{"id":2990,"date":"2024-06-19T21:19:57","date_gmt":"2024-06-19T13:19:57","guid":{"rendered":"https:\/\/opentrons.com.cn\/?post_type=knowledge2&#038;p=2990"},"modified":"2024-09-02T18:57:26","modified_gmt":"2024-09-02T10:57:26","slug":"shiyong-opentrons-dbzchgzzjxjy","status":"publish","type":"post","link":"https:\/\/opentrons.com.cn\/zh-TW\/news\/shiyong-opentrons-dbzchgzzjxjy\/","title":{"rendered":"\u4f7f\u7528 Opentrons \u86cb\u767d\u8cea\u7d14\u5316\u5de5\u4f5c\u7ad9\u9032\u884c\u57fa\u65bc Ni-NTA \u78c1\u73e0\u7684\u86cb\u767d\u8cea\u7d14\u5316"},"content":{"rendered":"<!-- wp:paragraph -->\n<p><strong>\u4f5c\u8005<\/strong><\/p>\n<!-- \/wp:paragraph -->\n\n<!-- wp:paragraph -->\n<p>BO\u4eba &nbsp;&nbsp;Lin,PhD,kin\u90a3\u65e5 &nbsp;&nbsp;Watson,PhD 1open TR ONS&nbsp;&nbsp;lab works,Inc.<\/p>\n<!-- \/wp:paragraph -->\n\n<!-- wp:paragraph -->\n<p><strong>\u6458\u8981<\/strong>\u672c\u6b21\u6e2c\u8a66\u7814\u7a76\u7684\u662f\u5728 Opentrons OT-2 \u81ea\u52d5\u5316\u79fb\u6db2\u5de5\u4f5c\u7ad9\u4e0a\u9032 \u884c\u57fa\u65bc\u78c1\u73e0\u7684\u56fa\u5b9a\u5316\u91d1\u5c6c\u89aa\u548c\u5c64\u6790\u6cd5 (IMAC) \u3002 \u6211\u5011\u4f7f\u7528PierceTM Ni-NTA \u78c1\u6027\u74ca\u8102\u7cd6\u5fae\u73e0(Thermo Fisher Scientific, USA) \u6e2c\u8a66\u4e86\u8a72\u65b9\u6848\uff0c\u7528\u65bc\u5728OT-2 \u4e0a\u63d0\u53d6\u76ee\u6a19\u86cb\u767d(\u5e36\u6709His \u6a19\u7c64\u7684GAPDH), \u4f7f\u7528\u88c2\u89e3\u7de9\u885d\u6db2\u7a00\u91cb\u7684\u91cd\u7d44GAPDH \u86cb\u767d\u3001HEp-2 \u7d30\u80de\u88c2\u89e3\u6db2\u53ca\u83cc\u7d30\u80de\u88fd\u5099\u7684\u6a23\u672c\u3002<\/p>\n<!-- \/wp:paragraph -->\n\n<!-- wp:paragraph -->\n<p><strong>\u95dc\u9375\u767c\u73fe<\/strong>\u5728Opentrons \u86cb\u767d\u8cea\u7d14\u5316\u5de5\u4f5c\u7ad9\u4e0a\u53ef\u4f7f\u7528 Ni-NTA \u78c1\u73e0\u9032\u884c\u91cd\u7d44 GAPDH \u86cb\u767d\u7d14\u5316\u3002 \u25a0 \u5be6\u9a57\u7d50\u679c\u8b49\u660e\u6240\u5f97\u86cb\u767d\u70ba\u76ee\u6a19\u86cb\u767d\uff0c\u5177\u6709\u826f\u597d\u7684\u4e00\u81f4\u6027\u548c \u7279\u7570\u6027\u3002 \u25a0i \u6b64\u65b9\u6848\u53ef\u4ee5\u8655\u7406\u591a\u905496\u500b\u6a23\u54c1\uff0c\u4e26\u6700\u5927\u9650\u5ea6\u5730\u6e1b\u5c11\u9700\u8981\u624b\u52d5\u64cd\u4f5c\u7684\u6642\u9593\u3002<\/p>\n<!-- \/wp:paragraph -->\n\n<!