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mRNA\u304cCas9\u30bf\u30f3\u30d1\u30af\u8cea\u306b\u7ffb\u8a33\u3055\u308c\u3001\u30ac\u30a4\u30c9RNA\u3068\u30b3\u30f3\u30d7\u30ec\u30c3\u30af\u30b9\u3092\u5f62\u6210\u3057\u3066\u3001\u6838\u306b\u79fb\u884c\u3057\u307e\u3059\u3002\r\nProtein\u65b9\u6cd5\u3067\u3082\u3001\u30ac\u30a4\u30c9RNA\u3068Cas9\u30bf\u30f3\u30d1\u30af\u8cea\u3092\u540c\u6642\u306b\u5c0e\u5165\u3057\u307e\u3059\u3002\u7d30\u80de\u8cea\u306b\u5c0e\u5165\u3055\u308c\u305f\u30ac\u30a4\u30c9RNA\u3068Cas9\u30bf\u30f3\u30d1\u30af\u8cea\u304c\u30b3\u30f3\u30d7\u30ec\u30c3\u30af\u30b9\u3092\u5f62\u6210\u5f8c\u3001\u6838\u306b\u79fb\u884c\u3057\u307e\u3059\u3002\r\n3\u3064\u306e\u65b9\u6cd5\u3044\u305a\u308c\u3082\u3001\u6838\u5185\u79fb\u884c\u5f8c\u306f\u30ac\u30a4\u30c9RNA\u304c\u76ee\u7684\u907a\u4f1d\u5b50\u306b\u7d50\u5408\u3057\u3066\u3001\u30b2\u30ce\u30e0\u3092\u5207\u65ad\u3057\u307e\u3059\u3002\r\n\r\nA\u3055\u3093\uff1a\r\n\u306a\u308b\u307b\u3069\u3002\r\n\u3067\u3001\u7d50\u5c40\u3069\u308c\u304c\u4e00\u756a\u304a\u52e7\u3081\u306a\u3093\u3067\u3059\u304b\uff1f\r\n\r\n\u5148\u8f29\uff1a\r\n\u3046\u3093\u3001\u305d\u308c\u304c\u77e5\u308a\u305f\u3044\u3088\u306d\u3002\u30b2\u30ce\u30e0\u7de8\u96c6\u306e\u52b9\u7387\u304c\u9ad8\u3044\u65b9\u6cd5\u306f\u3001Protein\u65b9\u6cd5\u3060\u3088\r\n\r\n\u300c\u30b2\u30ce\u30e0\u7de8\u96c6\u52b9\u7387\u304c\u9ad8\u3044\u300d\u3068\u306f\u3001\u300c\u5207\u65ad\u3057\u305f\u5f8c\u306b\u5909\u7570\u304c\u5165\u308b\u52b9\u7387\u304c\u9ad8\u3044\u300d\u3053\u3068\u3092\u6307\u3057\u307e\u3059\u3002\r\n\u30b2\u30ce\u30e0\u7de8\u96c6\u52b9\u7387\u304c\u9ad8\u3044\u65b9\u6cd5\u306f\u3001Protein\u65b9\u6cd5\u3067\u3059\u3002\r\n\u7b2c2\u4e16\u4ee3\u306eRNA\u65b9\u6cd5\u306fDNA\uff08Vector\uff09\u65b9\u6cd5\u306e\u30b2\u30ce\u30e0\u7de8\u96c6\u52b9\u7387\u3092\u6539\u5584\u3057\u305f\u65b9\u6cd5\u306e\u305f\u3081\u3001DNA\u65b9\u6cd5\u3088\u308a\u3082RNA\u65b9\u6cd5\u306e\u65b9\u304c\u30b2\u30ce\u30e0\u7de8\u96c6\u52b9\u7387\u306f\u9ad8\u3044\u3067\u3059\u3002\r\n\r\nA\u3055\u3093\uff1a\r\n\u3068\u3053\u308d\u3067\u3001Protein\u65b9\u6cd5\u306e\u30c8\u30e9\u30f3\u30b9\u30d5\u30a7\u30af\u30b7\u30e7\u30f3\u52b9\u7387\u306f\u3069\u3046\u306a\u3093\u3067\u3057\u3087\u3046\u3002\r\n\u30aa\u30d5\u30bf\u30fc\u30b2\u30c3\u30c8\u52b9\u679c\u3082\u6c17\u306b\u306a\u308a\u307e\u3059\u3002\r\n\r\n\u305d\u3046\u3001\u4f9b\u4e0e\u65b9\u6cd5\u3092\u9078\u3076\u3068\u304d\u306f\u3001\u30b2\u30ce\u30e0\u7de8\u96c6\u52b9\u7387\u306e\u307b\u304b\u3001\u30c8\u30e9\u30f3\u30b9\u30d5\u30a7\u30af\u30b7\u30e7\u30f3\u52b9\u7387\u3084\u30aa\u30d5\u30bf\u30fc\u30b2\u30c3\u30c8\u52b9\u679c\u3082\u8003\u3048\u306a\u304f\u3066\u306f\u306a\u308a\u307e\u305b\u3093\u3002\u30c8\u30e9\u30f3\u30b9\u30d5\u30a7\u30af\u30b7\u30e7\u30f3\u52b9\u7387\u3067\u8ecd\u914d\u304c\u4e0a\u304c\u308b\u306e\u306fDNA\uff08vector\uff09\u65b9\u6cd5\u3067\u3059\u3002\u9078\u629e\u30de\u30fc\u30ab\u30fc\uff08\u86cd\u5149\u30bf\u30f3\u30d1\u30af\u8cea\u3084\u85ac\u5264\u62b5\u6297\u30bf\u30f3\u30d1\u30af\u8cea\uff09\u304c\u63b2\u8f09\u3055\u308c\u3066\u3044\u308b\u306e\u3067\u3001\u5c0e\u5165\u3055\u308c\u305f\u7d30\u80de\u306e\u6fc3\u7e2e\u3084\u5c0e\u5165\u