LOCUS 40924_2353 4592 bp DNA circular SYN 18-DEC-2018 DEFINITION Cloning vector P1-EYFP-Triplex-28-gRNA2-28-pA, complete sequence. ACCESSION . VERSION . KEYWORDS . SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 4592) AUTHORS Nissim L, Perli SD, Fridkin A, Perez-Pinera P, Lu TK. TITLE Multiplexed and programmable regulation of gene networks with an integrated RNA and CRISPR/Cas toolkit in human cells JOURNAL Mol. Cell 54 (4), 698-710 (2014) PUBMED 24837679 REFERENCE 2 (bases 1 to 4592) AUTHORS Perli SD. TITLE Direct Submission JOURNAL Submitted (04-MAY-2014) Electrical Engineering and Computer Science, Massachusetts Institute of Technology, 77, Massachusetts Avenue, Cambridge, MA 02139, USA REFERENCE 3 (bases 1 to 4592) TITLE Direct Submission REFERENCE 4 (bases 1 to 4592) AUTHORS . TITLE Direct Submission COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Mol. Cell"; date: "2014"; volume: "54"; issue: "4"; pages: "698-710" COMMENT SGRef: number: 2; type: "Journal Article"; journalName: "Submitted (04-MAY-2014) Electrical Engineering and Computer Science, Massachusetts Institute of Technology, 77, Massachusetts Avenue, Cambridge, MA 02139, USA" COMMENT SGRef: number: 3; type: "Journal Article" FEATURES Location/Qualifiers source 1..4592 /mol_type="other DNA" /organism="synthetic DNA construct" rep_origin complement(99..687) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" CDS complement(861..1718) /label=AmpR /note="beta-lactamase" promoter complement(1719..1823) /label=AmpR promoter rep_origin 1850..2305 /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" misc_feature 2496..2587 /label=pause site /note="RNA polymerase II transcriptional pause signal from the human alpha-2 globin gene" misc_feature 2601..2962 /label=8x gRNA1 binding sites /note="8x gRNA1 binding sites" regulatory 2969..3030 /label=Minimal Ade MLP /note="Minimal Ade MLP" /regulatory_class="promoter" CDS 3037..3738 /label=PhiYFP /note="Phialidium yellow fluorescent proteim" misc_feature 3751..3860 /label=MALAT1_3 triplex /note="MALAT1_3 triplex" misc_feature 3911..3938 /label=Csy4 rpt /note="Csy4 rpt" protein_bind 3916..3930 /label=Csy4 site /bound_moiety="Pseudomonas aeruginosa Csy4 (Cas6f)" /note="cleavage site for the Csy4 endoribonuclease (Haurwitz et al., 2010)" misc_RNA 3959..4034 /label=gRNA scaffold /note="guide RNA scaffold for the Streptococcus pyogenes CRISPR/Cas9 system" misc_feature 4043..4070 /label=Csy4 rpt /note="Csy4 rpt" protein_bind 4048..4062 /label=Csy4 site /bound_moiety="Pseudomonas aeruginosa Csy4 (Cas6f)" /note="cleavage site for the Csy4 endoribonuclease (Haurwitz et al., 2010)" polyA_signal complement(4146..4267) /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" ORIGIN 1 cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa 61 aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct 121 gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa 181 agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg 241 cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcaatgctca 301 cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa 361 ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg 421 gtaagacacg acttatcgcc actggcagca gccactggta acaggattag cagagcgagg 481 tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta cactagaagg 541 acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc 601 tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag 661 attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac 721 gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc 781 ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag 841 taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt 901 ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag 961 ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca 1021 gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact 1081 ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca 1141 gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg 1201 tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc 1261 atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg 1321 gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca 1381 tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt 1441 atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc 1501 agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc 1561 ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca 1621 tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa 1681 aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat 1741 tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa 1801 aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga cgcgccctgt 1861 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc 1921 agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc 1981 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg 2041 cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga 2101 tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc 2161 caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg 2221 ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt 2281 aacaaaatat taacgtttac aatttcccat tcgccattca ggctgcgcaa ctgttgggaa 2341 gggcgatcgg tgcgggcctc ttcgctatta cgccagccca agctaccatg ataagtaagt 2401 aatattaagg tacgggaggt acttggagcg gccgcaataa aaatctttat ctcattacat 2461 ctgtgtgttg gttttttgtg tgaatcgata gtactaacat acgctctcca tcaaaacaaa 2521 acgaaacaaa acaaactagc aaaataggct gtccccagtg caagtgcagg tgccagaaca 2581 tttctctatc gatagctagc tgaaatatta atgtcgcgac tattaatatt ttcaggctag 2641 ccatgcttcg ctacacgcga ctattaatat tttcaggcta gccatgcttc gctacacgcg 2701 actattaata ttttcaagct agccatgctt cgctacacgc gactattaat attttcaggc 2761 tagccatgct tcgctaacgc gactattaat attttcaggc tagtcatgct tcgctacacg 2821 cgactattaa tattttcagg ctagccatgc ttcgctacac gcgactatta atattttcag 2881 gctagccatg cttcgctaca cgcgactatt aatattttca ggctagccat gcttcgctac 2941 acgcgactat taatattttc aggctagcgg ggggctataa aagggggtgg gggcgttcgt 3001 cctgctatct agcgtcgcgt tgaccatggc gccaccatga gcagcggcgc cctgctgttc 3061 cacggcaaga tcccctacgt ggtggagatg gagggcgatg tggatggcca caccttcagc 3121 atccgcggta agggctacgg cgatgccagc gtgggcaagg tggatgccca gttcatctgc 3181 accaccggcg atgtgcccgt gccctggagc accctggtga ccaccctgac ctacggcgcc 3241 cagtgcttcg ccaagtacgg ccccgagctg aaggatttct acaagagctg catgcccgat 3301 ggctacgtgc aggagcgcac catcaccttc gagggcgatg gcaatttcaa gacccgcgcc 3361 gaggtgacct tcgagaatgg cagcgtgtac aatcgcgtga agctgaatgg ccagggcttc 3421 aagaaggatg gccacgtgct gggcaagaat ctggagttca atttcacccc ccactgcctg 3481 tacatctggg gcgatcaggc caatcacggc ctgaagagcg ccttcaagat ctgccacgag 3541 atcgccggca gcaagggcga tttcatcgtg gccgatcaca cccagatgaa tacccccatc 3601 ggcggcggcc ccgtgcacgt gcccgagtac caccacatga gctaccacgt gaagctgagc 3661 aaggatgtga ccgatcaccg cgataatatg agcctgacgg agaccgtgcg cgccgtggat 3721 tgccgcaaga cctacctgtg ataaaccggt gattcgtcag tagggttgta aaggtttttc 3781 ttttcctgag aaaacaacct tttgttttct caggttttgc tttttggcct ttccctagct 3841 ttaaaaaaaa aaaagcaaaa ctcaccgagg cagttccata ggatggcaag atcctggtat 3901 cggtctgcga gttcactgcc gtataggcag ctaagaaagt aagtcggagt actgtcctgt 3961 tttagagcta gaaatagcaa gttaaaataa ggctagtccg ttatcaactt gaaaaagtgg 4021 caccgagtcg gtgctttttt tcgttcactg ccgtataggc agctaagaaa caaacaggaa 4081 tcgaatgcaa ccggcgcagg aacactgcca gcgcatcaac ggccggccgc ttcgagcaga 4141 catgataaga tacattgatg agtttggaca aaccacaact agaatgcagt gaaaaaaatg 4201 ctttatttgt gaaatttgtg atgctattgc tttatttgta accattataa gctgcaataa 4261 acaagttaac aacaacaatt gcattcattt tatgtttcag gttcaggggg aggtgtggga 4321 ggttttttaa agcaagtaaa acctctacaa atgtggtaaa atcgataagg atcgagtatg 4381 tcgaccgatg cccttgagag ccttcaaccc agtcagctcc ttccggtggg cgcggggcat 4441 gactatcgtc gccgcactta tgactgtctt ctttatcatg caactcgtag gacaggtgcc 4501 ggcagcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc 4561 gagcggtatc agctcactca aaggcggtaa ta //