-- wp:paragraph -->\n<p><strong>\u7c21\u4ecb<\/strong>\u86cb\u767d\u8cea\u7d14\u5316\u7684\u76ee\u7684\u662f\u70ba\u4e86\u7372\u5f97\u9ad8\u7d14\u5ea6\u7684\u7a69\u5b9a\u6d3b\u6027\u86cb\u767d\u8cea\uff0c\u4ee5\u4f9b\u4e0b\u6e38\u5206\u6790\u3001\u7814\u7a76\u6216\u6cbb\u7642\u4f7f\u7528\u3002\u56fa\u5b9a\u5316\u91d1\u5c6c\u89aa\u548c\u5c64\u6790\u6cd5 (IMAC)\u5229\u7528\u91d1\u5c6c\u96e2\u5b50\u8403\u53d6\u5177\u6709\u57fa\u56e0\u5de5\u7a0b\u6a19\u7c64\u7684\u91cd\u7d44\u86cb\u767d\uff0c\u80fd\u5920\u87af\u5408\u91d1 \u5c6c\u96e2\u5b50\u7684\u52dd\u80bd\u93c8\u3002\u93b3-\u4e09\u4e59\u9187\u80fa\u56db\u4e59\u9178 (Ni-NTA) \u8207\u74ca\u8102\u7cd6\u6a39\u8102 \u6216\u78c1\u73e0\u7d50\u5408\u662f\u5e38\u7528\u7684 IMAC \u5de5\u5177\uff0c\u7528\u65bc\u7d14\u5316\u591a\u7d44\u6c28\u9178 (His) \u6a19 \u8a18\u7684\u86cb\u767d\u8cea\u3002\u7531\u65bcHis\u6b98\u57fa\u53ef\u4ee5\u6574\u5408\u93b3\u96e2\u5b50(Ni?+),His \u6a19\u8a18\u7684\u86cb\u767d\u5c0d\u56fa\u5b9a\u4e86 Ni2* \u7684\u8f09\u9ad4\u8868\u73fe\u51fa\u5f37\u70c8\u7684\u89aa\u548c\u529b\u3002\u5728 Opentrons \u86cb\u767d\u8cea\u7d14\u5316\u5de5\u4f5c\u7ad9\u4e0a\u4f7f\u7528 Pierce Ni-NTA \u74ca\u8102\u7cd6\u78c1\u73e0\u8403\u53d6\u5e36\u6709 His\u6a19\u7c64\u7684GAPDH\u3002<\/p>\n<!-- \/wp:paragraph -->\n\n<!-- wp:paragraph -->\n<p><strong>\u6750\u6599\u8207\u65b9\u6cd5<\/strong>IMAC Protocol \u5be6\u9a57\u6d41\u7a0b\u6982\u8ff0IMAC protocol \u7684\u6d41\u7a0b\u5206\u70ba\u5e7e\u500b\u90e8\u5206\uff1a\u7b2c\u4e00\u90e8\u5206\uff1a\u6a23\u54c1\/\u78c1\u73e0\u6df7\u5408\u7269\u7684\u88fd\u5099\u3002\u76ee\u6a19\u6355\u7372\u90e8\u5206\u4e0d\u5728OT-2\u5e73\u53f0\u4e0a\u9032\u884c\u3002\u7b2c\u4e8c\u90e8\u5206\uff1a\u6d17\u6ecc\u548c\u6d17\u812b\u3002\u5177\u9ad4\u6d41\u7a0b\u5982\u57161\u6240\u793a\u3002\u7b2c\u4e00\u90e8\u5206\u548c\u7b2c\u4e8c\u90e8\u5206\u90fd\u5728OT-2 \u4e0a\u57f7\u884c\u3002\u91dd\u5c0d96\u500b\u6a23\u54c1\uff0c\u7b2c\u4e00 \u90e8\u5206\u904b\u884c\u6642\u9593\u70ba57\u5206\u9418\uff0c\u96a8\u5f8c\u5728\u6416\u5e8a\u4e0a\u9032\u884c30\u5206\u9418\u7684\u76ee\u6a19\u6355\u7372\u3002\u7b2c\u4e8c\u90e8\u5206\u9700\u898178\u5206\u9418\u5b8c\u6210\u3002<\/p>\n<!-- \/wp:paragraph -->\n\n<!