{"id":195,"date":"2012-10-16T13:44:12","date_gmt":"2012-10-16T12:44:12","guid":{"rendered":"http:\/\/blogs.lshtm.ac.uk\/alsfordlab\/?page_id=195"},"modified":"2015-06-30T23:08:25","modified_gmt":"2015-06-30T22:08:25","slug":"stem-loop-rnai-cassette","status":"publish","type":"page","link":"https:\/\/blogs.lshtm.ac.uk\/alsfordlab\/cell-lines-plasmids\/prpa-series-for-2t1-landing-pad-cell-line\/stem-loop-rnai-cassette\/","title":{"rendered":"Stem-loop RNAi cassette"},"content":{"rendered":"<p><span style=\"font-family: arial\">Cassettes sequenced in full, February 2008<\/span><\/p>\n<p><span style=\"font-family: arial\">Two versions are available in the <a title=\"pRPa series (for 2T1 \u2013 \u2018landing pad\u2019 cell line)\" href=\"http:\/\/blogs.lshtm.ac.uk\/alsfordlab\/cell-lines-plasmids\/prpa-series-for-2t1-landing-pad-cell-line\/\">pRPa<\/a> backbone, containing different multiple cloning sites separated by a 468 bp stuffer (pBluescript 117-585, including 125 bp of the \u03b2-galactosidase \u03b1-fragment)<\/span><\/p>\n<p><em>NB: The <a title=\"pRP series\" href=\"http:\/\/blogs.lshtm.ac.uk\/alsfordlab\/cell-lines-plasmids\/prp-series\/\">pRP<\/a> version of the stem-loop RNAi construct is only available with a <\/em>HYG<em> selectable marker and the MCS1\/2 RNAi expression cassette<\/em><\/p>\n<p><span style=\"font-family: arial\">All of the restriction sites listed are unique.<\/span><\/p>\n<p><span style=\"font-family: arial\"><b>MCS<sup>1\/2<\/sup><\/b><\/span><\/p>\n<p><span><i>Acc<\/i>65<\/span><span style=\"font-family: arial\">I <i>Sma<\/i>I <i>Afl<\/i>II <i>Bam<\/i>HI<\/span><\/p>\n<pre>   1  GGTACCCGGG CTTAAGGATC CGAGATAGGG TTGAGTGTTG TTCCAGTTTG\r\n  51  GAACAAGAGT CCACTATTAA AGAACGTGGA CTCCAACGTC AAAGGGCGAA\r\n 101  AAACCGTCTA TCAGGGCGAT GGCCCACTAC GTGAACCATC ACCCTAATCA\r\n 151  AGTTTTTTGG GGTCGAGGTG CCGTAAAGCA CTAAATCGGA ACCCTAAAGG\r\n 201  GAGCCCCCGA TTTAGAGCTT GACGGGGAAA GCCGGCGAAC GTGGCGAGAA\r\n 251  AGGAAGGGAA GAAAGCGAAA GGAGCGGGCG CTAGGGCGCT GGCAAGTGTA\r\n 301  GCGGTCACGC TGCGCGTAAC CACCACACCC GCCGCGCTTA ATGCGCCGCT\r\n 351  ACAGGGCGCG TCCATTCGCC ATTCAGGCTG CGCAACTGTT GGGAAGGGCG\r\n 401  ATCGGTGCGG GCCTCTTCGC TATTACGCCA GCTGGCGAAA GGGGGATGTG\r\n 451  CTGCAAGGCG ATTAAGTTGG GTAACGCCAG GGTTTTCTAG AAGCTTCTCG\r\n 501  AGGGCCC<\/pre>\n<p><span style=\"font-family: arial\"><i>Xba<\/i>I <i>Hin<\/i>dIII <i>Xho<\/i>I <\/span><span><i>Bsp<\/i>120I<\/span><\/p>\n<p>&nbsp;<\/p>\n<p><span style=\"font-family: arial\"><b>MCS<sup>3\/4<\/sup><\/b><\/span><\/p>\n<p><span style=\"font-family: arial\">Acc65I <i>Age<\/i>I <i>Cla<\/i>I <i>Mfe<\/i>I <i>Sma<\/i>I<br \/>\n<\/span><span style=\"font-family: arial\">Note: <i>Age<\/i>I &amp; <i>Xma<\/i>I (isoschizomer of <i>Sma<\/i>I) give compatible overhangs; <i>Cla<\/i>I site blocked by Dam methylation<\/span><\/p>\n<pre>   1  GGTACCGGTA TCGATCAATT GCCCGGGTCC