LOCUS Exported 5434 bp ds-DNA circular SYN 17-OCT-2014 DEFINITION . ACCESSION . VERSION . KEYWORDS piG0071_sensor_pluxRRR12y_bxb1_mCherry SOURCE synthetic DNA construct ORGANISM recombinant plasmid REFERENCE 1 (bases 1 to 5434) AUTHORS Daniel Gerngross TITLE Direct Submission JOURNAL Exported 17 Oct 2014 from SnapGene 2.5.0 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5434 /organism="recombinant plasmid" /mol_type="other DNA" terminator 1..126 /note="T0" terminator 27..121 /note="lambda t0 terminator" /note="transcription terminator from phage lambda" misc_feature 127..196 /note="R2oDNA spacer" /note="R2oDNA 70bp spacer 13" terminator 203..230 /note="T7Te" /note="phage T7 early transcription terminator" misc_feature complement(238..308) /note="taR12y" promoter complement(309..363) /note="lux rR (R0062)" misc_feature 364..433 /note="spacer (B0040)" promoter 434..488 /note="lux rR (R0062)" 3'UTR 489..540 /note="crR12y" RBS 523..534 /note="RBS (B0034)" CDS 541..2043 /codon_start=1 /note="bxb1" /translation="MRALVVIRLSRVTDATTSPERQLESCQQLCAQRGWDVVGVAEDLD VSGAVDPFDRKRRPNLARWLAFEEQPFDVIVAYRVDRLTRSIRHLQQLVHWAEDHKKLV VSATEAHFDTTTPFAAVVIALMGTVAQMELEAIKERNRSAAHFNIRAGKYRGSLPPWGY LPTRVDGEWRLVPDPVQRERILEVYHRVVDNHEPLHLVAHDLNRRGVLSPKDYFAQLQG REPQGREWSATALKRSMISEAMLGYATLNGKTVRDDDGAPLVRAEPILTREQLEALRAE LVKTSRAKPAVSTPSLLLRVLFCAVCGEPAYKFAGGGRKHPRYRCRSMGFPKHCGNGTV AMAEWDAFCEEQVLDLLGDAERLEKVWVAGSDSAVELAEVNAELVDLTSLIGSPAYRAG SPQREALDARIAALAARQEELEGLEARPSGWEWRETGQRFGDWWREQDTAAKNTWLRSM NVRLTFDVRGGLTRTIDFGDLQEYEQHLRLGSVVERLHTGMS" misc_feature 2047..2066 /note="35 nt upstream of ATG from pET vectors" /note="35 nt upstream of ATG from pET vectors (pET RBS) --> introduction of SphI site for library cloning (SphI-HF!!)" misc_feature 2082..2795 /note="mCherry" misc_feature 2097..2102 /note="introduction" /note="introduction of two point mutations to disrupt second inframe RBS in the mCherry gene (2x GAG --> GAA for E)" terminator 2860..2976 /note="T1" terminator 2869..2955 /gene="Escherichia coli rrnB" /note="T1" /note="transcription terminator T1 from the E. coli rrnB gene" primer_bind 2983..3002 /note="AB179" primer_bind 3168..3187 /note="AB5" rep_origin 3321..4148 /direction=RIGHT /note="p15A" rep_origin 3569..3571 /direction=RIGHT /note="P15 ori from pACYC184" rep_origin complement(3572..4116) /direction=LEFT /note="p15A ori" /note="Plasmids containing the medium-copy-number p15A origin of replication can be propagated in E. coli cells that contain a second plasmid with the ColE1 origin." primer_bind 4220..4239 /note="AB180" oriT 4250..4358 /note="incP origin of transfer" CDS complement(4514..5329) /note="KamR" ORIGIN 1 aaatcgtaat tattggggac ccctggattc tcaccaataa aaaacgcccg gcggcaaccg 61 agcgttctga acaaatccag atggagttct gaggtcatta ctggatctat caacaggagt 121 ccaagatatt cgtcacagta gttcaacctc agatacagaa accgtagatt gggattcagg 181 aagttaccct attgtttata aacgcagaaa ggcccacccg aaggtgagcc agtgtgatct 241 agagatatat ggtagtagta agttaatttt cattaaccac cactaccaat cacctcctgg 301 atttagcttt tattcgacta