LOCUS Exported 8003 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION 3-color stripe (inverted reporters). ACCESSION . VERSION . KEYWORDS pJ2042.2_invRep SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 8003) AUTHORS Santos-Moreno J, Tasiudi E, Stelling J, Schaerli Y TITLE Multistable and dynamic CRISPRi-based synthetic circuits. JOURNAL Nat Commun. 2020 Jun 2;11(1):2746. doi: 10.1038/s41467-020-16574-1. PUBMED 32488086 REFERENCE 2 (bases 1 to 8003) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..8003 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 65..349 /gene="araBAD" /label=araBAD promoter /note="promoter of the L-arabinose operon of E. coli; the araC regulatory gene is transcribed in the opposite direction (Guzman et al., 1995)" primer_bind 272..291 /label=pBAD Forward /note="For vectors with E. coli araBAD promoter, forward primer" protein_bind 415..429 /label=Csy4 site /bound_moiety="Pseudomonas aeruginosa Csy4 (Cas6f)" /note="cleavage site for the Csy4 endoribonuclease (Haurwitz et al., 2010)" protein_bind 537..551 /label=Csy4 site /bound_moiety="Pseudomonas aeruginosa Csy4 (Cas6f)" /note="cleavage site for the Csy4 endoribonuclease (Haurwitz et al., 2010)" protein_bind 617..631 /label=Csy4 site /bound_moiety="Pseudomonas aeruginosa Csy4 (Cas6f)" /note="cleavage site for the Csy4 endoribonuclease (Haurwitz et al., 2010)" protein_bind 743..757 /label=Csy4 site /bound_moiety="Pseudomonas aeruginosa Csy4 (Cas6f)" /note="cleavage site for the Csy4 endoribonuclease (Haurwitz et al., 2010)" protein_bind 829..843 /label=Csy4 site /bound_moiety="Pseudomonas aeruginosa Csy4 (Cas6f)" /note="cleavage site for the Csy4 endoribonuclease (Haurwitz et al., 2010)" RBS 848..870 /note="efficient ribosome binding site from bacteriophage T7 gene 10 (Olins and Rangwala, 1989)" CDS 878..1531 /codon_start=1 /product="monomeric Kusabira-Orange 2 fluorescent protein" /label=mKO2 /translation="MVSVIKPEMKMRYYMDGSVNGHEFTIEGEGTGRPYEGHQEMTLRV TMAEGGPMPFAFDLVSHVFCYGHRVFTKYPEEIPDYFKQAFPEGLSWERSLEFEDGGSA SVSAHISLRGNTFYHKSKFTGVNFPADGPIMQNQSVDWEPSTEKITASDGVLKGDVTMY LKLEGGGNHKCQMKTTYKAAKEILEMPGDHYIGHRLVRKTEGNITEQVEDAVAHS" protein_bind 2440..2454 /label=Csy4 site /bound_moiety="Pseudomonas aeruginosa Csy4 (Cas6f)" /note="cleavage site for the Csy4 endoribonuclease (Haurwitz et al., 2010)" RBS 2459..2481 /note="efficient ribosome binding site from bacteriophage T7 gene 10 (Olins and Rangwala, 1989)" protein_bind 3913..3927 /label=Csy4 site /bound_moiety="Pseudomonas aeruginosa Csy4 (Cas6f)" /note="cleavage site for the Csy4 endoribonuclease (Haurwitz et al., 2010)" protein_bind 4039..4053 /label=Csy4 site /bound_moiety="Pseudomonas aeruginosa Csy4 (Cas6f)" /note="cleavage site for the Csy4 endoribonuclease (Haurwitz et al., 2010)" protein_bind 