-- wp:paragraph -->\n<p><strong>OT-2 \u5e73\u53f0\u4e0a\u7684 Ni-NTA \u5be6\u9a57\u6d41\u7a0b<\/strong>\u7b2c\u4e00\u90e8\u5206\u548c\u7b2c\u4e8c\u90e8\u5206\u7684\u8a2d\u5099\u4f48\u5c40\u5716\u5982\u4e0b(\u898b\u57162)\u3002\u5354\u8b70\u7684\u7b2c \u4e00\u90e8\u5206\u548c\u7b2c\u4e8c\u90e8\u5206\u5747\u5728 OT-2 \u5e73\u53f0\u4e0a\u9032\u884c\uff0c\u4e26\u4f7f\u7528\u78c1\u73e0\u7d14\u5316\u6a21 \u584a\u5c07\u78c1\u73e0\u5f9e\u6eb6\u6db2\u4e2d\u5206\u96e2\u51fa\u4f86\u3002\u6a23\u54c1\u8655\u7406\u904e\u7a0b\u4e2d\uff0c\u5c07500 \u03bcL \u4eba\u985e \u6e90 His\u6a19\u8a18\u7684 GAPDH \u6eb6\u6db2\u820712.5 \u03bcL \u6c89\u6fb1\u7684\u78c1\u73e0\u6df7\u5408\uff0c\u4e26\u5728 \u5ba4\u6eab\u4e0b\u4ee5800 rpm \u7684\u901f\u5ea6\u5b75\u80b230\u5206\u9418\u3002\u7d93\u904e2 \u500b\u6d17\u6ecc\u6b65\u9a5f \u5f8c\uff0c\u76ee\u6a19\u86cb\u767d\u5728250 \u03bcL \u6d17\u812b\u7de9\u885d\u6db2\u4e2d\u4ee5800 rpm \u548c\u5ba4\u6eab\u7684\u689d \u4ef6\u5728\u6416\u5e8a\u4e0a\u6d17\u812b10\u5206\u9418\u3002\u53d620 \u03bcL \u6d17\u812b\u7269\u9032\u884c SDS-PAGE \u548c Western Blot \u5206\u6790\uff0c\u4f7f\u7528\u55ae\u682a\u6297-GAPDH \u6297\u9ad4 (Thermo Fisher Scientific,USA)\u548c\u8207\u87a2\u5149\u67d3\u6599\u5076\u806f\u7684\u4e8c\u6297 (LI-COR Biosciences,USA) \u9032\u884c\u6aa2\u6e2c\u3002<\/p>\n<!-- \/wp:paragraph -->\n\n<!-- wp:image {\"id\":2991,\"sizeSlug\":\"full\",\"linkDestination\":\"none\"} -->\n<figure class=\"wp-block-image size-full\"><img src=\"https:\/\/opentrons.com.cn\/wp-content\/uploads\/2024\/06\/image-64.png\" alt=\"\u86cb\u767d\u8d28\u7eaf\u5316\u5de5\u4f5c\u7ad9\u4e0a\u57fa\u4e8e Ni-NTA \u78c1\u73e0\u7684\u86cb\u767d\u8d28\u7eaf\u5316\u5de5\u4f5c\u6d41\u7a0b\u6982\u8ff0\" class=\"wp-image-2991\"><\/figure>\n<!-- \/wp:image -->\n\n<!-- wp:image {\"id\":2992,\"width\":\"726px\",\"height\":\"auto\",\"sizeSlug\":\"full\",\"linkDestination\":\"none\"} -->\n<figure class=\"wp-block-image size-full is-resized\"><img src=\"https:\/\/opentrons.com.cn\/wp-content\/uploads\/2024\/06\/image-65.png\" alt=\"\u86cb\u767d\u8d28\u7eaf\u5316\u5de5\u4f5c\u7ad9\u4e0a\u8fd0\u884c Ni-NTA  \u5e94\u7528\u534f\u8bae\u7684\u7532\u677f\u5e03\u5c40\u56fe\" class=\"wp-image-2992\" style=\"width:726px;height:auto\"><\/figure>\n<!-- \/wp:image -->\n\n<!