LOCUS Exported 7924 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION synthetic circular DNA ACCESSION . VERSION . KEYWORDS pBbA5k-EPL55 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7924) AUTHORS Dunlop MJ, Dossani ZY, Szmidt HL, Chu HC, Lee TS, Keasling JD, Hadi MZ, Mukhopadhyay A TITLE Engineering microbial biofuel tolerance and export using efflux pumps. JOURNAL Mol Syst Biol. 2011 May 10;7:487. doi: 10.1038/msb.2011.21. PUBMED 21556065 REFERENCE 2 (bases 1 to 7924) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7924 /organism="synthetic DNA construct" /mol_type="other DNA" regulatory 3886..3895 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 4267..4284 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" terminator 4320..4391 /label=rrnB T1 terminator /note="transcription terminator T1 from the E. coli rrnB gene" terminator 4407..4434 /label=T7Te terminator /note="phage T7 early transcription terminator" primer_bind complement(4462..4479) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(4596..5141) /direction=LEFT /label=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 complement(4676..4695) /label=pBR322ori-F /note="pBR322 origin, forward primer" terminator 5255..5349 /label=lambda t0 terminator /note="transcription terminator from phage lambda" CDS complement(5380..6174) /codon_start=1 /gene="aph(3')-II (or nptII)" /product="aminoglycoside phosphotransferase from Tn5" /label=NeoR/KanR /note="confers resistance to neomycin, kanamycin, and G418 (Geneticin(R))" /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF" primer_bind complement(5491..5510) /label=Neo-F /note="Neomycin resistance gene, forward primer" primer_bind 6101..6120 /label=Neo-R /note="Neomycin resistance gene, reverse primer" CDS complement(6359..7441) /codon_start=1 /gene="lacI" /product="lac repressor" /label=lacI /note="The lac repressor binds to the lac operator to inhibit transcription in E. coli. This inhibition can be relieved by adding lactose or isopropyl-beta-D-thiogalactopyranoside (IPTG)." /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR ALADSLMQLARQVSRLESGQ" primer_bind 7403..7422 /label=LacI-R /note="LacI, reverse primer" promoter complement(7442..7519) /gene="lacI" /label=lacI promoter primer_bind 7725..7744 /label=pBRrevBam /note="pBR322 vectors, tet region, downstream of BamHI, reverse primer" promoter 7830..7860 /label=lac UV5 promoter /note="E. coli lac promoter with an ""up"" mutation" protein_bind 7868..7884 /label=lac operator /bound_moiety="lac repressor encoded by lacI" /note="The lac repressor binds to the lac operator to inhibit transcription in E. coli. This inhibition can be relieved by adding lactose or isopropyl-beta-D-thiogalactopyranoside (IPTG)." ORIGIN 1 gtgatttcaa acaaacacct gcccgacagc cgcctgctgg cgctgctgct aaccagtatt 61 tttgtacttt ccggatgcag cgatccggca ccgcaacaac aaaaccagcg cccggcgcct 121 caggtggggg tgtatgaagt ggaagaacaa ccactggccc tgtcttccgc cctgcctggc 181 cgcacccgcg cctggcggac tgccgaggtt cgcccacagg tgaatggcat tattgaagag 241 cgactgttcg ttgaaggcag cgtagtggat