{"id":9814,"date":"2025-04-13T03:27:43","date_gmt":"2025-04-12T19:27:43","guid":{"rendered":"https:\/\/fanyiming.life\/?p=9814"},"modified":"2025-12-21T01:03:47","modified_gmt":"2025-12-20T17:03:47","slug":"crispr-screen%e7%bb%86%e8%8a%82","status":"publish","type":"post","link":"https:\/\/fanyiming.life\/?p=9814","title":{"rendered":"CRISPR Screen\u7ec6\u8282"},"content":{"rendered":"\n
\n

\u672c\u6587\u7531Gemini 2.5 Pro\u6839\u636e\u6211\u7684\u8349\u7a3f\u586b\u5145\u5b8c\u6210\uff0c\u6d89\u53ca\u7684\u516c\u5f0f\u63a8\u5bfc\u6ca1\u6709\u5199\u5168\uff0c\u9700\u8981\u7684\u8bfb\u8005\u53ef\u4ee5\u590d\u5236\u540e\u53bb\u95eeAI\u3002<\/p>\n<\/blockquote>\n\n\n\n

\n\n\n\n

CRISPR\u7b5b\u9009\u662f\u529f\u80fd\u57fa\u56e0\u7ec4\u5b66\u7814\u7a76\u7684\u5229\u5668\uff0c\u4f46\u5176\u6210\u529f\u4f9d\u8d56\u4e8e\u5bf9\u5b9e\u9a8c\u7ec6\u8282\u7684\u7cbe\u51c6\u628a\u63a7\u3002\u672c\u6587\u805a\u7126\u4e09\u4e2a\u5173\u952e\u73af\u8282\uff1asgRNA\u6587\u5e93\u5747\u5300\u6027<\/strong>\u3001\u7ec6\u80de\u8d77\u59cb\u91cf\u4f18\u5316\uff08MOI\u4e0e\u8986\u76d6\u5ea6\u5e73\u8861\uff09<\/strong>\u53ca\u7ec6\u80de\u4f20\u4ee3\u74f6\u9888\u7ba1\u7406<\/strong>\uff0c\u5206\u4eab\u6838\u5fc3\u8981\u70b9\u4e0e\u5b9e\u8df5\u5efa\u8bae\uff0c\u52a9\u60a8\u83b7\u5f97\u53ef\u9760\u7684\u7b5b\u9009\u7ed3\u679c\u3002<\/p>\n\n\n\n

\u4e00\u3001 \u4fdd\u8bc1\u8d77\u70b9\u516c\u5e73\uff1asgRNA\u8d28\u7c92\u6587\u5e93\u7684\u5747\u5300\u6027<\/strong><\/h2>\n\n\n\n

\u7406\u60f3\u7684CRISPR\u7b5b\u9009\u59cb\u4e8e\u4e00\u4e2a\u5404\u79cdsgRNA\u8d28\u7c92\u62f7\u8d1d\u6570\u5747\u7b49\u7684\u6587\u5e93\u3002<\/p>\n\n\n\n

