{"id":145,"date":"2023-02-16T01:52:32","date_gmt":"2023-02-15T17:52:32","guid":{"rendered":"http:\/\/39.105.184.3\/?p=145"},"modified":"2025-12-21T01:06:34","modified_gmt":"2025-12-20T17:06:34","slug":"%e5%9f%ba%e4%ba%8ecrispr-cas9%e5%92%8ccre-loxp%e7%9a%84%e6%88%90%e5%b9%b4%e5%b0%8f%e9%bc%a0%e5%9f%ba%e5%9b%a0%e6%95%b2%e9%99%a4","status":"publish","type":"post","link":"https:\/\/fanyiming.life\/?p=145","title":{"rendered":"\u57fa\u4e8eCRISPR\/Cas9\u548cCre-LoxP\u7684\u6210\u5e74\u5c0f\u9f20\u57fa\u56e0\u6572\u9664"},"content":{"rendered":"\n<p>\u5bf9\u5c0f\u9f20\u8fdb\u884c\u57fa\u56e0\u6572\u9664\u662f\u7814\u7a76\u76ee\u7684\u57fa\u56e0\u529f\u80fd\u7684\u5e38\u7528\u65b9\u6cd5\u3002\u4f20\u7edf\u7684\u3001\u4ece\u53d7\u7cbe\u5375\u6216\u80da\u80ce\u65f6\u671f\u8fdb\u884c\u57fa\u56e0\u64cd\u4f5c\uff0c\u6765\u83b7\u5f97\u57fa\u56e0\u6572\u9664\u5c0f\u9f20\u7684\u65b9\u6cd5\u5f80\u5f80\u8017\u65f6\u957f\u8fbe\u51e0\u5e74\u3002AAV\uff08\u817a\u76f8\u5173\u75c5\u6bd2\uff09\u9012\u9001\u7cfb\u7edf\u7ed3\u5408CRISPR\/Cas9\u53ef\u4ee5\u5b9e\u73b0\u5bf9\u6210\u5e74\u5c0f\u9f20\u7684\u57fa\u56e0\u6572\u9664\uff0c\u4ece\u800c\u5927\u5927\u7f29\u77ed\u83b7\u5f97\u57fa\u56e0\u6572\u9664\u5c0f\u9f20\u7684\u65f6\u95f4\u3002AAV\u8fd8\u5177\u6709\u4f4e\u514d\u75ab\u539f\u6027\u3001\u4e0d\u540c\u8840\u6e05\u578b\u5b9e\u73b0\u9776\u5411\u4e0d\u540c\u7ec4\u7ec7\u6216\u5668\u5b98\u7684\u4f18\u70b9\u3002<\/p>\n\n\n\n<p>\u4e0e\u6b64\u540c\u65f6\uff0cAAV\u9012\u9001\u7cfb\u7edf\u6709\u4e00\u4e2a\u56fa\u6709\u7f3a\u9677\uff0c\u5176\u8f7d\u4f53\u6240\u80fd\u5bb9\u7eb3\u7684\u63d2\u5165\u5e8f\u5217\u6700\u5927\u53ea\u67094.7Kb\u3002\u4f7f\u7528CRISPR\/Cas9\u8fdb\u884c\u57fa\u56e0\u6572\u9664\uff0c\u9700\u8981\u5c06\u542b\u6709Cas9\u7f16\u7801\u5e8f\u5217\u548csgRNA\u7684\u8f7d\u4f53\u8fdb\u884c\u9012\u9001\u3002\u7136\u800c\uff0c<em>Streptococcus pyogenes<\/em> (SpCas9) \u548c gRNA together\u5c31\u5360\u4e864.2kb\uff0c\u5bf9AAV\u7684\u5bb9\u91cf\u4ea7\u751f\u4e86\u6311\u6218\u3002\u56e0\u6b64\uff0c\u5982\u4f55\u5bf9Cas9\u8fdb\u884c\u6709\u6548\u7684\u9012\u9001\u4e00\u76f4\u662fCRISPR\/Cas9\u5728\u52a8\u7269\u4f53\u5185\u53d1\u6325\u4f5c\u7528\u7684\u96be\u70b9\u3002\u8fc7\u53bb\u7684\u65b9\u6cd5\u5305\u62ec\uff1a\u5728\u4e0d\u540c\u7269\u79cd\u4e2d\u5bfb\u627e\u5e8f\u5217\u66f4\u77ed\u66f4\u6709\u6548\u7684Cas9\u3001\u5c06Cas9\u5206\u6bb5\u8868\u8fbe\u5728\u4e24\u4e2aAAV\u8f7d\u4f53\u7b49\u3002<\/p>\n\n\n\n<p>\u5317\u5927\u738b\u6653\u4f1f\u8bfe\u9898\u7ec4\u4e8e2021\u5e749\u6708\u5728STAR