{"id":13992,"date":"2018-01-05T11:20:35","date_gmt":"2018-01-05T10:20:35","guid":{"rendered":"http:\/\/www.neb-online.de\/en\/genome-editing\/herstellung-von-homologous-repair-templates\/"},"modified":"2023-12-19T14:37:26","modified_gmt":"2023-12-19T13:37:26","slug":"designing-homologous-repair-templates","status":"publish","type":"page","link":"https:\/\/www.neb-online.de\/en\/genome-editing\/designing-homologous-repair-templates\/","title":{"rendered":"Designing Homologous Repair Templates"},"content":{"rendered":"

[vc_row][vc_column][vc_basic_grid post_type=”post” max_items=”3″ element_width=”12″ item=”4123″ grid_id=”vc_gid:1702992988176-57b35e10-06a3-3″ taxonomies=”145, 69, 70″][\/vc_column][\/vc_row][vc_row][vc_column width=”2\/3″][vc_column_text]Modifications of Cas9-target sites can be achieved by supplying a homologous repair template in addition to sgRNA and Cas9. Homologous repair templates are plasmids containing 2-3 kb of homology surrounding the target sequence that are then altered to have the desired mutations or \u201cknock-ins\u201d such as selectable markers or fluorescent tags.<\/p>\n

For initial amplification of the region of homology, the homologous region can be isolated by PCR amplification from genomic DNA with a high-fidelity polymerase and then cloned into your plasmid backbone.<\/p>\n

NEB suggests the use Q5 High Fidelity DNA Polymerase<\/a> products for this amplification followed by the use of the PCR Cloning Kit (NEB #E1202)<\/a> to clone this product into a plasmid. Further modifications can then be introduced using NEBuilder HiFi DNA Assembly Master Mix (NEB #E2621)<\/a> or by site-directed mutagenesis using the Q5 Site-Directed Mutagenesis Kit (NEB #E0554)<\/a>.[\/vc_column_text][\/vc_column][vc_column width=”1\/3″][vc_single_image image=”4798″ img_size=”full” onclick=”link_image”][\/vc_column][\/vc_row][vc_row bwd_show_when_type=”before” bwd_show_when_date=”12\/31\/2023 23:00″ bwd_tbc4vd_show_weekday=””][vc_column][vc_separator][vc_column_text]<\/p>\n

Product table<\/h2>\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n
PRODUCT<\/th>\nProd. #<\/th>\nSIZE<\/th>\nPRICE<\/th>\n<\/tr>\n
Q5\u00ae<\/sup> High-Fidelity DNA Polymerase<\/td>\nM0491S<\/td>\n100 units<\/td>\n108 \u20ac<\/strong><\/td>\n\"In<\/a><\/td>\n\"Produktinformationen\"<\/a><\/td>\n<\/tr>\n
M0491L<\/td>\n500 units<\/td>\n474 \u20ac<\/strong><\/td>\n\"In<\/a><\/td>\n\"Produktinformationen\"<\/a><\/td>\n<\/tr>\n
Q5\u00ae<\/sup> Hot Start High-Fidelity DNA Polymerase<\/td>\nM0493S<\/td>\n100 units<\/td>\n126 \u20ac<\/strong><\/td>\n\"In<\/a><\/td>\n\"Produktinformationen\"<\/a><\/td>\n<\/tr>\n
M0493L<\/td>\n500 units<\/td>\n570 \u20ac<\/strong><\/td>\n\"In<\/a><\/td>\n\"Produktinformationen\"<\/a><\/td>\n<\/tr>\n
NEB PCR Cloning Kit<\/td>\nE1202S<\/td>\n20 rxns<\/td>\n318 \u20ac<\/strong><\/td>\n\"In<\/a><\/td>\n\"Produktinformationen\"<\/a><\/td>\n<\/tr>\n
NEB PCR Cloning Kit (Without Competent Cells)<\/td>\nE1203S<\/td>\n20 rxns<\/td>\n166 \u20ac<\/strong><\/td>\n\"In<\/a><\/td>\n\"Produktinformationen\"<\/a><\/td>\n<\/tr>\n
Q5\u00ae<\/sup> Site-Directed Mutagenesis Kit<\/td>\nE0554S<\/td>\n10 rxns<\/td>\n218 \u20ac<\/strong><\/td>\n\"In<\/a><\/td>\n\"Produktinformationen\"<\/a><\/td>\n<\/tr>\n
Q5\u00ae<\/sup> Site-Directed Mutagenesis Kit (Without Competent Cells)<\/td>\nE0552S<\/td>\n10 rxns<\/td>\n149 \u20ac<\/strong><\/td>\n\"In<\/a><\/td>\n\"Produktinformationen\"<\/a><\/td>\n<\/tr>\n
NEBuilder\u00ae<\/sup> HiFi DNA Assembly Master Mix<\/td>\nE2621S<\/td>\n10 rxns<\/td>\n172 \u20ac<\/strong><\/td>\n\"In<\/a><\/td>\n\"Produktinformationen\"<\/a><\/td>\n<\/tr>\n
E2621L<\/td>\n50 rxns<\/td>\n688 \u20ac<\/strong><\/td>\n\"In<\/a><\/td>\n\"Produktinformationen\"<\/a><\/td>\n<\/tr>\n
E2621X<\/td>\n250 rxns<\/td>\n2.752 \u20ac<\/strong><\/td>\n\"In<\/a><\/td>\n\"Produktinformationen\"<\/a><\/td>\n<\/tr>\n
NEBuilder\u00ae<\/sup> HiFi DNA Assembly Cloning Kit<\/td>\nE5520S<\/td>\n10 rxns<\/td>\n204 \u20ac<\/strong><\/td>\n\"In<\/a><\/td>\n\"Produktinformationen\"<\/a><\/td>\n<\/tr>\n<\/tbody>\n<\/table>\n

