LOCUS 40924_505 3928 bp DNA circular SYN 17-DEC-2018 DEFINITION Cloning vector CMVp-HHRibo-gRNA1-HDVRibo-pA, complete sequence. ACCESSION . VERSION . KEYWORDS . SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 3928) 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 3928) 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 3928) TITLE Direct Submission REFERENCE 4 (bases 1 to 3928) 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..3928 /mol_type="other DNA" /organism="synthetic DNA construct" promoter complement(620..724) /label=AmpR promoter rep_origin 751..1206 /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" enhancer 1395..1698 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 1699..1902 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" ncRNA 1936..1972 /label=hammerhead ribozyme misc_RNA 1993..2068 /label=gRNA scaffold /note="guide RNA scaffold for the Streptococcus pyogenes CRISPR/Cas9 system" ncRNA 2073..2140 /label=HDV ribozyme polyA_signal 2389..2436 /label=HSV TK poly(A) signal /note="herpes simplex virus thymidine kinase polyadenylation signal (Cole and Stacy, 1985)" rep_origin complement(2928..3516) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" CDS complement(join(3690..3928,1..619)) /codon_start=1 /label=AmpR /note="beta-lactamase" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" ORIGIN 1 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg 61 ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca 121 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg 181 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca 241 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg 301 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct 361 cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca 421 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca 481 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg 541 tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac 601 ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt 661 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc 721 cgcgcacatt tccccgaaaa gtgccacctg acgcgccctg tagcggcgca ttaagcgcgg 781 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc 841 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa 901 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac 961 ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt 1021 tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca 1081 accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt 1141 taaaaaatga gctgatttaa caaaaattta acgcgaattt taacaaaata ttaacgttta 1201 caatttccca ttcgccattc aggctgcgca actgttggga agggcgatcg gtgcgggcct 1261 cttcgctatt acgccagccc aagctaccat gataagtaag taatattaag gtacgggagg 1321 tacttggagg tacctagtta ttaatagtaa tcaattacgg ggtcattagt tcatagccca 1381 tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac 1441 gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact 1501 ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa 1561 gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg 1621 cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta 1681 gtcatcgcta ttaccatggt gatgcggttt tggcagtaca tcaatgggcg tggatagcgg 1741 tttgactcac ggggatttcc aagtctccac cccattgacg tcaatgggag tttgttttgg 1801 caccaaaatc aacgggactt tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg 1861 ggcggtaggc gtgtacggtg ggaggtctat ataagcagag ctggtttatg aaccgtcaga 1921 tccgagctcc gactactgat gagtccgtga ggacgaaacg agtaagctcg tctagtcgcg 1981 tgtagcgaag cagttttaga gctagaaata gcaagttaaa ataaggctag tccgttatca 2041 acttgaaaaa gtggcaccga gtcggtgctt ttggccggca tggtcccagc ctcctcgctg 2101 gcgccggctg ggcaacatgc ttcggcatgg cgaatgggac cccgggatgc tagctaagcg 2161 ggactctggg gttcgaaatg accgaccaag cgacgcccaa cctgccatca cgagatttcg 2221 attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct 2281 ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccaccct agggggaggc 2341 taactgaaac acggaaggag acaataccgg aaggaacccg cgctatgacg gcaataaaaa 2401 gacagaataa aacgcacggt gttgggtcgt ttgttcataa acgcggggtt cggtcccagg 2461 gctggcactc tgtcgatacc ccaccgagac gccattgggg ccaatacgcc cgcgtttctt 2521 ccttttcccc accccacccc ccaagttcgg gtgaaggccc agggctcgca gccaacgtcg 2581 gggcggcagg ccctgccata gcctcagctc gagtatgtcg accgatgccc ttgagagcct 2641 tcaacccagt cagctccttc cggtgggcgc ggggcatgac tatcgtcgcc gcacttatga 2701 ctgtcttctt tatcatgcaa ctcgtaggac aggtgccggc agcgctcttc cgcttcctcg 2761 ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag 2821 gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa 2881 ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc 2941 cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca 3001 ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg 3061 accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct 3121 caatgctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt 3181 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag 3241 tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc 3301 agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac 3361 actagaagga cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga 3421 gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc 3481 aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg 3541 gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca 3601 aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt 3661 atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca 3721 gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg 3781 atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca 3841 ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt 3901 cctgcaactt tatccgcctc catccagt //