gaggggtcag tgctgtacca gatcgatgac 301 gctccctacc gggccgccct ggcccgggcc aatgccagcc tgaccaatgc ccgcaacctg 361 gccaagcgct acgaaaacct ggtgggcacc cgttccatca gccagcagca gtatgacgac 421 gccctggctg ccctgcgcag cgccgaggcg gaatacgaag ccgccagcat caacatgaac 481 tacacccggg tgaaagcccc gatcaccggc aagatcagcc gttcgcgagt cagcgaaggc 541 gccctggtga ccagcggcca gcagcaggaa atggccacca tcacccaact ggacccgatc 601 tacgtggaca tcgtgcaacc ggtcacccgc cttctggacc tgcagagcgc tatcgaagcc 661 gggctgattg aagccgatga gaacgacgtg gttgaagtct cgctggtgct ggagaacggc 721 caggactatc cgcacaccgg tttgctgaag ttctccgaag tggtggtcga tgagggcacc 781 ggctcggtca ccctgcgggc tgagttcccg aatccggatg gcaagctgct gccgggtatg 841 tttgtgcgcg cgctggtgga agaaggcacc cgcccgaacg ccatcctggt gccccagcaa 901 gcggtgtcac gcgatacccg cggcgtacca accgtctggg tgctggccga ggacaacacg 961 gtaagcgtcc gccagattga aaccgagcgc accgtgggca acgcctggct ggtgcaaaag 1021 gggctgcagg ccggtgaaca ggtggtgacc gagggcgttc agcgcctggc gccgggcatt 1081 cccgtgcagc cggcaccggc agccaatgtc gacctgaaaa ccgaattcgc aacccaggcc 1141 gaataaggag ctccggacat gccccgattt tttatcgaac gccccatatt tgcctgggtc 1201 gtcgccctga tgctgatgct cggcggcgga ctggcagtaa agaacctcgc cgttaaccag 1261 tttccggatg tcgccccgcc ggcgattgcc ttgtcggtga attaccccgg cgcttccgcc 1321 cagaccgtgc aggacacggt ggtccaggtg atcgagcaac agctcaacgg cctggacggc 1381 ttgcgctaca tctcctccga gagcaactcg gatggcagca tgaccatcat tgccaccttc 1441 gaacagggca ccgacccaga catcgcccag gtgcaggtgc aaaacaaact gcagctggcc 1501 aacccgctgc tgcccgaaga agtgcagcgt caaggcatcc gggtgtctaa gtacaaggtg 1561 aacttcttca ccgtatttgc gctgaccagc cctgatggca agtacaccca gggcgacctg 1621 gccgattaca tcgtgtccaa tatccaggac ccggttgcgc gaacccaggg cgtcggtgac 1681 tttctgctgt tcggctccca gtacgccatg cgcctgtggc tcgacccgga aaaactcaac 1741 agctaccagc tgaccccgca ggacgtgatc aattcggtgc gcgcccagaa cgtgcaggtc 1801 tctgccggcc agctcggtgg cctgcccacc gcagaaggcg tgcagctgca ggccaccgtt 1861 atcggcaagc agcgcatgaa gaccgccgaa gaattcgaaa acatcctgct gaaagtgaac 1921 ccggacggct cccaggtgcg cctggccgat gtcgccgagg tgaatctggg caacgagaac 1981 tacgccacca ccggtaaata caacggcgcg ccggcagccg gtatggccct gcgcctggcc 2041 accggcgcca accagctgga aaccgctggc cgggtaaaag aaaccctggc agaactggaa 2101 cgctttctgc ctgaaggcgt agagatcgtc ttcccctacg acaccacacc ggtcgtgtcg 2161 gcgtctatcg agaccgtcgc catgaccctg atcgaagccg tggtgctggt gttcctggtg 2221 atgttcctgt tcctgcaaag ctggcgcgcc accatcattc ccacactggc ggtgccggtg 2281 gttctgctgg ctacctttgg cgtactgtac gcctttggct tcaccgtcaa cgtgatgacc 2341 atgtttgcca tggtcctggc catcggtttg ctggtggacg acgccatcgt agtggtggaa 2401 aacgtcgagc gactgatgga agaagagggg ctgtcgccca aggaagccgc caagaaatcc 2461 atggaccaga tctccggcgc actgctgggc atcggcctgg tgatttcggc ggtgttcctg 2521 cccatggcct tttttggcgg ctccaccggg gtgatctacc ggcagttctc ggtcaccatc 2581 atctcggcaa tgtcgttctc tgtgctggtg gcatttattt tcaccccggc cctgtgcgcc 2641 accttgctga aacccggcga ccagcacgtc aggaaaggct ttttcggctg gtttaaccgc 2701 acctttgacc gcagcgcaga ccgctacaaa tcaggcgtca gttacctgat caaacgtaag 2761 ggccgcttta tgggcgtgta cctgttgctg gtcgtggctg tgggcttcct gttcaagggc 2821 ctgcccaccg cattcctgcc ggacgaagat cagggcgtga tgattgtgat