4125..4139 /label=Csy4 site /bound_moiety="Pseudomonas aeruginosa Csy4 (Cas6f)" /note="cleavage site for the Csy4 endoribonuclease (Haurwitz et al., 2010)" RBS 4144..4166 /note="efficient ribosome binding site from bacteriophage T7 gene 10 (Olins and Rangwala, 1989)" CDS 4174..4887 /codon_start=1 /product="GFP variant that folds robustly even when fused to poorly folded proteins (Pedelacq et al., 2006)" /label=superfolder GFP /translation="MRKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK FICTTGKLPVPWPTLVTTLTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTISFKDDG TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE FVTAAGITHGMDELYK" terminator 5237..5264 /label=rrnB T2 terminator /note="transcription terminator T2 from the E. coli rrnB gene" CDS complement(5452..6267) /codon_start=1 /gene="aph(3')-Ia" /product="aminoglycoside phosphotransferase" /label=KanR /note="confers resistance to kanamycin in bacteria or G418 (Geneticin(R)) in eukaryotes" /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF" primer_bind 6175..6194 /label=Kan-R /note="Kanamycin resistance gene, reverse primer" promoter complement(6268..6359) /gene="bla" /label=AmpR promoter rep_origin complement(6377..7012) /direction=LEFT /label=ColA ori /note="Plasmids containing the ColA origin of replication can be propagated in E. coli cells that contain additional plasmids with compatible origins." primer_bind complement(6522..6541) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(7163..38) /codon_start=1 /gene="araC" /product="L-arabinose regulatory protein" /label=araC /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE EKVNDVAVKLS" ORIGIN 1 tatcccggca gcaggggatc attttgcgct tcagccatac ttttcatact cccgccattc 61 agagaagaaa ccaattgtcc atattgcatc agacattgcc gtcactgcgt cttttactgg 121 ctcttctcgc taaccaaacc ggtaaccccg cttattaaaa gcattctgta acaaagcggg 181 accaaagcca tgacaaaaac gcgtaacaaa agtgtctata atcacggcag aaaagtccac 241 attgattatt tgcacggcgt cacactttgc tatgccatag catttttatc cataagatta 301 gcggttccta cctgacgctt tttatcgcaa ctctctactg tttctccata ccgaattcat 361 aggatagatt ctggaaactt taccgtccga gctccagcct gcggtccggg ttcactgccg 421 tataggcaga agctagactc tagtggtttc agagctatgc tggaaacagc atagcaagtt 481 gaaataaggc tagtccgtta tcaacttgaa aaagtggcac cgagtcggtg cgttcactgc 541 cgtataggca gtcgctggga cgcccgggga ctacacttac gaaactattg attgctcagc 601 ctgcggtccg ggttcactgc cgtataggca gatcagtgtg tactaagtac tgtttcagag 661 ctatgctgga aacagcatag caagttgaaa taaggctagt ccgttatcaa cttgaaaaag 721 tggcaccgag tcggtgcgtt cactgccgta taggcagtcg ctgggacgcc cgctcgagca 781 ataaacagtt gatagggctt ctccgttaca gcctgcggtc cgggttcact gccgtatagg 841 cagtaatttt gtttaacttt aagaaggaga tatacatatg gtttcggtta tcaaaccaga 901 gatgaaaatg cgttactata tggatggttc agtaaatggt cacgaattta ctattgaggg 