GAGATAGGGT TGAGTGTTGT\r\n  51  TCCAGTTTGG AACAAGAGTC CACTATTAAA GAACGTGGAC TCCAACGTCA\r\n 101  AAGGGCGAAA AACCGTCTAT CAGGGCGATG GCCCACTACG TGAACCATCA\r\n 151  CCCTAATCAA GTTTTTTGGG GTCGAGGTGC CGTAAAGCAC TAAATCGGAA\r\n 201  CCCTAAAGGG AGCCCCCGAT TTAGAGCTTG ACGGGGAAAG CCGGCGAACG\r\n 251  TGGCGAGAAA GGAAGGGAAG AAAGCGAAAG GAGCGGGCGC TAGGGCGCTG\r\n 301  GCAAGTGTAG CGGTCACGCT GCGCGTAACC ACCACACCCG CCGCGCTTAA\r\n 351  TGCGCCGCTA CAGGGCGCGT CCATTCGCCA TTCAGGCTGC GCAACTGTTG\r\n 401  GGAAGGGCGA TCGGTGCGGG CCTCTTCGCT ATTACGCCAG CTGGCGAAAG\r\n 451  GGGGATGTGC TGCAAGGCGA TTAAGTTGGG TAACGCCAGG GTTTTCAGAT\r\n 501  CTTAAGGCCG GCCTCGAGTC TAGAGGGCCC<\/pre>\n<p><span style=\"font-family: arial\"><i>Bgl<\/i>II <i>Afl<\/i>II <i>Fse<\/i>I <i>Xho<\/i>I <i>Xba<\/i>I <i>Bsp<\/i>120I<\/span><\/p>\n","protected":false},"excerpt":{"rendered":"<p>Cassettes sequenced in full, February 2008 Two versions are available in the pRPa backbone, containing&#8230;<\/p>\n","protected":false},"author":116,"featured_media":0,"parent":188,"menu_order":0,"comment_status":"closed","ping_status":"closed","template":"onecolumn-page.php","meta":{"footnotes":""},"class_list":["post-195","page","type-page","status-publish","hentry","odd"],"yoast_head":"<!-- This site is optimized with the Yoast SEO plugin v26.9 - https:\/\/yoast.com\/product\/yoast-seo-wordpress\/ -->\n<title>Stem-loop RNAi cassette - Alsford Lab<\/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:\/\/blogs.lshtm.ac.uk\/alsfordlab\/cell-lines-plasmids\/prpa-series-for-2t1-landing-pad-cell-line\/stem-loop-rnai-cassette\/\" \/>\n<meta property=\"og:locale\" content=\"en_GB\" \/>\n<meta property=\"og:type\" content=\"article\" \/>\n<meta property=\"og:title\" content=\"Stem-loop RNAi cassette - Alsford Lab\" \/>\n<meta property=\"og:description\" content=\"Cassettes sequenced in full, February 2008 Two versions are available in the pRPa backbone, containing...\" \/>\n<meta property=\"og:url\" content=\"https:\/\/blogs.lshtm.ac.uk\/alsfordlab\/cell-lines-plasmids\/prpa-series-for-2t1-landing-pad-cell-line\/stem-loop-rnai-cassette\/\" \/>\n<meta property=\"og:site_name\" content=\"Alsford Lab\" \/>\n<meta property=\"article:modified_time\" content=\"2015-06-30T22:08:25+00:00\" \/>\n<meta name=\"twitter:label1\" content=\"Estimated reading time\" \/>\n\t<meta name=\"twitter:data1\" content=\"1 minute\" \/>\n<script type=\"application\/ld+json\" class=\"yoast-schema-graph\">{\"@context\":\"https:\/\/schema.org\",\"@graph\":[{\"@type\":\"WebPage\",\"@id\":\"https:\/\/blogs.lshtm.ac.uk\/alsfordlab\/cell-lines-plasmids\/prpa-series-for-2t1-landing-pad-cell-line\/stem-loop-rnai-cassette\/\",\"url\":\"https:\/\/blogs.lshtm.ac.uk\/alsfordlab\/cell-lines-plasmids\/prpa-series-for-2t1-landing-pad-cell-line\/stem-loop-rnai-cassette\/\",\"name\":\"Stem-loop RNAi cassette - Alsford