    \n
  • \u9a8c\u8bc1\u5148\u884c\uff1a<\/strong> \u5fc5\u987b\u901a\u8fc7NGS\u6d4b\u5e8f<\/strong>\u4e25\u683c\u9a8c\u8bc1\u521d\u59cb\u6587\u5e93\u7684\u5747\u5300\u6027\u3002<\/li>\n\n\n\n
  • \u6269\u589e\u6311\u6218\u4e0e\u5bf9\u7b56\uff1a<\/strong> \u6587\u5e93\u6269\u589e\u6613\u5f15\u5165\u74f6\u9888\u6548\u5e94<\/strong>\uff0c\u5bfc\u81f4sgRNA\u4e30\u5ea6\u504f\u659c\u3002\u4e3a\u7ef4\u6301\u5747\u5300\u6027\uff1a\n
      \n
    • \u8d85\u91cf\u8f6c\u5316\uff1a<\/strong> \u91c7\u7528\u9ad8\u6548\u7387\u8f6c\u5316\uff08\u5982\u7535\u8f6c\uff09\uff0c\u786e\u4fdd\u8f6c\u5316\u5b50\u603b\u6570\u8fdc\u8d85sgRNA\u79cd\u7c7b\u6570\uff08>100\u500d\u8986\u76d6\u5ea6\uff09\u3002<\/li>\n\n\n\n
    • \u5408\u5e76\u57f9\u517b\uff1a<\/strong> \u5173\u952e\uff1a\u907f\u514d\u6311\u5355\u514b\u9686\u3002<\/strong> \u5e94\u5408\u5e76\u6240\u6709\u8f6c\u5316\u5b50\uff08\u522e\u677f\u6216\u6db2\u4f53\u8f6c\u5316\u540e\u76f4\u63a5\u57f9\u517b\uff09\u8fdb\u884c\u5927\u89c4\u6a21\u57f9\u517b\u3002<\/li>\n\n\n\n
    • \u5747\u5300\u62bd\u63d0\uff1a<\/strong> \u62bd\u63d0\u524d\u5145\u5206\u6df7\u5300\u83cc\u6db2\uff0c\u786e\u4fdd\u8d28\u7c92\u6837\u672c\u4ee3\u8868\u6027\u3002<\/li>\n<\/ul>\n<\/li>\n\n\n\n
    • \u518d\u6b21\u9a8c\u8bc1\uff1a<\/strong> \u6269\u589e\u540e\u5fc5\u987b\u518d\u6b21NGS\u6d4b\u5e8f<\/strong>\u8bc4\u4f30\u5747\u5300\u6027\u3002<\/li>\n\n\n\n
    • \u8861\u91cf\u6307\u6807\uff1a<\/strong>\n
        \n
      • \u504f\u659c\u5ea6 (Skew Ratio)\uff1a<\/strong> \u6700\u9ad81\/4 sgRNA reads\u603b\u6570 \/ \u6700\u4f4e1\/4 sgRNA reads\u603b\u6570\u3002\u6bd4\u503c\u8d8a\u63a5\u8fd11\u8d8a\u597d\uff08\u76ee\u6807<10\uff0c\u7406\u60f3<5\uff09\u3002<\/li>\n\n\n\n
      • \u5b8c\u6574\u6027 (Representation)\uff1a<\/strong> \u786e\u8ba4\u6240\u6709\u8bbe\u8ba1\u7684sgRNA\u5747\u88ab\u68c0\u51fa\uff08reads > 0\uff09\u3002<\/li>\n<\/ul>\n<\/li>\n<\/ul>\n\n\n\n

        \u4e8c\u3001 \u7cbe\u786e\u8ba1\u7b97\u7ec6\u80de\u91cf\uff1a\u5e73\u8861\u4f4eMOI\u4e0e\u9ad8\u8986\u76d6\u5ea6<\/strong><\/h2>\n\n\n\n

        \u6587\u5e93\u5bfc\u5165\u7ec6\u80de\u9700\u5e73\u8861\u4e24\u5927\u539f\u5219\uff1a<\/p>\n\n\n\n