961 cgagggtacg ggacgcccat acgaggggca ccaggaaatg actttacgcg tcacaatggc 1021 tgaaggcggg cctatgccgt ttgcgttcga tcttgttagt catgtctttt gttacggtca 1081 ccgtgtattt actaaatacc ccgaggaaat tccagactat ttcaaacaag ccttcccgga 1141 aggtttgtct tgggagcgca gtttagagtt tgaagacggt ggctcggcca gcgtgtcagc 1201 tcatattagt cttcgcggca atacatttta tcacaagtca aagttcaccg gcgtgaactt 1261 ccccgcagac ggcccaatca tgcagaatca aagtgttgat tgggaaccgt ccacagagaa 1321 gattacagct tccgatggag tcttaaaggg cgatgtaacc atgtacttaa aattagaagg 1381 gggagggaac cataaatgtc agatgaagac tacctataag gccgcaaaag agattcttga 1441 aatgcccgga gaccactaca ttgggcatcg tttggtccgt aagacagaag gaaatattac 1501 tgaacaggtc gaagacgctg tggcacacag catgtcccgc cgtaatactg acgccatcac 1561 aatccacagc atcctggatt ggattgaaga caacttggag tcgccgttga gtttagaaaa 1621 agttagtgaa cgtagtggtt actcaaagtg gcaccttcag cgcatgttta agaaggaaac 1681 gggtcattca ttgggtcaat atattcgttc tcgcaagatg actgaaattg cccagaaatt 1741 gaaagagtct aatgaaccta ttttgtacct ggcggagcgt tacggctttg aaagtcagca 1801 aacccttaca cgtaccttca agaattactt tgacgttcca ccacacaaat atcgtatgac 1861 caacatgcag ggtgagtcac gttttttgca tccgttgaat cattacaatt cctaataatc 1921 gctgggacgc ccgccatggt tcagccaaaa aacttaagac cgccggtctt gtccactacc 1981 ttgcagtaat gcggtggaca ggatcggcgg ttttcttttc tcttctcaat tcttctgacc 2041 tgtaacgaat aatagatagt aaagtagtct ccgattgagt tttctctgcc gagtcccacc 2101 cagttctgtg atttcagtaa gttggtaatt gatacactgt tgcgagaact gctgcctggt 2161 agtagatagg ttgttattga gtaagaaggt aaagtgaacg aaatccctga aactgagact 2221 gtagaaaata agcttcagcc tgcggtccgg ttgacggcta gctcagtcct aggtacagtg 2281 ctagctcgct gggacgcccg agatagccgt tacacaggtg acacttattt cagcctgcgg 2341 tccggccaca ctagagtcta gcttgagatc gctgggacgc ccgggatcca agagatttct 2401 acacgattga gcactgtctc agcctgcggt ccgggttcac tgccgtatag gcagtaattt 2461 tgtttaactt taagaaggag atatacatat gaatcagtca ttcatctcgg acatcttata 2521 tgccgacatc gaatcgaagg ctaaggaact tacagtcaat tccaacaata ctgtccagcc 2581 ggtcgcgctt atgcgcttag gagttttcgt tcccaaacct tccaagagca aaggagaaag 2641 taaggaaatt gacgccacca aagccttctc tcaactggag attgctaaag cagagggcat 2701 ggttagtaaa ggagaagaaa ataacatggc actgattaag gagaacatgc acatgaagct 2761 gtacatggag ggcaccgtga acaaccacca cttcaagtgc acatccgagg gcgaaggcaa 2821 gccctacgag ggcacccaga ccatgagaat caaggccgtc gagggcggcc ctctcccctt 2881 cgccttcgac atcctggcta ccagcttcat gtacggcagc aaaaccttca tcaaccacac 2941 ccagggcatc cccgacttct ttaagcagtc cttccctgag ggcttcacat gggagagagt 3001 caccacatac gaagacgggg gcgtgctgac cgctacccag gacaccagcc tccaggacgg 3061 ctgcctcatc tacaacgtca agatcagagg ggtgaacttc ccatccaacg gccctgtgat 3121 gcagaagaaa acactcggct gggaggcctc caccgagacc ctgtaccccg ctgacggcgg 3181 cctggaaggc agagccgaca tggccctgaa gctcgtgggc gggggccacc tgatctgcaa 3241 cttgaagacc acatacagat ccaagaaacc cgctaagaac ctcaagatgc ccggcgtcta 3301 