52b9\u7387\u306e\u78ba\u8a8d\u304c\u3057\u3084\u3059\u3044\u3067\u3059\u3002\u4e00\u65b9\u3001Protein\u65b9\u6cd5\u306fDNA\u65b9\u6cd5\u3068\u6bd4\u3079\u3066\u3001\u4e00\u822c\u7684\u306b\u30c8\u30e9\u30f3\u30b9\u30d5\u30a7\u30af\u30b7\u30e7\u30f3\u52b9\u7387\u306f\u9ad8\u304f\u3042\u308a\u307e\u305b\u3093\u3002\u305d\u306e\u305f\u3081\u3001\u57f9\u990a\u7d30\u80de\u306b\u52b9\u7387\u3088\u304fCas9\u30bf\u30f3\u30d1\u30af\u8cea\u304c\u5c0e\u5165\u3067\u304d\u308b\u306e\u304b\u3001\u3042\u3089\u304b\u3058\u3081\u78ba\u8a8d\u304c\u5fc5\u8981\u3067\u3059\u3002\r\n\r\n\u307e\u305f\u3001\u30aa\u30d5\u30bf\u30fc\u30b2\u30c3\u30c8\u52b9\u679c\u306fProtein\u65b9\u6cd5\u304c\u6700\u3082\u4f4e\u3044\u3067\u3059\u3002\u3053\u308c\u306f\u3001\u30bf\u30f3\u30d1\u30af\u8cea\u304c\u7d30\u80de\u5185\u306b\u6b8b\u308a\u3065\u3089\u304f\u3001\u30bf\u30fc\u30b2\u30c3\u30c8\u4ee5\u5916\u306e\u907a\u4f1d\u5b50\u306b\u4f5c\u7528\u3057\u3065\u3089\u3044\u305f\u3081\u3060\u3068\u8a00\u308f\u308c\u3066\u3044\u307e\u3059\u3002\r\n\r\n\u5148\u8f29\uff1a\r\n3\u3064\u306e\u65b9\u6cd5\u3092\u307e\u3068\u3081\u3066\u307f\u305f\u3088\r\n\r\n \t\u30b2\u30ce\u30e0\u7de8\u96c6\u52b9\u7387\u306e\u9ad8\u3055\uff1aProtein\uff1eRNA\uff1eDNA(Vector)\r\n \t\u30aa\u30d5\u30bf\u30fc\u30b2\u30c3\u30c8\u52b9\u679c\u306e\u4f4e\u3055\uff1aProtein\uff1eRNA\uff1eDNA(Vector)\r\n \t\u30c8\u30e9\u30f3\u30b9\u30d5\u30a7\u30af\u30b7\u30e7\u30f3\u52b9\u7387\u306e\u9ad8\u3055\u203b\uff1aDNA(Vector)\uff1eRNA\uff1eProtein\r\n\u203b\u6bd4\u8f03\u304c\u96e3\u3057\u304f\u3001\u76f4\u63a5\u6bd4\u8f03\u3057\u305f\u30c7\u30fc\u30bf\u304c\u306a\u3044\u305f\u3081\u3001\u539f\u7406\u304b\u3089\u63a8\u6e2c\u3055\u308c\u308b\u9806\u756a\u3067\u3059\u3002\r\n\r\nA\u3055\u3093\uff1a\r\n\u3058\u3083\u3042\u3001Protein\u65b9\u6cd5\u3067\u8a66\u3057\u3066\u307f\u307e\u3059\uff01\r\n\u3048\u3063\u3068\u3001\u5b9f\u9a13\u306b\u5fc5\u8981\u306a\u306e\u306f\u2026\r\n\r\n\u5148\u8f29\uff1a\r\n\u307e\u305a\u306f\u5927\u4e8b\u306a2\u3064\u3002\r\n\u30ac\u30a4\u30c9RNA\u3068Cas9 Protein\u3060\u306d\u3002\r\n\r\n\u4f9b\u4e0e\u65b9\u6cd5\u304c\u6c7a\u307e\u3063\u305f\u3089\u3001\u88681\u3092\u53c2\u8003\u306b\u3057\u3066Cas9\u3068\u30ac\u30a4\u30c9RNA\u3092\u7528\u610f\u3057\u3066\u304a\u304d\u307e\u3057\u3087\u3046\u3002\r\n\u88681\u3000Cas9\u3068\u30ac\u30a4\u30c9RNA\u306e\u6e96\u5099\u65b9\u6cd5\r\n\r\n\r\n\r\nCas9\r\n\u30ac\u30a4\u30c9RNA\r\n\u5099\u8003\r\n\r\n\r\n\r\n\r\nDNA\u65b9\u6cd5\r\n(Vector)\r\nInvitrogen\u2122 GeneArt\u2122 CRISPR vector\u30b7\u30b9\u30c6\u30e0\r\nInvitrogen\u2122 TrueDesign Genome Editor\u306a\u3069\u3067\u914d\u5217\u3092\u30c7\u30b6\u30a4\u30f3\u3057\u3001Primer\u5408\u6210\u306a\u3069\u3067\u5165\u624b\u203b\r\nCRISPR vector\u306f\u54fa\u4e73\u985e\u7d30\u80de\u7528\u306evector\u306e\u305f\u3081\u3001\u54fa\u4e73\u985e\u7d30\u80de\u4ee5\u5916\u306e\u30b5\u30f3\u30d7\u30eb\u306e\u5834\u5408\u306f\u3001\u305d\u306e\u7d30\u80de\u7a2e\u306b\u5bfe\u5fdc\u3057\u305fvector\u3092\u3054\u7528\u610f\u304f\u3060\u3055\u3044\u3002\r\n\r\n\r\nRNA\u65b9\u6cd5\r\nInvitrogen\u2122 GeneArt\u2122 Cas9 mRNA\r\nInvitrogen\u2122 TrueGuide RNA\uff08\u30c7\u30b6\u30a4\u30f3\u30b5\u30a4\u30c8\uff09\u3067SpCas9\u7528\u30ac\u30a4\u30c9RNA\u304c\u5165\u624b\u53ef\u80fd\r\nCas9 