-- wp:paragraph -->\n<p><strong>\u7d50\u679c<\/strong>GAPDH \u5728\u91cd\u8907\u5be6\u9a57\u4e2d\u8403\u53d6\u5f97\u5230 \u4e00 \u81f4\u7684\u6a23\u54c1\u8655\u7406\u7d50\u679c(\u900f\u904eWestern blot \u5206\u6790\u7684\u86cb\u767d\u5e36\u8a0a\u865f\u6e2c\u5b9a\uff0c CV=6%) ( \u57163)\u3002\u90e8\u5206\u8403\u53d6\u7269\u900f\u904e\u4f7f\u7528 QubitTM \u86cb \u767d \u8cea \u5206 \u6790 (ThermoFisher Scientific,USA) \u9032\u884c\u5b9a\u91cf\uff0c\u8b49\u660e\u4e86\u9ad8\u86cb\u767d\u8cea\u7522\u91cf( \u5716 4A) \u3002 \u6b64 \u5916 \uff0c \u4f7f \u7528Opentrons \u86cb\u767d\u8cea\u7d14\u5316\u5de5\u4f5c\u7ad9\u9032\u884c \u7684\u86cb\u767d\u8cea\u7d14\u5316\u904e\u7a0b\u4e0d\u6703\u7834\u58de\u86cb\u767d\u8cea\u7684\u9176\u6d3b\u6027\uff0c\u9019\u662f\u901a\u904eGAPDH \u6d3b \u6027 \u5206 \u6790 (Sigma-Aldrich,St.Louis,MO,USA)\u6aa2 \u6e2c \u5f97 \u51fa \u7684 ( \u5716 4B)\u3002<\/p>\n<!-- \/wp:paragraph -->\n\n<!-- wp:image {\"id\":2993,\"sizeSlug\":\"full\",\"linkDestination\":\"none\",\"align\":\"center\"} -->\n<figure class=\"wp-block-image aligncenter size-full\"><img src=\"https:\/\/opentrons.com.cn\/wp-content\/uploads\/2024\/06\/image-66.png\" alt=\"His \u6807\u8bb0\u7684GAPDH\u63d0\u53d6\u7ed3\u679c\u7b26\u5408\u9884\u671f\" class=\"wp-image-2993\"><\/figure>\n<!-- \/wp:image -->\n\n<!-- wp:paragraph -->\n<p>Ni-NTA \u4ecb\u5c0e\u7684\u6572\u4f4e\u5be6\u9a57\u5c0d\u65bc\u5728\u7d30\u80de\u6216\u7d44\u7e54\u6a23\u672c\u4e2d\u5206\u96e2 \u5916\u6e90\u6027\u8868\u73fe\u7684 His \u6a19\u8a18\u86cb\u767d\u8cea\u975e\u5e38\u6709\u7528\u3002\u4e0a\u4e00\u500b\u65b9\u6848\u7528\u65bc\u7d14 \u5316 2 0 0 \u03bcLHEp-2 \u7d30\u80de\u88c2\u89e3\u6db2\u4e2d\u7684 His \u6a19 \u8a18 GAPDH\u3002\u7d50\u679c\u518d\u6b21\u8b49\u660e His-GAPDH \u6210\u529f\u88ab\u8403\u53d6\uff0c\u8207\u975e\u6a19\u8a18 \u86cb\u767d\u7684\u7d50\u5408\u76f8\u6bd4\uff0cNi-NTA \u78c1\u73e0\u7684\u89aa\u548c\u529b\u7279\u7570\u6027\u975e\u5e38\u9ad8 (\u57165)\u3002\u5728\u5927\u898f\u6a21\u86cb\u767d\u8cea\u751f\u7522\u4e2d\uff0c\u5e38\u7528\u5927\u8178\u687f\u83cc\u9032\u884c\u91cd\u7d44\u86cb\u767d\u8868\u73fe\u3002\u900f\u904e\u88c2\u89e3\u6709\u6216\u6c92\u6709 IPTG \u8a98\u5c0e\u7684 His\u6a19\u8a18 GAPDH \u8868\u73fe\u7684\u7d30\u83cc\u7d30\u80de\u4f86\u88fd\u5099\u7d30\u80de\u88c2\u89e3\u6db2\u3002\u7d50\u679c\u9032\u4e00\u6b65\u78ba\u8a8d\u4e86\u6b64\u65b9\u6848\u6210\u529f\u5730\u4f7f\u7528 Ni-NTA \u78c1\u73e0\u9032\u884c\u4e86\u5e36\u6709 His \u6a19 \u8a18 \u7684 \u86cb \u767d \u8cea \u7d14 \u5316 ( \u5716 6 ) \u3002\u5728\u9019\u9805\u7814\u7a76\u4e2d\u8655\u7406\u4e8696\u500b\u6a23\u672c\uff0c\u986f\u793a\u7684\u7d50\u679c\u50c5\u70ba\u572896 \u5b54\u6a23\u54c1\u677f\u7684\u6700\u5f8c \u4e00\u5217\u4e2d\u76848\u500b\u6a23\u54c1\u3002\u540c\u6642\uff0c\u5be6\u9a57\u7d50\u679c\u4e5f\u5c0d\u4e0d\u540c\u6a23\u672c\u898f\u6a21\u7684\u904b\u884c\u6642\u9593\u9032\u884c\u4e86\u8a55\u4f30(\u57167)\u3002<\/p>\n<!-- \/wp:paragraph -->\n\n<!-- wp:image {\"id\":2994,\"width\":\"349px\",\"height\":\"auto\",\"sizeSlug\":\"full\",\"linkDestination\":\"none\"} -->\n<figure class=\"wp-block-image size-full is-resized\"><img src=\"https:\/\/opentrons.com.cn\/wp-content\/uploads\/2024\/06\/image-67.png\" alt=\"\u5b9e\u9a8c\u4ea7\u51fa\u7684\u86cb\u767d\u8d28\u4ea7\u91cf\u9ad8\" class=\"wp-image-2994\" style=\"width:349px;height:auto\"><\/figure>\n<!-- \/wp:image -->\n\n<!-- wp:paragraph -->\n<p><\/p>\n<!-- \/wp:paragraph -->\n\n<!-- wp:paragraph -->\n<p><\/p>\n<!-- \/wp:paragraph -->\n\n<!-- wp:paragraph -->\n<p><strong>\u7d50\u8ad6<\/strong>\u672c\u6b21\u7814\u7a76\u5728 Opentrons \u86cb\u767d\u8cea\u7d14\u5316\u5de5\u4f5c\u7ad9\u4e0a\u904b\u884c\u4e86\u4e00\u7a2e\u57fa\u65bc\u78c1\u73e0\u7684 His \u6a19\u8a18\u91cd\u7d44 GAPDH \u86cb\u767d\u8cea\u7d14\u5316\u7684\u81ea\u52d5\u5316\u5be6\u9a57\u6d41\u7a0b\u89e3 \u6c7a\u65b9\u6848\u3002\u5be6\u9a57\u8b49\u660e\uff0c Opentrons \u86cb\u767d\u8cea\u7d14\u5316\u5de5\u4f5c\u7ad9\u53ef\u4ee5\u5728\u4e2d\u5230\u9ad8\u901a\u91cf\u7684\u8a2d\u5b9a\u4e2d\u7a69\u5b9a\u4e14\u6709\u6548\u7387\u5730\u8655\u7406\u6a23\u672c\uff0c\u6e1b\u5c11\u64cd\u4f5c\u6642\u9593\uff0c\u540c\u6642\u63d0\u4f9b\u7b26\u5408\u9810\u671f\u7684\u53ef\u91cd\u8907\u6027\u7684\u7522\u51fa\u3002<\/p>\n<!-- \/wp:paragraph -->\n\n<!-- wp:paragraph -->\n<p>\u539f\u6587\u7db2\u5740\uff1a<strong><a href=\"https:\/\/opentrons.com.cn\/wp-content\/uploads\/2023\/11\/Automated-Ni-NTA-bead-based-protein-purification-on-the-OT -2-liquid-handler-CN.pdf\" target=\"_blank\" 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