taacaaacca ttttcttgcg taaacctgta cgatcctaca 361 ggtaggttct gttaagtaac tgaacccaat gtcgttagtg acgcttacct cttaagaggt 421 cactgaccta acaacctgta ggatcgtaca ggtttacgca agaaaatggt ttgttatagt 481 cgaataaaga attctaccat tcacctcttg gatttagcta ttaaagagga gaaaggtacc 541 gtgagagccc tggtagtcat ccgcctgtcc cgcgtcaccg atgctacgac ttcaccggag 601 cgtcagctgg agtcttgcca gcagctctgc gcccagcgcg gctgggacgt cgtcggggta 661 gcggaggatc tggacgtctc cggggcggtc gatccgttcg accggaagcg cagaccgaac 721 ctggcccggt ggctagcgtt cgaggagcaa ccgtttgacg tgatcgtggc gtaccgggta 781 gatcggttga cccgatcgat ccggcatctt cagcagctgg tccactgggc cgaggaccac 841 aagaagctgg tcgtctccgc gaccgaagcg cacttcgata cgacgacgcc gtttgcggcg 901 gtcgtcatcg cgcttatggg aacggtggcg cagatggaat tagaagcgat caaagagcgg 961 aaccgttcgg ctgcgcattt caatatccgc gccgggaaat accgaggatc cctgccgccg 1021 tggggatacc tgcctacgcg cgtggacggg gagtggcggc tggtgccgga ccctgtgcag 1081 cgagagcgca tcctcgaggt gtatcaccgc gtcgtcgaca accacgagcc gctgcatctg 1141 gtggcccacg acctgaaccg gcgtggtgtc ctgtcgccga aggactactt cgcgcagctg 1201 caaggccgcg agccgcaggg ccgggagtgg tcggctaccg cgctgaagcg atcgatgatc 1261 tccgaggcga tgctcgggta cgcgactctg aacggtaaga ccgtccgaga cgacgacgga 1321 gccccgctgg tgcgggctga gccgatcctg acccgtgagc agctggaggc gctgcgcgcc 1381 gagctcgtga agacctcccg ggcgaagccc gcggtgtcta ccccgtcgct gctgctgcgg 1441 gtgttgttct gcgcggtgtg cggggagccc gcgtacaagt tcgccggggg aggacgtaag 1501 cacccgcgct accgctgccg ctcgatgggg ttcccgaagc actgcgggaa cggcacggtg 1561 gcgatggccg agtgggacgc gttctgcgag gagcaggtac tggatctgct cggggacgcg 1621 gagcgtctgg agaaagtctg ggtagcgggc tcggactccg cggtcgaact cgcggaggtg 1681 aacgcggagc tggtggacct gacgtcgctg atcggctccc cggcctaccg ggcgggctct 1741 ccgcagcgag aagcactgga tgcccgtatt gcggcgctgg ccgcgcggca agaggagctg 1801 gagggcctgg aggctcgccc gtctggctgg gagtggcgcg agaccgggca gcggttcggg 1861 gactggtggc gggagcagga caccgcggca aagaacacct ggcttcggtc gatgaacgtt 1921 cggctgacgt tcgacgtccg cggcgggctg actcgcacga tcgacttcgg ggatcttcag 1981 gagtacgagc agcatctcag gctcggcagc gtggtcgaac ggctacacac cgggatgtcg 2041 taataaataa ttttgtttaa ctttaatgag gaggtatacg catgcatagc aagggcgaag 2101 aagataacat ggccatcatc aaggagttca tgcgcttcaa ggtgcacatg gagggctccg 2161 tgaacggcca cgagttcgag atcgagggcg agggcgaggg ccgcccctac gagggcaccc 2221 agaccgccaa gctgaaggtg accaagggtg gccccctgcc cttcgcctgg gacatcctgt 2281 cccctcagtt catgtacggc tccaaggcct acgtgaagca ccccgccgac atccccgact 2341 acttgaagct gtccttcccc gagggcttca agtgggagcg cgtgatgaac ttcgaggacg 2401 gcggcgtggt gaccgtgacc caggactcct ccttgcagga cggcgagttc atctacaagg 2461 tgaagctgcg cggcaccaac ttcccctccg acggccccgt aatgcagaag aagaccatgg 2521 gctgggaggc ctcctccgag cggatgtacc ccgaggacgg cgccctgaag ggcgagatca 2581 agcagaggct gaagctgaag gacggcggcc actacgacgc tgaggtcaag accacctaca 2641 aggccaagaa gcccgtgcag ctgcccggcg cctacaacgt caacatcaag ttggacatca 2701 cctcccacaa cgaggactac accatcgtgg aacagtacga acgcgccgag ggccgccact 2761 ccaccggcgg catggacgag ctgtacaaga ggccttaata agaattcgcg cggccgcggc 2821 ctaggcggcc tcctgtgtga aattgttatc cgctttaatt aaaggcatca aataaaacga 2881 