LEAPER plasmid sequence "Armstrong"

Inserted LEAPER gRNA sequence: CTTTAAAGAACAAAAGAACCCTTAGTAGTGTTGACACCGACGTAAAGTGGTTCTTACACCAAATGTCAGT

https://www.addgene.org/91906/

LOCUS Exported 2978 bp ds-DNA circular SYN 25-FEB-2021 DEFINITION Backbone for cloning LwCas13a guides under U6 promoter. ACCESSION . VERSION . KEYWORDS pC016 - LwCas13a guide expression backbone with U6 promoter SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 2978) AUTHORS Abudayyeh OO, Gootenberg JS, Essletzbichler P, Han S, Joung J, Belanto JJ, Verdine V, Cox DBT, Kellner MJ, Regev A, Lander ES, Voytas DF, Ting AY, Zhang F TITLE RNA targeting with CRISPR-Cas13. JOURNAL Nature. 2017 Oct 4. doi: 10.1038/nature24049. PUBMED 28976959 REFERENCE 2 (bases 1 to 2978) AUTHORS . TITLE Direct Submission JOURNAL Exported Thursday, Feb 25, 2021 from SnapGene 5.2.4 https://www.snapgene.com COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Nature. 2017 Oct 4. doi: 10.1038/nature24049." FEATURES Location/Qualifiers source 1..2978 /organism="synthetic DNA construct" /mol_type="other DNA" rep_origin 138..726 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" /note="color: #ffff00" primer_bind 627..646 /label=pBR322ori-F /note="pBR322 origin, forward primer" /note="color: black; sequence: GGGAAACGCCTGGTATCTTT" promoter 789..1029 /label=U6 promoter /note="RNA polymerase III promoter for human U6 snRNA" /note="color: #ffffff; direction: RIGHT" primer_bind 789..809 /label=hU6-F /note="Human U6 promoter, forward primer" /note="color: black; sequence: GAGGGCCTATTTCCCATGATT" primer_bind 960..979 /label=LKO.1 5' /note="Human U6 promoter, forward primer" /note="color: black; sequence: GACTATCATATGCTTACCGT" misc_feature 1053..1122 /label=LEAPER GRNA /note="color: #a6acb3" rep_origin 1243..1698 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" /note="color: #ffff00" primer_bind complement(1330..1349) /label=F1ori-R /note="F1 origin, reverse primer" /note="color: black; sequence: AGGGAAGAAAGCGAAAGGAG" primer_bind 1540..1561 /label=F1ori-F /note="F1 origin, forward primer" /note="color: black; sequence: GTGGACTCTTGTTCCAAACTGG" primer_bind complement(1715..1734) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" /note="color: black; sequence: CGGCATCAGAGCAGATTGTA" primer_bind 1834..1856 /label=pGEX 3' /note="pGEX vectors, reverse primer" /note="color: black; sequence: CCGGGAGCTGCATGTGTCAGAGG" primer_bind complement(1894..1912) /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" /note="color: black; sequence: AATAGGCGTATCACGAGGC" promoter 1980..2084 /gene="bla" /label=AmpR promoter /note="color: #ffffff; direction: RIGHT" CDS 2085..2945 /codon_start=1 /gene="bla" /product="beta-lactamase" /label=AmpR /note="confers resistance to ampicillin, carbenicillin, and related antibiotics" /note="This feature has 2 segments: 1: 2085 .. 2153 / #ccffcc / signal sequence 2: 2154 .. 2945 / #ccffcc Cleavage site after base 2153" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" primer_bind complement(2303..2322) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" /note="color: black; sequence: ATAATACCGCGCCACATAGC" ORIGIN 1 attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat 61 ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa 121 aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca 181 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt 241 ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg 301 tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc 361 ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga 421 cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc 481 agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc 541 gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca 601 ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg 661 tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta 721 tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct 781 cagctagcga gggcctattt cccatgattc cttcatattt gcatatacga tacaaggctg 841 ttagagagat aattggaatt aatttgactg taaacacaaa gatattagta caaaatacgt 901 gacgtagaaa gtaataattt cttgggtagt ttgcagtttt aaaattatgt tttaaaatgg 961 actatcatat gcttaccgta acttgaaagt atttcgattt cttggcttta tatatcttgt 1021 ggaaaggacg aaacaccgat ttagactacc ccctttaaag aacaaaagaa cccttagtag 1081 tgttgacacc gacgtaaagt ggttcttaca ccaaatgtca gtaaaaacga aggggactaa 1141 aacgggtctt cgagaagacc ttttttttga attctgatgc ggtattttct ccttacgcat 1201 ctgtgcggta tttcacaccg catacgtcaa agcaaccata gtacgcgccc tgtagcggcg 1261 cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgcct 1321 tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc 1381 gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg 1441 accccaaaaa acttgatttg ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg 1501 tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg 1561 gaacaacact caactctatc tcgggctatt cttttgattt ataagggatt ttgccgattt 1621 cggtctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa 1681 tattaacgtt tacaatttta tggtgcactc tcagtacaat ctgctctgat gccgcatagt 1741 taagccagcc ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc 1801 cggcatccgc ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt 1861 caccgtcatc accgaaacgc gcgagacgaa agggcctcgt gatacgccta tttttatagg 1921 ttaatgtcat gataataatg gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc 1981 gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac 2041 aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt 2101 tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag 2161 aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg 2221 aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa 2281 tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc 2341 aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag 2401 tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa 2461 ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc 2521 taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg 2581 agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa 2641 caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg caacaattaa 2701 tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg 2761 gctggtttat tgctgataaa tctggagccg gtgagcgtgg aagccgcggt atcattgcag 2821 cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg 2881 caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt 2941 ggtaactgtc agaccaagtt tactcatata tactttag //