mRNA\u306fSpCas9 mRNA\u3067\u3001\u6838\u79fb\u884c\u30b7\u30b0\u30ca\u30eb\u304c\u4ed8\u4e0e\r\n\r\n\r\nProtein\u65b9\u6cd5\r\nTrueCut Cas9 Protein\r\nInvitrogen\u2122 TrueCut Cas9 \u2026","hashtag":"ThermoFisherJP","subscribe":"content","subscribe_url":"","activestatus":"1","singular":"1","twitter_popup":"1","refresh":"0","nonce":"512ab7dbaa","postid":"13935","servertime":"1732740731","ajaxurl":"https:\/\/www.thermofisher.com\/blog\/wp-admin\/admin-ajax.php"}; /* ]]> */ </script> <script type="text/javascript" src="https://www.thermofisher.com/blog/learning-at-the-bench/wp-content/plugins/mashsharer/assets/js/mashsb.min.js?ver=4.0.47" id="mashsb-js"></script> <script type="text/javascript" src="https://www.thermofisher.com/blog/learning-at-the-bench/wp-content/plugins/mashshare-google-analytics/assets/js/mashga.min.js?ver=1.0.9" id="mashga-js"></script> <script type="text/javascript" id="mashnet-js-extra"> /* <![CDATA[ */ var mashnet = {"body":"Check out this article I just read:","subject":"Check out this article from Accelerating 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\tDNA\uff08Vector\uff09\u65b9\u6cd5\uff1a\r\n\u7b2c\u4e00\u4e16\u4ee3\u3002Cas9\u3068\u30ac\u30a4\u30c9RNA\u4e21\u65b9\u306e\u914d\u5217\u3092DNA\uff08Vector\uff09\u306b\u7d44\u307f\u8fbc\u3093\u3067\u3001\u7d30\u80de\u306b\u4f9b\u4e0e\u3059\u308b\u65b9\u6cd5\r\n \tRNA\uff08Cas9 mRNA\uff09\u65b9\u6cd5\uff1a\r\n\u7b2c\u4e8c\u4e16\u4ee3\u3002Cas9\u3068\u30ac\u30a4\u30c9RNA\u3092\u3001\u305d\u308c\u305e\u308cRNA\u306e\u72b6\u614b\u3067\u7d30\u80de\u306b\u4f9b\u4e0e\u3059\u308b\u65b9\u6cd5\r\n \tProtein\uff08Cas9 protein\uff09\u65b9\u6cd5\uff1a\r\nCas9 \u3092Cas9 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\t\u30b2\u30ce\u30e0\u7de8\u96c6\u52b9\u7387\u306e\u9ad8\u3055\uff1aProtein\uff1eRNA\uff1eDNA(Vector)\r\n \t\u30aa\u30d5\u30bf\u30fc\u30b2\u30c3\u30c8\u52b9\u679c\u306e\u4f4e\u3055\uff1aProtein\uff1eRNA\uff1eDNA(Vector)\r\n \t\u30c8\u30e9\u30f3\u30b9\u30d5\u30a7\u30af\u30b7\u30e7\u30f3\u52b9\u7387\u306e\u9ad8\u3055\u203b\uff1aDNA(Vector)\uff1eRNA\uff1eProtein\r\n\u203b\u6bd4\u8f03\u304c\u96e3\u3057\u304f\u3001\u76f4\u63a5\u6bd4\u8f03\u3057\u305f\u30c7\u30fc\u30bf\u304c\u306a\u3044\u305f\u3081\u3001\u539f\u7406\u304b\u3089\u63a8\u6e2c\u3055\u308c\u308b\u9806\u756a\u3067\u3059\u3002\r\n\r\nA\u3055\u3093\uff1a\r\n\u3058\u3083\u3042\u3001Protein\u65b9\u6cd5\u3067\u8a66\u3057\u3066\u307f\u307e\u3059\uff01\r\n\u3048\u3063\u3068\u3001\u5b9f\u9a13\u306b\u5fc5\u8981\u306a\u306e\u306f\u2026\r\n\r\n\u5148\u8f29\uff1a\r\n\u307e\u305a\u306f\u5927\u4e8b\u306a2\u3064\u3002\r\n\u30ac\u30a4\u30c9RNA\u3068Cas9 Protein\u3060\u306d\u3002\r\n\r\n\u4f9b\u4e0e\u65b9\u6cd5\u304c\u6c7a\u307e\u3063\u305f\u3089\u3001\u88681\u3092\u53c2\u8003\u306b\u3057\u3066Cas9\u3068\u30ac\u30a4\u30c9RNA\u3092\u7528\u610f\u3057\u3066\u304a\u304d\u307e\u3057\u3087\u3046\u3002\r\n\u88681\u3000Cas9\u3068\u30ac\u30a4\u30c9RNA\u306e\u6e96\u5099\u65b9\u6cd5\r\n\r\n\r\n\r\nCas9\r\n\u30ac\u30a4\u30c9RNA\r\n\u5099\u8003\r\n\r\n\r\n\r\n\r\nDNA\u65b9\u6cd5\r\n(Vector)\r\nInvitrogen\u2122 GeneArt\u2122 CRISPR vector\u30b7\u30b9\u30c6\u30e0\r\nInvitrogen\u2122 TrueDesign Genome