Lab\",\"isPartOf\":{\"@id\":\"https:\/\/blogs.lshtm.ac.uk\/alsfordlab\/#website\"},\"datePublished\":\"2012-10-16T12:44:12+00:00\",\"dateModified\":\"2015-06-30T22:08:25+00:00\",\"breadcrumb\":{\"@id\":\"https:\/\/blogs.lshtm.ac.uk\/alsfordlab\/cell-lines-plasmids\/prpa-series-for-2t1-landing-pad-cell-line\/stem-loop-rnai-cassette\/#breadcrumb\"},\"inLanguage\":\"en-GB\",\"potentialAction\":[{\"@type\":\"ReadAction\",\"target\":[\"https:\/\/blogs.lshtm.ac.uk\/alsfordlab\/cell-lines-plasmids\/prpa-series-for-2t1-landing-pad-cell-line\/stem-loop-rnai-cassette\/\"]}]},{\"@type\":\"BreadcrumbList\",\"@id\":\"https:\/\/blogs.lshtm.ac.uk\/alsfordlab\/cell-lines-plasmids\/prpa-series-for-2t1-landing-pad-cell-line\/stem-loop-rnai-cassette\/#breadcrumb\",\"itemListElement\":[{\"@type\":\"ListItem\",\"position\":1,\"name\":\"Cell Lines &amp; Plasmids\",\"item\":\"https:\/\/blogs.lshtm.ac.uk\/alsfordlab\/cell-lines-plasmids\/\"},{\"@type\":\"ListItem\",\"position\":2,\"name\":\"pRPa series\",\"item\":\"https:\/\/blogs.lshtm.ac.uk\/alsfordlab\/cell-lines-plasmids\/prpa-series-for-2t1-landing-pad-cell-line\/\"},{\"@type\":\"ListItem\",\"position\":3,\"name\":\"Stem-loop RNAi cassette\"}]},{\"@type\":\"WebSite\",\"@id\":\"https:\/\/blogs.lshtm.ac.uk\/alsfordlab\/#website\",\"url\":\"https:\/\/blogs.lshtm.ac.uk\/alsfordlab\/\",\"name\":\"Alsford Lab\",\"description\":\"Trypanosome molecular and cellular biology\",\"potentialAction\":[{\"@type\":\"SearchAction\",\"target\":{\"@type\":\"EntryPoint\",\"urlTemplate\":\"https:\/\/blogs.lshtm.ac.uk\/alsfordlab\/?s={search_term_string}\"},\"query-input\":{\"@type\":\"PropertyValueSpecification\",\"valueRequired\":true,\"valueName\":\"search_term_string\"}}],\"inLanguage\":\"en-GB\"}]}<\/script>\n<!-- \/ Yoast SEO plugin. -->","yoast_head_json":{"title":"Stem-loop RNAi cassette - Alsford Lab","robots":{"index":"index","follow":"follow","max-snippet":"max-snippet:-1","max-image-preview":"max-image-preview:large","max-video-preview":"max-video-preview:-1"},"canonical":"https:\/\/blogs.lshtm.ac.uk\/alsfordlab\/cell-lines-plasmids\/prpa-series-for-2t1-landing-pad-cell-line\/stem-loop-rnai-cassette\/","og_locale":"en_GB","og_type":"article","og_title":"Stem-loop RNAi cassette - Alsford Lab","og_description":"Cassettes sequenced in full, February 2008 Two versions are available in the pRPa backbone, containing...","og_url":"https:\/\/blogs.lshtm.ac.uk\/alsfordlab\/cell-lines-plasmids\/prpa-series-for-2t1-landing-pad-cell-line\/stem-loop-rnai-cassette\/","og_site_name":"Alsford Lab","article_modified_time":"2015-06-30T22:08:25+00:00","twitter_misc":{"Estimated reading time":"1 minute"},"schema":{"@context":"https:\/\/schema.org","@graph":[{"@type":"WebPage","@id":"https:\/\/blogs.lshtm.ac.uk\/alsfordlab\/cell-lines-plasmids\/prpa-series-for-2t1-landing-pad-cell-line\/stem-loop-rnai-cassette\/","url":"https:\/\/blogs.lshtm.ac.uk\/alsfordlab\/cell-lines-plasmids\/prpa-series-for-2t1-landing-pad-cell-line\/stem-loop-rnai-cassette\/","name":"Stem-loop