          \n
        • \u539f\u5219\u4e00\uff1a\u4f4eMOI (Multiplicity of Infection)<\/strong>\n
            \n
          • \u76ee\u6807\uff1a<\/strong> \u6700\u5927\u5316\u5355sgRNA\u6574\u5408\u7ec6\u80de\u6bd4\u4f8b\uff0c\u51cf\u5c11\u591a\u91cd\u611f\u67d3\u5e72\u6270\u3002<\/li>\n\n\n\n
          • \u8bc4\u4f30 (\u57fa\u4e8e\u6cca\u677e\u5206\u5e03, \u03bb=MOI):<\/strong> \u5173\u952e\u770b\u591a\u91cd\u611f\u67d3\u7ec6\u80de\u5728\u6240\u6709\u88ab\u611f\u67d3\u7ec6\u80de<\/strong>\u4e2d\u7684\u5360\u6bd4 [P(k\u22652) \/ P(k\u22651)]\u3002\n
              \n
            • MOI = 0.3:<\/strong> P(k\u22651) \u2248 25.9%, P(k\u22652) \u2248 3.7%. \u6bd4\u4f8b \u2248 14.3%<\/strong>\u3002<\/li>\n\n\n\n
            • MOI = 0.1:<\/strong> P(k\u22651) \u2248 9.5%, P(k\u22652) \u2248 0.45%. \u6bd4\u4f8b \u2248 4.7%<\/strong>\u3002<\/li>\n<\/ul>\n<\/li>\n\n\n\n
            • \u5efa\u8bae\uff1a<\/strong> MOI\u63a7\u5236\u57280.1-0.3\uff0c\u663e\u8457\u964d\u4f4e\u6709\u6548\u7b5b\u9009\u7fa4\u4f53\u4e2d\u7684\u591a\u91cd\u611f\u67d3\u6bd4\u4f8b\u3002<\/li>\n<\/ul>\n<\/li>\n\n\n\n
            • \u539f\u5219\u4e8c\uff1a\u9ad8sgRNA\u8986\u76d6\u5ea6 (High sgRNA Coverage)<\/strong>\n
                \n
              • \u76ee\u6807\uff1a<\/strong> \u786e\u4fdd\u6bcf\u6761sgRNA\u6709\u8db3\u591f\u7ec6\u80de\u4ee3\u8868\uff0c\u62b5\u6297\u7b5b\u9009\u8fc7\u7a0b\u4e2d\u7684\u968f\u673a\u4e22\u5931\u3002<\/li>\n\n\n\n
              • \u5efa\u8bae\uff1a<\/strong> \u8d77\u59cb\u8986\u76d6\u5ea6\u81f3\u5c11300-500x\uff0c\u751a\u81f3\u66f4\u9ad8\u3002<\/li>\n<\/ul>\n<\/li>\n\n\n\n
              • \u8ba1\u7b97\u521d\u59cb\u7ec6\u80de\u91cf (N):<\/strong>\n
                  \n
                • N = (sgRNA\u79cd\u7c7b\u6570 S) * (\u671f\u671b\u8986\u76d6\u5ea6 C) \/ (\u6709\u6548\u611f\u67d3\u7387 E)<\/code><\/li>\n\n\n\n
                • \u4f4eMOI\u4e0b\uff0cE\u53ef\u8fd1\u4f3c\u4e3aMOI\u3002<\/em><\/li>\n\n\n\n
                • \u793a\u4f8b:<\/strong> 8\u4e07sgRNA\uff0c500x\u8986\u76d6\u5ea6\uff0cMOI=0.1 \u2192 N = 80k * 500 \/ 0.1 = 4\u4ebf<\/strong> \u6700\u4f4e\u8d77\u59cb\u7ec6\u80de\u6570\u3002<\/li>\n<\/ul>\n<\/li>\n<\/ul>\n\n\n\n

                  \u4e09\u3001 \u7cbe\u51c6\u75c5\u6bd2\u6ef4\u5ea6\uff1aMOI\u6ef4\u5b9a\u4e0e\u5e94\u7528<\/strong><\/h2>\n\n\n\n

                  \u75c5\u6bd2\u611f\u67d3\u6548\u7387\u591a\u53d8\uff0c\u7cbe\u786e\u6ef4\u5b9a\u5fc5\u4e0d\u53ef\u5c11\u3002<\/p>\n\n\n\n