ctatgtggac agaagactgg aaagaatcaa ggaggccgac aaagagacct acgtcgagca 3361 gcacgaggtg gctgtggcca gatactgcga cctccctagc aaactggggc acagagtcta 3421 ataatcgctg ggacgcccgc ctgcaggctc ggtaccaaat tccagaaaag aggcctcccg 3481 aaaggggggc cttttttcgt tttggtccta atagataaag gataggtctg gtagtgttgt 3541 tcgttctcgc aggtaaatca ataatactca gcagttccgt agacttttca gtgggacagg 3601 gtagcgataa cagatagatt gtaataagac acagtaggtg ctcgtagttg cgtgaagaga 3661 accgctcagg aaatccagtc agaagtattg gtaatcgttg aaaactcagt cgaccagcct 3721 gcggtccggt ttacggctag ctcagtccta ggtattatgc tagctcgctg ggacgcccga 3781 gtgacgactg cgaagtaacc tctatttatc agcctgcggt ccggccaagt acttagtaca 3841 cactgattcg ctgggacgcc cggtagtgct tatcagaccc aatactgttg aacagcctgc 3901 ggtccgggtt cactgccgta taggcagtct caagctagac tctagtggtt tcagagctat 3961 gctggaaaca gcatagcaag ttgaaataag gctagtccgt tatcaacttg aaaaagtggc 4021 accgagtcgg tgcgttcact gccgtatagg cagtcgctgg gacgcccgga cgtcctatta 4081 cactcgtcgt tggaaactga agatcagcct gcggtccggg ttcactgccg tataggcagt 4141 aattttgttt aactttaaga aggagatata catatgcgta aaggcgaaga actgtttacc 4201 ggtgtggttc cgattctggt ggaactggac ggcgatgtta atggtcataa attcagtgtt 4261 cgcggcgaag gtgaaggcga tgcgacgaac ggcaaactga ccctgaaatt tatctgcacc 4321 acgggtaaac tgccggtccc gtggccgacg ctggtgacca cgctgaccta tggcgttcaa 4381 tgttttgcgc gttacccgga tcacatgaaa cagcacgact ttttcaaatc ggccatgccg 4441 gaaggctatg tgcaggaacg tacgattagc tttaaagacg atggtacgta taaaacccgc 4501 gcggaagtga aattcgaagg cgataccctg gttaaccgta tcgaactgaa aggtatcgat 4561 ttcaaagaag acggcaatat tctgggtcat aaactggaat ataacttcaa ttcccacaac 4621 gtgtacatca ccgcggataa acagaaaaac ggcattaaag ccaatttcaa aatccgccat 4681 aatgtggaag atggtagcgt tcagctggcc gaccactatc agcaaaacac gccgattggt 4741 gatggcccgg tcctgctgcc ggacaatcac tacctgagta cccagtccgt gctgtcaaaa 4801 gatccgaacg aaaaacgtga ccacatggtc ctgctggaat ttgtgacggc tgcgggtatc 4861 acccacggca tggacgaact gtataaaatg tcccgccgta atactgacgc catcacaatc 4921 cacagcatcc tggattggat tgaagactaa taatcgctgg gacgcccggc ggccgcggaa 4981 acacagaaaa aagcccgcac ctgacagtgc gggctttttt tttcgaccaa aggtagcgaa 5041 cgacgagtca ctgttgagga taaatacttt ctctactagg cgcctgttac acaggtcctc 5101 agcggcgcgc ctttgtcggt gaacgctctc ctgagtagga caaatccgcc gggagcggat 5161 ttgaacgttg tgaagcaacg gcccggaggg tggcgggcag gacgcccgcc ataaactgcc 5221 aggcatcaaa ctaagcagaa ggccatcctg acggatggcc tttttgcgtt tcagatctac 5281 cggtaaacca gcaatagaca taagcggcta tttaacgacc ctgccctgaa ccgacgacaa 5341 gctgacgacc gggtctccgc aagtggcact tttcggggaa atgtgcgcgg aacccctatt 5401 tgtttatttt tctaaataca ttcaaatatg tatccgctca tgaattaatt cttagaaaaa 5461 ctcatcgagc atcaaatgaa actgcaattt attcatatca ggattatcaa taccatattt 5521 ttgaaaaagc cgtttctgta atgaaggaga aaactcaccg aggcagttcc ataggatggc 5581 aagatcctgg tatcggtctg cgattccgac tcgtccaaca tcaatacaac ctattaattt 5641 cccctcgtca