Editor\u306a\u3069\u3067\u914d\u5217\u3092\u30c7\u30b6\u30a4\u30f3\u3057\u3001Primer\u5408\u6210\u306a\u3069\u3067\u5165\u624b\u203b\r\nCRISPR vector\u306f\u54fa\u4e73\u985e\u7d30\u80de\u7528\u306evector\u306e\u305f\u3081\u3001\u54fa\u4e73\u985e\u7d30\u80de\u4ee5\u5916\u306e\u30b5\u30f3\u30d7\u30eb\u306e\u5834\u5408\u306f\u3001\u305d\u306e\u7d30\u80de\u7a2e\u306b\u5bfe\u5fdc\u3057\u305fvector\u3092\u3054\u7528\u610f\u304f\u3060\u3055\u3044\u3002\r\n\r\n\r\nRNA\u65b9\u6cd5\r\nInvitrogen\u2122 GeneArt\u2122 Cas9 mRNA\r\nInvitrogen\u2122 TrueGuide RNA\uff08\u30c7\u30b6\u30a4\u30f3\u30b5\u30a4\u30c8\uff09\u3067SpCas9\u7528\u30ac\u30a4\u30c9RNA\u304c\u5165\u624b\u53ef\u80fd\r\nCas9 mRNA\u306fSpCas9 mRNA\u3067\u3001\u6838\u79fb\u884c\u30b7\u30b0\u30ca\u30eb\u304c\u4ed8\u4e0e\r\n\r\n\r\nProtein\u65b9\u6cd5\r\nTrueCut Cas9 Protein\r\nInvitrogen\u2122 TrueCut Cas9 \u2026"}; /* ]]> */ </script> <script type="text/javascript" src="https://www.thermofisher.com/blog/learning-at-the-bench/wp-content/plugins/mashshare-networks/assets/js/mashnet.min.js?ver=2.5.3" id="mashnet-js"></script> <script type="text/javascript" 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Scientific">Thermo Fisher Scientific</a></p></div><div class="widget-area header-widget-area"><section id="nav_menu-2" class="widget widget_nav_menu"><div class="widget-wrap"><nav class="nav-header"><ul id="menu-main-menu" class="menu genesis-nav-menu"><li id="menu-item-5525" class="menu-item menu-item-type-custom menu-item-object-custom menu-item-has-children menu-item-5525"><a><span >メニュー</span></a> <ul class="sub-menu"> <li id="menu-item-5528" class="menu-item menu-item-type-taxonomy menu-item-object-category current-post-ancestor current-menu-parent current-post-parent menu-item-5528"><a href="https://www.thermofisher.com/blog/learning-at-the-bench/mol-bio/"><span >分子生物学実験関連</span></a></li> <li id="menu-item-5529" class="menu-item menu-item-type-taxonomy menu-item-object-category menu-item-5529"><a href="https://www.thermofisher.com/blog/learning-at-the-bench/analysis/"><span >分析・測定関連</span></a></li> <li id="menu-item-5531" class="menu-item menu-item-type-taxonomy 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>お問い合わせ</span></a></li> </ul></nav></div></section> </div><div class="breadcrumb"><span class="breadcrumb-link-wrap"><a href="https://www.thermofisher.com/blog/?">Accelerating Science</a></span><span class="breadcrumb-link-wrap"><a class="breadcrumb-link" href="https://www.thermofisher.com/blog/learning-at-the-bench/"><span class="breadcrumb-link-text-wrap">Learning at the Bench</span></a><meta ></span> <span aria-label="breadcrumb separator">/</span> <span class="breadcrumb-link-wrap"><a class="breadcrumb-link" href="https://www.thermofisher.com/blog/learning-at-the-bench/mol-bio/"><span class="breadcrumb-link-text-wrap">分子生物学実験関連</span></a><meta ></span> <span aria-label="breadcrumb separator">/</span> <span class="breadcrumb-current-title">そういうことだったのか ! ゲノム編集実験（CRISPR/Cas9）～第3回　3つのCRIPSR/Cas9実験系～</span></div></div></header><div class="site-inner"><div class="content-sidebar-wrap"><main class="content"><article class="post-13935 post type-post status-publish format-standard has-post-thumbnail category-mol-bio