| Autor: |
Christopher Sifuentes, Andrea Farina, Grace M. Moore, Erika S. Buechelmaier, Jonathan E. Leeman, Rama Rao Damerla, Neeman Mohibullah, Kyrie S. Anderson, Rahul Majumdar, Daniel S. Higginson, Himanshi Narang, Agnes Viale, Pavithran T. Ravindran, Simon N. Powell, Rekha Soni, Manisha Jalan, Suleman S. Hussain, Yi Li, Edin Hamzic, Xiaoqing Rong-Mullins |
| Rok vydání: |
2020 |
| Předmět: |
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| Zdroj: |
SSRN Electronic Journal. |
| ISSN: |
1556-5068 |
| Popis: |
Double strand break (DSB) repair mainly occurs through 3 pathways: non-homologous end-joining (NHEJ), alternative end-joining (Alt-EJ), and homologous recombination (HR). We present an assay system that enables simultaneous measurement of all three pathways using Cas9-generated DSBs and next generation sequencing to profile and quantify pathway choice. The assay system has provided several insights. First, absence of the key Alt-EJ factor Pol q only abrogates ~50% of total Alt-EJ. Second, single-strand templated repair (SSTR) requires BRCA1 and MRE11 activity, but not BRCA2, establishing that SSTR commonly used in genome editing is not conventional HR. Third, BRCA1 promotes Alt-EJ usage at two-ended DSBs in contrast to BRCA2. These fundamental differences between BRCA1 and BRCA2 deficiency have implications for therapeutic targeting of HR-deficient cancers. This assay can be used in any system which permits Cas9 delivery and, importantly, allows rapid genotype-to-phenotype correlation in isogenic cell line pairs. |
| Databáze: |
OpenAIRE |
| Externí odkaz: |
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