                    \n
                  • \u6ef4\u5b9a\u5fc5\u8981\u6027\uff1a<\/strong> \u4e0d\u540c\u75c5\u6bd2\u6279\u6b21\u3001\u50a8\u5b58\u6761\u4ef6\u3001\u7ec6\u80de\u72b6\u6001\u5747\u5f71\u54cd\u6548\u7387\uff0c\u52a1\u5fc5\u5728\u4f7f\u7528\u524d\u5bf9\u7279\u5b9a\u7ec6\u80de\u7cfb\u8fdb\u884cMOI\u6ef4\u5b9a<\/strong>\u3002<\/li>\n\n\n\n
                  • MOI\u4e0e\u5b58\u6d3b\u7387\u5173\u7cfb:<\/strong> \u7b5b\u9009\u540e\u5b58\u6d3b\u7ec6\u80de\u6bd4\u4f8b(Y)\u4e0eMOI\u57fa\u4e8e\u6cca\u677e\u5206\u5e03\u5173\u8054\uff1aY = 1 – e^-MOI<\/strong>\u3002\u7531\u4e8eMOI\u4e0e\u75c5\u6bd2\u4f53\u79ef(X)\u6210\u6b63\u6bd4 (MOI=kX)\uff0c\u6545 Y = 1 – e^(-kX)<\/strong>\u3002<\/li>\n\n\n\n
                  • \u64cd\u4f5c\uff1a<\/strong> \u901a\u8fc7\u5b9e\u9a8c\u6d4b\u5b9a\u4e00\u7cfb\u5217X\u5bf9\u5e94\u7684Y\u503c\uff0c\u7ed8\u5236\u66f2\u7ebf\uff0c\u786e\u5b9a\u8fbe\u5230\u76ee\u6807MOI\uff08\u5982MOI=0.1\u5bf9\u5e94Y\u22489.5%\uff09\u6240\u9700\u7684\u75c5\u6bd2\u4f53\u79efX\u3002<\/li>\n<\/ul>\n\n\n\n

                    \u56db\u3001 \u7ec6\u80de\u4f20\u4ee3\u7684\u74f6\u9888\uff1a\u7ef4\u6301\u8986\u76d6\u5ea6\u7684\u6311\u6218<\/strong><\/h2>\n\n\n\n

                    \u5927\u89c4\u6a21\u7ec6\u80de\u57f9\u517b\u9700\u5f03\u53bb\u90e8\u5206\u7ec6\u80de\uff0c\u6b64\u8fc7\u7a0b\u5373\u4e3a\u74f6\u9888<\/strong>\uff0c\u6613\u4e22\u5931\u4f4e\u4e30\u5ea6sgRNA\u3002<\/p>\n\n\n\n

                      \n
                    • \u98ce\u9669\u91cf\u5316 (CV\u793a\u4f8b):<\/strong> \u5047\u8bbe\u4f20\u4ee3\u4fdd\u755950%\u7ec6\u80de\uff0c\u5206\u6790\u4ee3\u8868\u6570\u6ce2\u52a8\uff08Coefficient of Variation, CV = SD\/Mean\uff09\u3002\n
                        \n
                      • sgRNA\u4ee3\u8868\u6570=100:<\/strong> \u671f\u671b\u4fdd\u755950, SD\u22485, CV\u224810%<\/strong>\u3002<\/li>\n\n\n\n
                      • sgRNA\u4ee3\u8868\u6570=10:<\/strong> \u671f\u671b\u4fdd\u75595, SD\u22481.58, CV\u224831.6%<\/strong>\u3002<\/li>\n<\/ul>\n<\/li>\n\n\n\n
                      • \u7ed3\u8bba\uff1a<\/strong> \u4f4e\u4e30\u5ea6sgRNA\u7684\u76f8\u5bf9\u62bd\u6837\u8bef\u5dee(CV)\u6025\u5267\u589e\u5927\uff0c\u4e22\u5931\u98ce\u9669\u6781\u9ad8\u3002\u5173\u952e\u5728\u4e8e\u6bcf\u6b21\u4f20\u4ee3\u4fdd\u7559\u8db3\u591f\u7ec6\u80de\u6570\uff0c\u59cb\u7ec8\u7ef4\u6301\u5fc5\u8981\u7684sgRNA\u8986\u76d6\u5ea6<\/strong>\uff08\u4f8b\u5982\uff0c\u7ef4\u6301>300x\uff09\u3002<\/li>\n<\/ul>\n\n\n\n