tag-crispr-cas9 tag-genome-editing entry" aria-label="そういうことだったのか ! ゲノム編集実験（CRISPR/Cas9）～第3回　3つのCRIPSR/Cas9実験系～"><header class="entry-header"><h1 class="entry-title">そういうことだったのか ! ゲノム編集実験（CRISPR/Cas9）～第3回　3つのCRIPSR/Cas9実験系～</h1> <span class="post-info-author"> 作成者 <a href="https://www.thermofisher.com/blog/author/lgbadmin/" title="LatB Staff の投稿" rel="author">LatB Staff</a> </span> <span class="post-info-date"> 05.08.2023 </span> </header><div class="entry-content"><p>ゲノム編集の初心者Aさんによる「初めてのゲノム編集実験」。<a href="https://www.thermofisher.com/blog/learning-at-the-bench/gene-editing_bid_ts_1/?CID=bid_syn_gns_r04_jp_cp0000_pjt0000_bid00000_0so_blg_op_lgn_kt_s00_gene_editing_bid_ts_3_Social_LAB" target="_blank" rel="noopener">第1回</a>、<a href="https://www.thermofisher.com/blog/learning-at-the-bench/gene-editing_bid_ts_2/?CID=bid_syn_gns_r04_jp_cp0000_pjt0000_bid00000_0so_blg_op_lgn_kt_s00_gene_editing_bid_ts_3_Social_LAB" target="_blank" rel="noopener">第2回</a>を通して基礎的な内容を理解したAさんは、いよいよ実験を始めようとしています。<img decoding="async" class="aligncenter wp-image-13986" src="https://www.thermofisher.com/blog/learning-at-the-bench/wp-content/uploads/sites/13/2023/04/gene-editing_bid_ts_3-01-300x160.png" alt="" width="500" height="267" srcset="https://www.thermofisher.com/blog/learning-at-the-bench/wp-content/uploads/sites/13/2023/04/gene-editing_bid_ts_3-01-300x160.png 300w, https://www.thermofisher.com/blog/learning-at-the-bench/wp-content/uploads/sites/13/2023/04/gene-editing_bid_ts_3-01-1024x546.png 1024w, https://www.thermofisher.com/blog/learning-at-the-bench/wp-content/uploads/sites/13/2023/04/gene-editing_bid_ts_3-01-768x410.png 768w, https://www.thermofisher.com/blog/learning-at-the-bench/wp-content/uploads/sites/13/2023/04/gene-editing_bid_ts_3-01-1536x819.png 1536w, https://www.thermofisher.com/blog/learning-at-the-bench/wp-content/uploads/sites/13/2023/04/gene-editing_bid_ts_3-01.png 1787w" sizes="(max-width: 500px) 100vw, 500px" /></p> <div class="voice-text-right"> <p><span style="color: #3366ff">Aさん</span>：<br /> ガイドRNAがデザインできました！<br /> これで培養細胞のノックアウト実験ができますね</p> <p><span style="color: #ff00ff">先輩</span>：<br /> 待って。その前に<b>CIRSPR/Cas9の3つの実験系</b>からちゃんと選んだ？</p> <p><span style="color: #3366ff">Aさん</span>：<br /> 3つの実験系って？</p> <p>供与方法、つまり「Cas9とガイドRNAをどうやって細胞に与えるか」には、3つ方法があります。実験を行う前に、この3種類のどの方法で実施するか、選択しておきましょう。</p> <ul> <li>DNA（Vector）方法：<br /> 第一世代。Cas9とガイドRNA両方の配列をDNA（Vector）に組み込んで、細胞に供与する方法</li> <li>RNA（Cas9 mRNA）方法：<br /> 第二世代。Cas9とガイドRNAを、それぞれRNAの状態で細胞に供与する方法</li> <li>Protein（Cas9 protein）方法：<br /> Cas9 をCas9 タンパク質、ガイドRNAをRNAとして細胞に供与する方法</li> </ul> <p><span style="color: #3366ff">Aさん</span>：<br /> つまり、材料の作り方が違うんですね。<br /> ってことは、細胞内の動きも違うんですか？</p> <p><span style="color: #ff00ff">先輩</span>：<br /> いい質問だね。図1を見てみよう。3つの方法の細胞内挙動の違いが説明してあるよ</p> <div id="attachment_13982" style="width: 497px" class="wp-caption aligncenter"><img decoding="async" aria-describedby="caption-attachment-13982" class="wp-image-13982 size-full" src="https://www.thermofisher.com/blog/learning-at-the-bench/wp-content/uploads/sites/13/2023/04/gene-editing_bid_ts_3-02.png" alt="3つの方法で導入されたCas9とガイドRNAが細胞内で核内に移行するまで" width="487" height="291" srcset="https://www.thermofisher.com/blog/learning-at-the-bench/wp-content/uploads/sites/13/2023/04/gene-editing_bid_ts_3-02.png 487w, https://www.thermofisher.com/blog/learning-at-the-bench/wp-content/uploads/sites/13/2023/04/gene-editing_bid_ts_3-02-300x179.png 300w" sizes="(max-width: 487px) 100vw, 487px" /><p id="caption-attachment-13982" class="wp-caption-text">図1： 3つの方法で導入されたCas9とガイドRNAが<br />細胞内で核内に移行するまで</p></div> <p>DNA（Vector）方法では、vectorが細胞内に導入されると、核に移行したvectorからガイドRNAとCas9 mRNAが転写され、細胞質でCas9 タンパク質が翻訳されます。