids-research\/\" aria-label=\"Read more about G.P.Gupta &amp; Ramsden labs publish new dna repair research in Nucleic Acids Research\">Read more<\/a><\/p>\n","protected":false},"author":41619,"featured_media":18040,"comment_status":"closed","ping_status":"closed","sticky":false,"template":"","format":"standard","meta":{"_acf_changed":false,"layout":"","cellInformation":"","apiCallInformation":"","footnotes":"","_links_to":"","_links_to_target":""},"categories":[2],"tags":[10,363,179,13],"class_list":["post-18035","post","type-post","status-publish","format-standard","has-post-thumbnail","hentry","category-news","tag-news_faculty","tag-news_2021","tag-news_postdoc-scholars","tag-news_students","odd"],"acf":[],"yoast_head":"<!-- This site is optimized with the Yoast SEO plugin v26.8 - https:\/\/yoast.com\/product\/yoast-seo-wordpress\/ -->\n<title>G.P.Gupta &amp; Ramsden labs publish new dna repair research in Nucleic Acids Research | Biochemistry and Biophysics<\/title>\n<meta name=\"robots\" content=\"index, follow, max-snippet:-1, max-image-preview:large, max-video-preview:-1\" \/>\n<link rel=\"canonical\" href=\"https:\/\/www.med.unc.edu\/biochem\/news\/gpgupta-ramsden-new-pub-nucleic-acids-research-ramsden-gpgupta-labs-publish-new-dna-repair-research-in-nucleic-acids-research\/\" \/>\n<meta property=\"og:locale\" content=\"en_US\" \/>\n<meta property=\"og:type\" content=\"article\" \/>\n<meta property=\"og:title\" content=\"G.P.Gupta &amp; Ramsden labs publish new dna repair research in Nucleic Acids Research | Biochemistry and Biophysics\" \/>\n<meta property=\"og:description\" content=\"Congrats on the new publication: G.P.Gupta and Ramsden labs! &#8220;Marker-free quantification of repair pathway utilization at Cas9-induced double-strand breaks&#8221; Nucleic Acids Research. This was a collaborative effort with\u00a0UNC Biochemistry and Biophysics, UNC Lineberger Comprehensive Cancer Center, UNC Department of Radiation Oncology, The Curriculum in Genetics and Molecular Biology, and UNC Pharmacy. Interested in DSB repair &hellip; Read more\" \/>\n<meta property=\"og:url\" content=\"https:\/\/www.med.unc.edu\/biochem\/news\/gpgupta-ramsden-new-pub-nucleic-acids-research-ramsden-gpgupta-labs-publish-new-dna-repair-research-in-nucleic-acids-research\/\" \/>\n<meta property=\"og:site_name\" content=\"Biochemistry and Biophysics\" \/>\n<meta property=\"article:publisher\" content=\"https:\/\/www.facebook.com\/uncbiochemistryandbiophysics\/\" \/>\n<meta property=\"article:published_time\" content=\"2021-05-14T18:29:07+00:00\" \/>\n<meta property=\"article:modified_time\" content=\"2021-05-18T18:48:15+00:00\" \/>\n<meta property=\"og:image\" content=\"https:\/\/www.med.unc.edu\/biochem\/wp-content\/uploads\/sites\/795\/2021\/05\/gupta-ramsden.png\" \/>\n\t<meta property=\"og:image:width\" content=\"490\" \/>\n\t<meta property=\"og:image:height\" content=\"232\" \/>\n\t<meta property=\"og:image:type\" content=\"image\/png\" \/>\n<meta name=\"author\" content=\"Carolyn Clabo\" \/>\n<meta name=\"twitter:card\" content=\"summary_large_image\" \/>\n<meta name=\"twitter:creator\" content=\"@UNC_BCBP\" \/>\n<meta name=\"twitter:site\" content=\"@UNC_BCBP\" \/>\n<meta name=\"twitter:label1\" content=\"Written by\" \/>\n\t<meta name=\"twitter:data1\" content=\"Carolyn Clabo\" \/>\n\t<meta name=\"twitter:label2\" content=\"Est. reading time\" \/>\n\t<meta name=\"twitter:data2\" content=\"2 minutes\" \/>\n<script type=\"application\/ld+json\" class=\"yoast-schema-graph\">{\"@context\":\"https:\/\/schema.org\",\"@graph\":[{\"@type\":\"Article\",\"@id\":\"https:\/\/www.med.unc.edu\/biochem\/news\/gpgupta-ramsden-new-pub-nucleic-acids-research-ramsden-gpgupta-labs-publish-new-dna-repair-research-in-nucleic-acids-research\/#article\",\"isPartOf\":{\"@id\":\"https:\/\/www.med.unc.edu\/biochem\/news\/gpgupta-ramsden-new-pub-nucleic-acids-research-ramsden-gpgupta-labs-publish-new-dna-repair-research-in-nucleic-acids-research\/\"},\"author\":{\"name\":\"Carolyn Clabo\",\"@id\":\"https:\/\/www.med.unc.edu\/biochem\/#\/schema\/person\/9693a4e0a76e8208ca2105ae25587332\"},\"headline\":\"G.P.Gupta &amp; 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