aaggctcagt cgaaagactg ggcctttcgt tttatctgtt gtttgtcggt gaacgctctc 2941 ctgagtagga caaatccgcc gccctagaca gctgggcgcg ccaaatacgc ccggtagtga 3001 tcttatttca ttatggtgaa agttggaacc tcttacgtgc cgatcaacgt ctcattttcg 3061 ccaaaagttg gcccagggct tcccggtatc aacagggaca ccaggattta tttattctgc 3121 gaagtgatct tccgtcacag gtatttattc ggcgcaaagt gcgtcgggtg atgctgccaa 3181 cttactgatt tagtgtatga tggtgttttt gaggtgctcc agtggcttct gtttctatca 3241 gctgtccctc ctgttcagct actgacgggg tggtgcgtaa cggcaaaagc accgccggac 3301 atcagcgcta gcggagtgta tactggctta ctatgttggc actgatgagg gtgtcagtga 3361 agtgcttcat gtggcaggag aaaaaaggct gcaccggtgc gtcagcagaa tatgtgatac 3421 aggatatatt ccgcttcctc gctcactgac tcgctacgct cggtcgttcg actgcggcga 3481 gcggaaatgg cttacgaacg gggcggagat ttcctggaag atgccaggaa gatacttaac 3541 agggaagtga gagggccgcg gcaaagccgt ttttccatag gctccgcccc cctgacaagc 3601 atcacgaaat ctgacgctca aatcagtggt ggcgaaaccc gacaggacta taaagatacc 3661 aggcgtttcc cctggcggct ccctcgtgcg ctctcctgtt cctgcctttc ggtttaccgg 3721 tgtcattccg ctgttatggc cgcgtttgtc tcattccacg cctgacactc agttccgggt 3781 aggcagttcg ctccaagctg gactgtatgc acgaaccccc cgttcagtcc gaccgctgcg 3841 ccttatccgg taactatcgt cttgagtcca acccggaaag acatgcaaaa gcaccactgg 3901 cagcagccac tggtaattga tttagaggag ttagtcttga agtcatgcgc cggttaaggc 3961 taaactgaaa ggacaagttt tggtgactgc gctcctccaa gccagttacc tcggttcaaa 4021 gagttggtag ctcagagaac cttcgaaaaa ccgccctgca aggcggtttt ttcgttttca 4081 gagcaagaga ttacgcgcag accaaaacga tctcaagaag atcatcttat taatcagata 4141 aaatatttct agcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg 4201 ggttccgcgc acatttcccc gaaaagtgcc acctgctagg gccggcctac ggccagcctc 4261 gcagagcagg attcccgttg agcaccgcca ggtgcgaata agggacagtg aagaaggaac 4321 acccgctcgc gggtgggcct acttcaccta tcctgcccgg ctgacgccgt tggatacacc 4381 aaggaaagtc tacacgaacc ctttggcaaa atcctgtata tcgtgcgaaa aaggatggat 4441 ataccgaaaa aatcgctata atgaccccga agcagggtta tgcagcggaa aaggaccctt 4501 gtccaattaa ttattagaaa aattcatcca gcatcagatg aaattgcagt ttgttcatat 4561 ccggattatc aatgccatat ttctgaaaca gacgtttttg caggctcggg ctaaattcgc 4621 ccaggcagtt ccacagaatg gccagatcct gataacgatc cgcaatgccc acacggccca 4681 catcaatgca gccaatcagt ttgccttcat cgaaaatcag gttatccagg ctaaaatcgc 4741 cgtgggtcac cacgctatcc gggctaaacg gcagcagttt atgcatttct ttccacacct 4801 gttccaccgg ccagccgtta cgttcatcat caaaatcgct cgcatccacc aggccgttgt 4861 tcatacggct ctgcgcctgg gccagacgaa acacacgatc gctgttaaac gggcagttgc 4921 acaccggaat gctatgcaga cgacgcagaa acacggccag cgcatccaca atgttttcgc 4981 cgctatccgg atattcttcc agcacctgaa acgcggtttt gcccggaatc gcggtggtca 5041 gcagccacgc atcatccggg gtgcgaataa aatgtttaat ggtcggcagc ggcataaatt 5101 cggtcagcca gttcagacgc accatttcat cggtcacatc gttcgccacg ctgcctttgc 5161 catgtttcag aaacagttcc ggcgcatccg gtttgccata cagacgataa atggtcgcgc 5221 cgctctgacc cacgttatca cgcgcccatt tatagccata cagatccgca tccatgttgc 5281 tgttcagacg cggacggcta cagctcgttt cacgctgaat atggctcata acaccccttg 5341 tattactgtt tatgtaagca gacagtttta ttgttcatga tgatatattt ttatcttgtg 5401 caatgtaaca tcagagattt tgagacacaa attt //