                        \u7ed3\u8bed<\/strong><\/p>\n\n\n\n

                        CRISPR\u7b5b\u9009\u7684\u53ef\u9760\u6027\u6e90\u4e8e\u5bf9\u7ec6\u8282\u7684\u4e25\u8c28\u63a7\u5236\u3002\u7cbe\u7ec6\u7ba1\u7406\u6587\u5e93\u5747\u5300\u6027<\/strong>\u3001\u7ec6\u80de\u6570\u4e0eMOI\/\u8986\u76d6\u5ea6<\/strong>\u3001\u4ee5\u53ca\u7ec6\u80de\u4f20\u4ee3\u8fc7\u7a0b<\/strong>\uff0c\u662f\u83b7\u53d6\u9ad8\u8d28\u91cf\u3001\u53ef\u91cd\u590d\u6570\u636e\u7684\u57fa\u77f3\u3002<\/p>\n","protected":false},"excerpt":{"rendered":"

                        \u8fd9\u7bc7\u7b14\u8bb0\u805a\u7126CRISPR\u7b5b\u9009\u5b9e\u9a8c\u4e2d\u7684\u5173\u952e\u7ec6\u8282\uff0c\u65e8\u5728\u63d0\u5347\u7ed3\u679c\u7684\u53ef\u9760\u6027\u3002\u6587\u7ae0\u7cfb\u7edf\u9610\u8ff0\u4e86sgRNA\u6587\u5e93\u5747\u5300\u6027\u9a8c\u8bc1\u3001\u7ec6\u80de\u8d77\u59cb\u91cf\u4e0eMOI\/\u8986\u76d6\u5ea6\u7684\u7cbe\u786e\u8ba1\u7b97\u3001\u75c5\u6bd2\u6ef4\u5ea6\u6ef4\u5b9a\u4ee5\u53ca\u7ec6\u80de\u4f20\u4ee3\u8fc7\u7a0b\u4e2d\u7684\u74f6\u9888\u7ba1\u7406\u3002<\/p>\n","protected":false},"author":1,"featured_media":0,"comment_status":"open","ping_status":"closed","sticky":false,"template":"","format":"standard","meta":{"footnotes":""},"categories":[4],"tags":[],"class_list":["post-9814","post","type-post","status-publish","format-standard","hentry","category-note"],"_links":{"self":[{"href":"https:\/\/fanyiming.life\/index.php?rest_route=\/wp\/v2\/posts\/9814","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=9814"}],"version-history":[{"count":2,"href":"https:\/\/fanyiming.life\/index.php?rest_route=\/wp\/v2\/posts\/9814\/revisions"}],"predecessor-version":[{"id":13858,"href":"https:\/\/fanyiming.life\/index.php?rest_route=\/wp\/v2\/posts\/9814\/revisions\/13858"}],"wp:attachment":[{"href":"https:\/\/fanyiming.life\/index.php?rest_route=%2Fwp%2Fv2%2Fmedia&parent=9814"}],"wp:term":[{"taxonomy":"category","embeddable":true,"href":"https:\/\/fanyiming.life\/index.php?rest_route=%2Fwp%2Fv2%2Fcategories&post=9814"},{"taxonomy":"post_tag","embeddable":true,"href":"https:\/\/fanyiming.life\/index.php?rest_route=%2Fwp%2Fv2%2Ftags&post=9814"}],"curies":[{"name":"wp","href":"https:\/\/api.w.org\/{rel}","templated":true}]}}