その後、細胞質内でガイドRNAとCas9タンパク質がコンプレックスを形成して、Cas9 proteinの核移行シグナルによって核内に移行します。<br /> RNA方法は、ガイドRNAとCas9 mRNAを同時に導入します。細胞質に導入されたCas9 mRNAがCas9タンパク質に翻訳され、ガイドRNAとコンプレックスを形成して、核に移行します。<br /> Protein方法でも、ガイドRNAとCas9タンパク質を同時に導入します。細胞質に導入されたガイドRNAとCas9タンパク質がコンプレックスを形成後、核に移行します。<br /> 3つの方法いずれも、核内移行後はガイドRNAが目的遺伝子に結合して、ゲノムを切断します。</p> <p><span style="color: #3366ff">Aさん</span>：<br /> なるほど。<br /> で、結局どれが一番お勧めなんですか？</p> <p><span style="color: #ff00ff">先輩</span>：<br /> うん、それが知りたいよね。ゲノム編集の効率が高い方法は、Protein方法だよ</p> <p>「ゲノム編集効率が高い」とは、「切断した後に変異が入る効率が高い」ことを指します。<br /> ゲノム編集効率が高い方法は、Protein方法です。<br /> 第2世代のRNA方法はDNA（Vector）方法のゲノム編集効率を改善した方法のため、DNA方法よりもRNA方法の方がゲノム編集効率は高いです。</p> <p><span style="color: #3366ff">Aさん</span>：<br /> ところで、Protein方法のトランスフェクション効率はどうなんでしょう。<br /> オフターゲット効果も気になります。</p> <p>そう、供与方法を選ぶときは、ゲノム編集効率のほか、トランスフェクション効率やオフターゲット効果も考えなくてはなりません。トランスフェクション効率で軍配が上がるのはDNA（vector）方法です。選択マーカー（蛍光タンパク質や薬剤抵抗タンパク質）が掲載されているので、導入された細胞の濃縮や導入効率の確認がしやすいです。一方、Protein方法はDNA方法と比べて、一般的にトランスフェクション効率は高くありません。そのため、培養細胞に効率よくCas9タンパク質が導入できるのか、あらかじめ確認が必要です。<img loading="lazy" decoding="async" class="aligncenter size-full wp-image-13983" src="https://www.thermofisher.com/blog/learning-at-the-bench/wp-content/uploads/sites/13/2023/04/gene-editing_bid_ts_3-03-e1682407330444.jpg" alt="" width="389" height="260" srcset="https://www.thermofisher.com/blog/learning-at-the-bench/wp-content/uploads/sites/13/2023/04/gene-editing_bid_ts_3-03-e1682407330444.jpg 389w, https://www.thermofisher.com/blog/learning-at-the-bench/wp-content/uploads/sites/13/2023/04/gene-editing_bid_ts_3-03-e1682407330444-300x201.jpg 300w, https://www.thermofisher.com/blog/learning-at-the-bench/wp-content/uploads/sites/13/2023/04/gene-editing_bid_ts_3-03-e1682407330444-96x65.jpg 96w" sizes="(max-width: 389px) 100vw, 389px" /></p> <p>また、オフターゲット効果はProtein方法が最も低いです。これは、タンパク質が細胞内に残りづらく、ターゲット以外の遺伝子に作用しづらいためだと言われています。</p> <p><span style="color: #ff00ff">先輩</span>：<br /> 3つの方法をまとめてみたよ</p> <ul> <li>ゲノム編集効率の高さ：Protein＞RNA＞DNA(Vector)</li> <li>オフターゲット効果の低さ：Protein＞RNA＞DNA(Vector)</li> <li>トランスフェクション効率の高さ<sup>※</sup>：DNA(Vector)＞RNA＞Protein<br /> <span style="font-size: small">※比較が難しく、直接比較したデータがないため、原理から推測される順番です。</span></li> </ul> <p><span style="color: #3366ff"><img loading="lazy" decoding="async" class="aligncenter wp-image-14114 size-full" src="https://www.thermofisher.com/blog/learning-at-the-bench/wp-content/uploads/sites/13/2023/04/gene-editing_bid_ts_3-04-e1683527966161.png" alt="" width="3370" height="1823" srcset="https://www.thermofisher.com/blog/learning-at-the-bench/wp-content/uploads/sites/13/2023/04/gene-editing_bid_ts_3-04-e1683527966161.png 3370w, https://www.thermofisher.com/blog/learning-at-the-bench/wp-content/uploads/sites/13/2023/04/gene-editing_bid_ts_3-04-e1683527966161-300x162.png 