As of: 01.01.2023<\/span><\/em><\/p>\n

[\/vc_column_text][\/vc_column][\/vc_row][vc_row bwd_show_when_type=”after” bwd_show_when_date=”12\/31\/2023 23:00″ bwd_tbc4vd_show_weekday=””][vc_column][vc_separator][vc_column_text]<\/p>\n

Product table<\/h2>\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n
PRODUCT<\/th>\nProd. #<\/th>\nSIZE<\/th>\nINFO<\/th>\n<\/tr>\n
Q5\u00ae<\/sup> High-Fidelity DNA Polymerase<\/td>\nM0491S<\/td>\n100 units<\/td>\n\"In<\/a><\/td>\n\"Produktinformationen\"<\/a><\/td>\n<\/tr>\n
M0491L<\/td>\n500 units<\/td>\n\"In<\/a><\/td>\n\"Produktinformationen\"<\/a><\/td>\n<\/tr>\n
Q5\u00ae<\/sup> Hot Start High-Fidelity DNA Polymerase<\/td>\nM0493S<\/td>\n100 units<\/td>\n\"In<\/a><\/td>\n\"Produktinformationen\"<\/a><\/td>\n<\/tr>\n
M0493L<\/td>\n500 units<\/td>\n\"In<\/a><\/td>\n\"Produktinformationen\"<\/a><\/td>\n<\/tr>\n
NEB PCR Cloning Kit<\/td>\nE1202S<\/td>\n20 rxns<\/td>\n\"In<\/a><\/td>\n\"Produktinformationen\"<\/a><\/td>\n<\/tr>\n
NEB PCR Cloning Kit (Without Competent Cells)<\/td>\nE1203S<\/td>\n20 rxns<\/td>\n\"In<\/a><\/td>\n\"Produktinformationen\"<\/a><\/td>\n<\/tr>\n
Q5\u00ae<\/sup> Site-Directed Mutagenesis Kit<\/td>\nE0554S<\/td>\n10 rxns<\/td>\n\"In<\/a><\/td>\n\"Produktinformationen\"<\/a><\/td>\n<\/tr>\n
Q5\u00ae<\/sup> Site-Directed Mutagenesis Kit (Without Competent Cells)<\/td>\nE0552S<\/td>\n10 rxns<\/td>\n\"In<\/a><\/td>\n\"Produktinformationen\"<\/a><\/td>\n<\/tr>\n
NEBuilder\u00ae<\/sup> HiFi DNA Assembly Master Mix<\/td>\nE2621S<\/td>\n10 rxns<\/td>\n\"In<\/a><\/td>\n\"Produktinformationen\"<\/a><\/td>\n<\/tr>\n
E2621L<\/td>\n50 rxns<\/td>\n\"In<\/a><\/td>\n\"Produktinformationen\"<\/a><\/td>\n<\/tr>\n
E2621X<\/td>\n250 rxns<\/td>\n\"In<\/a><\/td>\n\"Produktinformationen\"<\/a><\/td>\n<\/tr>\n
NEBuilder\u00ae<\/sup> HiFi DNA Assembly Cloning Kit<\/td>\nE5520S<\/td>\n10 rxns<\/td>\n\"In<\/a><\/td>\n\"Produktinformationen\"<\/a><\/td>\n<\/tr>\n<\/tbody>\n<\/table>\n

As of: 01.01.2024<\/span><\/em><\/p>\n

[\/vc_column_text][\/vc_column][\/vc_row]<\/p>\n<\/div>","protected":false},"excerpt":{"rendered":"

[vc_row][vc_column][vc_basic_grid post_type=”post” max_items=”3″ element_width=”12″ item=”4123″ grid_id=”vc_gid:1702992988176-57b35e10-06a3-3″ taxonomies=”145, 69, 70″][\/vc_column][\/vc_row][vc_row][vc_column width=”2\/3″][vc_column_text]Modifications of Cas9-target sites can be achieved by supplying a homologous repair template in addition to sgRNA and Cas9. Homologous repair templates are plasmids containing 2-3 kb of homology surrounding the target sequence that are then altered to have the desired mutations or \u201cknock-ins\u201d such as… Read more »<\/a><\/p>\n","protected":false},"author":18,"featured_media":41240,"parent":2072,"menu_order":0,"comment_status":"closed","ping_status":"closed","template":"page-sidebar.php","meta":{"footnotes":""},"class_list":["post-13992","page","type-page","status-publish","has-post-thumbnail","hentry"],"yoast_head":"\nDesigning Homologous Repair Templates - New England Biolabs GmbH<\/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.neb-online.de\/en\/genome-editing\/designing-homologous-repair-templates\/\" \/>\n<meta property=\"og:locale\" content=\"en_US\" \/>\n<meta property=\"og:type\" content=\"article\" \/>\n<meta property=\"og:title\" content=\"Designing Homologous Repair Templates - New England Biolabs GmbH\" \/>\n<meta property=\"og:description\" content=\"[vc_row][vc_column][vc_basic_grid post_type=”post” max_items=”3″ element_width=”12″ item=”4123″ grid_id=”vc_gid:1702992988176-57b35e10-06a3-3″ taxonomies=”145, 69, 70″][\/vc_column][\/vc_row][vc_row][vc_column width=”2\/3″][vc_column_text]Modifications of Cas9-target sites can be achieved by supplying a homologous repair template in addition to sgRNA and Cas9. Homologous repair templates are plasmids containing 2-3 kb of homology surrounding the target sequence that are then altered to have the desired mutations or \u201cknock-ins\u201d such as... 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