Protocols\u6742\u5fd7\u53d1\u8868\u4e86\u4e00\u7bc7\u6587\u7ae0\u8bb0\u5f55\u4e00\u79cd\u7ed3\u5408\u4e86Cre-LoxP\u548cCRISPR\/Cas9\u3001\u7528AAV\u9012\u9001\uff0c\u5b9e\u73b0\u5bf9\u6210\u5e74\u5c0f\u9f20\u8fdb\u884c\u7ec4\u7ec7\u7279\u5f02\u6027\u57fa\u56e0\u6572\u9664\u7684\u65b9\u6cd5\u3002<\/p>\n\n\n\n<p>\u539f\u7406\u5927\u81f4\u4e3a\uff1a<\/p>\n\n\n\n<figure class=\"wp-block-image\"><img loading=\"lazy\" decoding=\"async\" decoding=\"async\" src=\"https:\/\/eamonimgbed.oss-cn-beijing.aliyuncs.com\/img\/202302162134381.png?x-oss-process=style\/screenshots\" alt=\"\" \/><\/figure>\n\n\n\n<p>\u5c06panel A\u6240\u793a\u7684\u7247\u6bb5\u6574\u5408\u5165\u91ce\u751f\u5c0f\u9f20\uff0cpanel B\u6240\u793a\u7247\u6bb5\u7528AAV\u9012\u9001\u3002\u5bf9\u4e8e\u4e0d\u540c\u7684\u9776\u57fa\u56e0\u6765\u8bf4\uff0c\u5de6\u8fb9\u7684\u5c0f\u9f20\u90fd\u662f\u7528\u7684\u540c\u4e00\u79cd\uff0c\u4f46\u662fB\u4e2d\u7684\u5e8f\u5217\u53ef\u4ee5\u6839\u636e\u9700\u8981\u81ea\u5b9a\u4e49\u3002\u4ee5\u56fe\u793a\u4e3a\u4f8b\uff0cTBG\u662f\u809d\u810f\u7279\u5f02\u6027\u542f\u52a8\u5b50\uff0cAAV\u611f\u67d3\u5c0f\u9f20\u540e\uff0cCre\u9176\u53ea\u5728\u809d\u810f\u4e2d\u8868\u8fbe\uff0c\u5728\u5c0f\u9f20\u7684\u809d\u810f\u4e2d\u5220\u53bb\u4e24\u4e2aloxp\u5305\u88f9\u7684stop signal\uff0cspCas9\u5f97\u4ee5\u8868\u8fbe\uff0c\u4e0eCre\u4e00\u540c\u8f6c\u5bfc\u5165\u52a8\u7269\u7684\u8fd8\u6709sgRNA\uff0c\u4e8e\u662f\u5b9e\u73b0\u5728\u809d\u810f\u4e2d\u5bf9\u8be5sgRNA\u9776\u5411\u7684\u7247\u6bb5\u8fdb\u884c\u5207\u5272\u3002spCas9\u540e\u9762\u8fd8\u63a5\u4e86GFP\u65b9\u4fbf\u5b9e\u9a8c\u8005\u89c2\u5bdfCre\u9176\u662f\u5426\u6210\u529f\u53d1\u6325\u4f5c\u7528\u3002<\/p>\n\n\n\n<hr class=\"wp-block-separator has-alpha-channel-opacity\" \/>\n\n\n\n<p>\u8be5\u65b9\u6cd5\u7684\u4f18\u70b9\u6709\uff1a<\/p>\n\n\n\n<ol class=\"wp-block-list\">\n<li>\u628aCas9\u4ece\u8f7d\u4f53\u4e0a\u62ef\u6551\u4e0b\u6765\uff0c\u76f8\u5f53\u4e8e\u7ed9AAV\u6269\u5bb9\u4e86\uff0c\u53ef\u4ee5\u52a0\u66f4\u957f\u7684\u7ec4\u7ec7\u7279\u5f02\u542f\u52a8\u5b50\u6216\u5176\u4ed6\u6709\u7279\u6b8a\u76ee\u7684\u7684\u5143\u4ef6\u3002<\/li>\n\n\n\n<li>GFP\u5bf9Cre\u9176\u7684\u6d3b\u6027\u8fdb\u884c\u4e86\u53ef\u89c6\u5316\u3002<\/li>\n\n\n\n<li>\u6709\u4e2a\u8fd9\u6837\u7684\u5c0f\u9f20\u4e5f\u8bb8\u4f1a\u66f4\u65b9\u4fbf\u5427\u3002\uff08\u6bd4\u5982\u76f8\u6bd4\u4e8e\u4f20\u7edf\u65b9\u6cd5\u4e0d\u7528\u4e3a\u6bcf\u4e2a\u57fa\u56e0\u5355\u72ec\u6784\u5efaflox\u5c0f\u9f20\u4e86\uff0c\u5f53\u7136\u56e0\u4e3a\u6ca1\u6709\u540c\u6e90\u91cd\u7ec4\u8fc7\u7a0b\uff0c\u7528\u9014\u4e5f\u5c40\u9650\u4e86\uff09<\/li>\n<\/ol>\n\n\n\n<p>\u539f\u6587\u94fe\u63a5\uff1a<\/p>\n\n\n\n<p><a href=\"https:\/\/pubmed.ncbi.nlm.nih.gov\/34189476\/\">Wang X, Xu BL, Chen XW. Acute gene inactivation in the adult mouse liver using the CRISPR-Cas9 technology. STAR Protoc. 