300w, https://www.thermofisher.com/blog/learning-at-the-bench/wp-content/uploads/sites/13/2023/04/gene-editing_bid_ts_3-04-e1683527966161-1024x554.png 1024w, https://www.thermofisher.com/blog/learning-at-the-bench/wp-content/uploads/sites/13/2023/04/gene-editing_bid_ts_3-04-e1683527966161-768x415.png 768w, https://www.thermofisher.com/blog/learning-at-the-bench/wp-content/uploads/sites/13/2023/04/gene-editing_bid_ts_3-04-e1683527966161-1536x831.png 1536w, https://www.thermofisher.com/blog/learning-at-the-bench/wp-content/uploads/sites/13/2023/04/gene-editing_bid_ts_3-04-e1683527966161-2048x1108.png 2048w" sizes="(max-width: 3370px) 100vw, 3370px" />Aさん</span>：<br /> じゃあ、Protein方法で試してみます！<br /> えっと、実験に必要なのは…</p> <p><span style="color: #ff00ff">先輩</span>：<br /> まずは大事な2つ。<br /> ガイドRNAとCas9 Proteinだね。</p> <p>供与方法が決まったら、表1を参考にしてCas9とガイドRNAを用意しておきましょう。</p> <table border="2" cellpadding="10%"> <caption>表1　Cas9とガイドRNAの準備方法</caption> <thead> <tr bgcolor="aliceblue"> <td style="text-align: center" width="75"></td> <td style="text-align: center" width="104"><b>Cas9</b></td> <td style="text-align: center" width="161"><b>ガイドRNA</b></td> <td style="text-align: center" width="226"><b>備考</b></td> </tr> </thead> <tbody> <tr> <td width="75">DNA方法<br /> (Vector)</td> <td width="104"><a href="https://www.thermofisher.com/order/catalog/product/A21174?CID=bid_syn_gns_r04_jp_cp0000_pjt0000_bid00000_0so_blg_op_lgn_kt_s00_gene_editing_bid_ts_3_Social_LAB" target="_blank" rel="noopener">Invitrogen™ GeneArt™ CRISPR vector</a>システム</td> <td width="161">Invitrogen™ TrueDesign Genome Editorなどで配列をデザインし、Primer合成などで入手<sup>※</sup></td> <td width="226">CRISPR vectorは哺乳類細胞用のvectorのため、哺乳類細胞以外のサンプルの場合は、その細胞種に対応したvectorをご用意ください。</td> </tr> <tr> <td width="75">RNA方法</td> <td width="104"><a href="https://www.thermofisher.com/order/catalog/product/A29378?CID=bid_syn_gns_r04_jp_cp0000_pjt0000_bid00000_0so_blg_op_lgn_kt_s00_gene_editing_bid_ts_3_Social_LAB" target="_blank" rel="noopener">Invitrogen™ GeneArt™ Cas9 mRNA</a></td> <td rowspan="2" width="161"><a href="https://www.thermofisher.com/jp/ja/home/life-science/genome-editing/crispr-libraries/trueguide-grnas.html?CID=bid_syn_gns_r04_jp_cp0000_pjt0000_bid00000_0so_blg_op_lgn_kt_s00_gene_editing_bid_ts_3_Social_LAB" target="_blank" rel="noopener">Invitrogen™ TrueGuide RNA</a>（デザインサイト）でSpCas9用ガイドRNAが入手可能</td> <td width="226">Cas9 mRNAはSpCas9 mRNAで、核移行シグナルが付与</td> </tr> <tr> <td width="75">Protein方法</td> <td width="104"><a href="https://www.thermofisher.com/order/catalog/product/A36496?CID=bid_syn_gns_r04_jp_cp0000_pjt0000_bid00000_0so_blg_op_lgn_kt_s00_gene_editing_bid_ts_3_Social_LAB" target="_blank" rel="noopener">TrueCut Cas9 Protein</a></td> <td width="226">Invitrogen™ TrueCut Cas9 ProteinはSpCas9 Proteinで、核移行シグナルが付与</td> </tr> </tbody> </table> <p><span style="font-size: small">※3’末端側に配列の付加が必要など、いくつか注意点があります。ガイドRNAを合成する前に、必ずCRISPR vectorのプロトコルから詳細を確認してください。</span></p> </div> <p>Aさんは表1を参考にして、Cas9 ProteinとガイドRNAを手に入れることにしました。次回は用意したCas9とガイドRNAを用いて、Aさんがいよいよトランスフェクションに挑みます。</p> <h2>まとめ</h2> <ol> <li>CRISPR/Cas9システムには3つの供与方法（DNA(Vector)、RNA、およびProtein方法)がある</li> <li>Protein方法がゲノム編集効率が高く、オフターゲット効果も低い</li> </ol> <p>Aさんが初めてのゲノム編集に挑戦するシリーズ<a href="https://www.thermofisher.com/blog/learning-at-the-bench/gene-editing_bid_ts_4/?CID=bid_syn_gns_r04_jp_cp0000_pjt0000_bid00000_0so_blg_op_lgn_kt_s00_gene_editing_bid_ts_3_Social_LAB" rel="noopener" target="_blank">第4回</a>では、トランスフェクションのポイントについてご紹介します。