aaaataaggt tatcaagtga gaaatcacca tgagtgacga ctgaatccgg 5701 tgagaatggc aaaagtttat gcatttcttt ccagacttgt tcaacaggcc agccattacg 5761 ctcgtcatca aaatcactcg catcaaccaa accgttattc attcgtgatt gcgcctgagc 5821 gagacgaaat acgcggtcgc tgttaaaagg acaattacaa acaggaatcg aatgcaaccg 5881 gcgcaggaac actgccagcg catcaacaat attttcacct gaatcaggat attcttctaa 5941 tacctggaat gctgttttcc cggggatcgc agtggtgagt aaccatgcat catcaggagt 6001 acggataaaa tgcttgatgg tcggaagagg cataaattcc gtcagccagt ttagtctgac 6061 catctcatct gtaacatcat tggcaacgct acctttgcca tgtttcagaa acaactctgg 6121 cgcatcgggc ttcccataca atcgatagat tgtcgcacct gattgcccga cattatcgcg 6181 agcccattta tacccatata aatcagcatc catgttggaa tttaatcgcg gcctagagca 6241 agacgtttcc cgttgaatat ggctcatact cttccttttt caatattatt gaagcattta 6301 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat 6361 aggcatgcta gcgcagaaac gtcctagaag atgccaggag gatacttagc agagagacaa 6421 taaggccgga gcgaagccgt ttttccatag gctccgcccc cctgacgaac atcacgaaat 6481 ctgacgctca aatcagtggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc 6541 ccctgatggc tccctcttgc gctctcctgt tcccgtcctg cggcgtccgt gttgtggtgg 6601 aggctttacc caaatcacca cgtcccgttc cgtgtagaca gttcgctcca agctgggctg 6661 tgtgcaagaa ccccccgttc agcccgactg ctgcgcctta tccggtaact atcatcttga 6721 gtccaacccg gaaagacacg acaaaacgcc actggcagca gccattggta actgagaatt 6781 agtggattta gatatcgaga gtcttgaagt ggtggcctaa cagaggctac actgaaagga 6841 cagtatttgg tatctgcgct ccactaaagc cagttaccag gttaagcagt tccccaactg 6901 acttaacctt cgatcaaacc gcctccccag gcggtttttt cgtttacaga gcaggagatt 6961 acgacgatcg taaaaggatc tcaagaagat cctttacgga ttcccgacac catcactcta 7021 gatttcagtg caatttatct cttcaaatgt agcacctgaa gtcagcccca tacgatataa 7081 gttgtaattc tcatgttagt catgccccgc gcccaccgga aggagctgac tgggttgctc 7141 ctagggtctg attcgttacc aattatgaca acttgacggc tacatcattc actttttctt 7201 cacaaccggc acggaactcg ctcgggctgg ccccggtgca ttttttaaat acccgcgaga 7261 aatagagttg atcgtcaaaa ccaacattgc gaccgacggt ggcgataggc atccgggtgg 7321 tgctcaaaag cagcttcgcc tggctgatac gttggtcctc gcgccagctt aagacgctaa 7381 tccctaactg ctggcggaaa agatgtgaca gacgcgacgg cgacaagcaa acatgctgtg 7441 cgacgctggc gatatcaaaa ttgctgtctg ccaggtgatc gctgatgtac tgacaagcct 7501 cgcgtacccg attatccatc ggtggatgga gcgactcgtt aatcgcttcc atgcgccgca 7561 gtaacaattg ctcaagcaga tttatcgcca gcagctccga atagcgccct tccccttgcc 7621 cggcgttaat gatttgccca aacaggtcgc tgaaatgcgg ctggtgcgct tcatccgggc 7681 gaaagaaccc cgtattggca aatattgacg gccagttaag ccattcatgc cagtaggcgc 7741 gcggacgaaa gtaaacccac tggtgatacc attcgcgagc ctccggatga cgaccgtagt 7801 gatgaatctc tcctggcggg aacagcaaaa tatcacccgg tcggcaaaca aattctcgtc 7861 cctgattttt caccaccccc tgaccgcgaa tggtgagatt gagaatataa cctttcattc 7921 ccagcggtcg gtcgataaaa aaatcgagat aaccgttggc ctcaatcggc gttaaacccg 7981 ccaccagatg ggcattaaac gag //