2021 Jun 17;2(3):100611. doi: 10.1016\/j.xpro.2021.100611. PMID: 34189476; PMCID: PMC8220398.<\/a><\/p>\n\n\n\n<p>\u53c2\u8003\u535a\u6587\uff1a<\/p>\n\n\n\n<figure class=\"wp-block-embed\"><div class=\"wp-block-embed__wrapper\">\nhttps:\/\/blog.addgene.org\/a-match-made-in-heaven-crispr\/cas9-and-aav\n<\/div><\/figure>\n\n\n\n<p>\u9e23\u8c22\uff1a\u5c0f\u9f99\u54e5<\/p>\n","protected":false},"excerpt":{"rendered":"<p>\u8fd9\u7bc7\u6587\u732e\u4ecb\u7ecd\u4e86\u4e00\u79cd\u5de7\u5999\u7ed3\u5408CRISPR\/Cas9\u4e0eCre-LoxP\u7cfb\u7edf\uff0c\u5229\u7528AAV\u9012\u9001\u5b9e\u73b0\u6210\u5e74\u5c0f\u9f20\u7ec4\u7ec7\u7279\u5f02\u6027\u57fa\u56e0\u6572\u9664\u7684\u521b\u65b0\u65b9\u6cd5\u3002\u5b83\u6709\u6548\u89e3\u51b3\u4e86AAV\u8f7d\u4f53\u5bb9\u91cf\u9650\u5236\u4e0eSpCas9\u7f16\u7801\u5e8f\u5217\u8fc7\u957f\u7684\u77db\u76fe\uff0c\u4e3a\u57fa\u56e0\u529f\u80fd\u7814\u7a76\u63d0\u4f9b\u4e86\u66f4\u9ad8\u6548\u3001\u66f4\u7075\u6d3b\u7684\u5de5\u5177\u3002<\/p>\n","protected":false},"author":1,"featured_media":0,"comment_status":"open","ping_status":"open","sticky":false,"template":"","format":"standard","meta":{"footnotes":""},"categories":[18],"tags":[20],"class_list":["post-145","post","type-post","status-publish","format-standard","hentry","category-18","tag-crispr"],"_links":{"self":[{"href":"https:\/\/fanyiming.life\/index.php?rest_route=\/wp\/v2\/posts\/145","targetHints":{"allow":["GET"]}}],"collection":[{"href":"https:\/\/fanyiming.life\/index.php?rest_route=\/wp\/v2\/posts"}],"about":[{"href":"https:\/\/fanyiming.life\/index.php?rest_route=\/wp\/v2\/types\/post"}],"author":[{"embeddable":true,"href":"https:\/\/fanyiming.life\/index.php?rest_route=\/wp\/v2\/users\/1"}],"replies":[{"embeddable":true,"href":"https:\/\/fanyiming.life\/index.php?rest_route=%2Fwp%2Fv2%2Fcomments&post=145"}],"version-history":[{"count":2,"href":"https:\/\/fanyiming.life\/index.php?rest_route=\/wp\/v2\/posts\/145\/revisions"}],"predecessor-version":[{"id":14180,"href":"https:\/\/fanyiming.life\/index.php?rest_route=\/wp\/v2\/posts\/145\/revisions\/14180"}],"wp:attachment":[{"href":"https:\/\/fanyiming.life\/index.php?rest_route=%2Fwp%2Fv2%2Fmedia&parent=145"}],"wp:term":[{"taxonomy":"category","embeddable":true,"href":"https:\/\/fanyiming.life\/index.php?rest_route=%2Fwp%2Fv2%2Fcategories&post=145"},{"taxonomy":"post_tag","embeddable":true,"href":"https:\/\/fanyiming.life\/index.php?rest_route=%2Fwp%2Fv2%2Ftags&post=145"}],"curies":[{"name":"wp","href":"https:\/\/api.w.org\/{rel}","templated":true}]}}