</p> <p>＜ゲノム編集実験シリーズ＞<br /> <a href="https://www.thermofisher.com/blog/learning-at-the-bench/gene-editing_bid_ts_1/?CID=bid_syn_gns_r04_jp_cp0000_pjt0000_bid00000_0so_blg_op_lgn_kt_s00_gene_editing_bid_ts_3_Social_LAB" target="_blank" rel="noopener">そういうことだったのか ! ゲノム編集実験（CRISPR/Cas9）～第1回　CRISPR/Cas9システムの原理～</a><br /> <a href="https://www.thermofisher.com/blog/learning-at-the-bench/gene-editing_bid_ts_2/?CID=bid_syn_gns_r04_jp_cp0000_pjt0000_bid00000_0so_blg_op_lgn_kt_s00_gene_editing_bid_ts_3_Social_LAB" target="_blank" rel="noopener">そういうことだったのか ! ゲノム編集実験（CRISPR/Cas9）～第2回　ガイドRNAのデザイン～</a><br /> そういうことだったのか ! ゲノム編集実験（CRISPR/Cas9）～第3回　3つのCRIPSR/Cas9実験系～<br /> <a href="https://www.thermofisher.com/blog/learning-at-the-bench/gene-editing_bid_ts_4/?CID=bid_syn_gns_r04_jp_cp0000_pjt0000_bid00000_0so_blg_op_lgn_kt_s00_gene_editing_bid_ts_3_Social_LAB" target="_blank" rel="noopener">そういうことだったのか ! ゲノム編集実験（CRISPR/Cas9）～第4回　トランスフェクション～</a><br /> <a href="https://www.thermofisher.com/blog/learning-at-the-bench/gene-editing_bid_ts_5/?CID=bid_syn_gns_r04_jp_cp0000_pjt0000_bid00000_0so_blg_op_lgn_kt_s00_gene_editing_bid_ts_3_Social_LAB" target="_blank" rel="noopener">そういうことだったのか ! ゲノム編集実験（CRISPR/Cas9）～第5回　変異の確認方法～</a><br /> <a href="https://www.thermofisher.com/blog/learning-at-the-bench/gene-editing_bid_ts_6/?CID=bid_syn_gns_r04_jp_cp0000_pjt0000_bid00000_0so_blg_op_lgn_kt_s00_gene_editing_bid_ts_3_Social_LAB" target="_blank" rel="noopener">そういうことだったのか ! ゲノム編集実験（CRISPR/Cas9）～第6回　オフターゲット効果～</a><br /> <a href="https://www.thermofisher.com/blog/learning-at-the-bench/gene-editing_bid_ts_7/?CID=bid_syn_gns_r04_jp_cp0000_pjt0000_bid00000_0so_blg_op_lgn_kt_s00_gene_editing_bid_ts_3_Social_LAB" target="_blank" rel="noopener">そういうことだったのか ! ゲノム編集実験（CRISPR/Cas9）～第7回　ノックイン～</a></p> <p>＜ゲノム編集の基礎的な内容＞<br /> <a href="https://www.thermofisher.com/blog/learning-at-the-bench/crispr-cas9_basic_bid_ts_1/?CID=bid_syn_gns_r04_jp_cp0000_pjt0000_bid00000_0so_blg_op_lgn_kt_s00_gene_editing_bid_ts_3_Social_LAB" target="_blank" rel="noopener">ゲノム編集の原理と手引き</a></p> <h3>CRISPR/Cas9 スターターパックで実際にゲノム編集を試してみよう！！</h3> <p><a class="btn btn-red" href="https://www.thermofisher.com/jp/ja/home/technical-resources/trainings/starter-pack.html?CID=bid_syn_gns_r04_jp_cp0000_pjt0000_bid00000_0so_blg_op_lgn_kt_s00_gene_editing_bid_ts_3_Social_LAB" target="_blank" rel="noopener noreferrer">詳しくはこちら</a></p> <p> </p> <p>研究用にのみ使用できます。診断用には使用いただけません。</p> <aside class="mashsb-container mashsb-main "><div class="mashsb_above_buttons">Share this article </div><div class="mashsb-box"><div class="mashsb-buttons"><a class="mashicon-facebook mash-large mashsb-noshadow" href="https://www.facebook.com/sharer.php?u=https%3A%2F%2Fwww.thermofisher.com%2Fblog%2Flearning-at-the-bench%2Fgene-editing_bid_ts_3%2F" target="_top" 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そういうことだったのか ! ゲノム編集実験（CRISPR/Cas9） ～第3回 3つのCRIPSR/Cas9実験系～ - Learning at the Bench

そういうことだったのか ! ゲノム編集実験（CRISPR/Cas9）～第3回　3つのCRIPSR/Cas9実験系～ - Learning at the Bench