RNAi cassette - Alsford Lab","isPartOf":{"@id":"https:\/\/blogs.lshtm.ac.uk\/alsfordlab\/#website"},"datePublished":"2012-10-16T12:44:12+00:00","dateModified":"2015-06-30T22:08:25+00:00","breadcrumb":{"@id":"https:\/\/blogs.lshtm.ac.uk\/alsfordlab\/cell-lines-plasmids\/prpa-series-for-2t1-landing-pad-cell-line\/stem-loop-rnai-cassette\/#breadcrumb"},"inLanguage":"en-GB","potentialAction":[{"@type":"ReadAction","target":["https:\/\/blogs.lshtm.ac.uk\/alsfordlab\/cell-lines-plasmids\/prpa-series-for-2t1-landing-pad-cell-line\/stem-loop-rnai-cassette\/"]}]},{"@type":"BreadcrumbList","@id":"https:\/\/blogs.lshtm.ac.uk\/alsfordlab\/cell-lines-plasmids\/prpa-series-for-2t1-landing-pad-cell-line\/stem-loop-rnai-cassette\/#breadcrumb","itemListElement":[{"@type":"ListItem","position":1,"name":"Cell Lines &amp; Plasmids","item":"https:\/\/blogs.lshtm.ac.uk\/alsfordlab\/cell-lines-plasmids\/"},{"@type":"ListItem","position":2,"name":"pRPa series","item":"https:\/\/blogs.lshtm.ac.uk\/alsfordlab\/cell-lines-plasmids\/prpa-series-for-2t1-landing-pad-cell-line\/"},{"@type":"ListItem","position":3,"name":"Stem-loop RNAi cassette"}]},{"@type":"WebSite","@id":"https:\/\/blogs.lshtm.ac.uk\/alsfordlab\/#website","url":"https:\/\/blogs.lshtm.ac.uk\/alsfordlab\/","name":"Alsford Lab","description":"Trypanosome molecular and cellular biology","potentialAction":[{"@type":"SearchAction","target":{"@type":"EntryPoint","urlTemplate":"https:\/\/blogs.lshtm.ac.uk\/alsfordlab\/?s={search_term_string}"},"query-input":{"@type":"PropertyValueSpecification","valueRequired":true,"valueName":"search_term_string"}}],"inLanguage":"en-GB"}]}},"_links":{"self":[{"href":"https:\/\/blogs.lshtm.ac.uk\/alsfordlab\/wp-json\/wp\/v2\/pages\/195","targetHints":{"allow":["GET"]}}],"collection":[{"href":"https:\/\/blogs.lshtm.ac.uk\/alsfordlab\/wp-json\/wp\/v2\/pages"}],"about":[{"href":"https:\/\/blogs.lshtm.ac.uk\/alsfordlab\/wp-json\/wp\/v2\/types\/page"}],"author":[{"embeddable":true,"href":"https:\/\/blogs.lshtm.ac.uk\/alsfordlab\/wp-json\/wp\/v2\/users\/116"}],"replies":[{"embeddable":true,"href":"https:\/\/blogs.lshtm.ac.uk\/alsfordlab\/wp-json\/wp\/v2\/comments?post=195"}],"version-history":[{"count":11,"href":"https:\/\/blogs.lshtm.ac.uk\/alsfordlab\/wp-json\/wp\/v2\/pages\/195\/revisions"}],"predecessor-version":[{"id":1216,"href":"https:\/\/blogs.lshtm.ac.uk\/alsfordlab\/wp-json\/wp\/v2\/pages\/195\/revisions\/1216"}],"up":[{"embeddable":true,"href":"https:\/\/blogs.lshtm.ac.uk\/alsfordlab\/wp-json\/wp\/v2\/pages\/188"}],"wp:attachment":[{"href":"https:\/\/blogs.lshtm.ac.uk\/alsfordlab\/wp-json\/wp\/v2\/media?parent=195"}],"curies":[{"name":"wp","href":"https:\/\/api.w.org\/{rel}","templated":true}]}}