ggtgcagctg 2881 cccaccaatg ccacgggcga gcgcaccgaa gccgtactgg cagaagctgg aaattatttg 2941 ctggaagaag agtccgaggt ggtcaagagc gtgatgtctg tgcggggctt caacttcgcc 3001 ggccgtggcc agaactccgg tattctgttc gtggacctga agccctttgc tgaccgggaa 3061 agctttgccc agagcgtgtt cgccctcgcg ggccgctccg gggcaaggtt cgcccagatc 3121 aaagacgcca tcgtgttccc gattgtgcca ccggccattc tggagctcgg caacgccacc 3181 ggctttgacc tctacctgaa agacaacgga gctatcggcc accacgccct gatggccgcc 3241 accaatgagt ttatcagcag agccaatgcc gcgccggagc tgaacatggt gcgccacaac 3301 ggcctgccgg acgagcccca ataccaggtg atcatcgatg acgaaaaagc ccggctgttg 3361 caggtgtcca ttgccgacat caatgccacc atgtctgccg cctggggttc aagctatgtg 3421 aacgacttcc tgcacaacgg ccgggtgaag aaagtctatg tgcagggcaa accggattca 3481 cgactggcac cggaagactt cgacaaatgg tttgtgcgca acgcccaggg tgagatggtg 3541 ccctttgccg cctttgccac tggcgagtgg gtgttcggtt cgccgcgcct gcagcggtat 3601 cagggcttac ccgctacgca aatccagggt gctccggcta acggttacag taccggtgat 3661 gccatggccg cactggaacg cattgccgcc gacctgcccc agggcctggg cctggaatac 3721 accggcctgt cgttcgagga aaaacaggcc ggcaaccagg cgatgatgct gtatctgctg 3781 tcgatcctgg tggtgttcct gtgcctggca gccctgtatg aaagctggtc gattccgttt 3841 gcggtgatca tgctggtgcc actgggcgtg cttggcgcgg tcctggccac catggccaga 3901 gggctgtcca acgatgtgtt cttccaggtg ggtatgctga ctaccatggg gctggccgcc 3961 aagaacgcca tcctgatcgt ggagttcgcc cgccagctct atgagcagga aggcaagccg 4021 ctgctgcagg ccaccgccga ggccgcccgg ctgcgcctga ggccgatcat catgacctcg 4081 ctggcgttca tcttcggggt actgcccatg gccatcgcca gcggtgcctc cagcgccagc 4141 cagcatgcca tcggcaccgc ggtggtcggc ggcacactgg ccgcgaccat cctggcgatc 4201 ttctttgtac cgctgttcta cgtgtttgtg gtgggcctga ccggaaagcg gaaatccgct 4261 gacgatcatc atcaccatca ccactgagga tccaaactcg agtaaggatc tccaggcatc 4321 aaataaaacg aaaggctcag tcgaaagact gggcctttcg ttttatctgt tgtttgtcgg 4381 tgaacgctct ctactagagt cacactggct caccttcggg tgggcctttc tgcgtttata 4441 cctagggata tattccgctt cctcgctcac tgactcgcta cgctcggtcg ttcgactgcg 4501 gcgagcggaa atggcttacg aacggggcgg agatttcctg gaagatgcca ggaagatact 4561 taacagggaa gtgagagggc cgcggcaaag ccgtttttcc ataggctccg cccccctgac 4621 aagcatcacg aaatctgacg ctcaaatcag tggtggcgaa acccgacagg actataaaga 4681 taccaggcgt ttccccctgg cggctccctc gtgcgctctc ctgttcctgc ctttcggttt 4741 accggtgtca ttccgctgtt atggccgcgt ttgtctcatt ccacgcctga cactcagttc 4801 cgggtaggca gttcgctcca agctggactg tatgcacgaa ccccccgttc agtccgaccg 4861 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gaaagacatg caaaagcacc 4921 actggcagca gccactggta attgatttag aggagttagt cttgaagtca tgcgccggtt 4981 aaggctaaac tgaaaggaca agttttggtg actgcgctcc tccaagccag ttacctcggt 5041 tcaaagagtt ggtagctcag agaaccttcg aaaaaccgcc ctgcaaggcg gttttttcgt 5101 tttcagagca agagattacg cgcagaccaa aacgatctca agaagatcat cttattaatc 5161 agataaaata tttctagatt tcagtgcaat ttatctcttc aaatgtagca cctgaagtca 5221 gccccatacg atataagttg ttactagtgc ttggattctc accaataaaa aacgcccggc 5281 ggcaaccgag cgttctgaac aaatccagat ggagttctga ggtcattact ggatctatca 5341 acaggagtcc aagcgagctc tcgaacccca gagtcccgct cagaagaact cgtcaagaag 5401 gcgatagaag gcgatgcgct gcgaatcggg agcggcgata ccgtaaagca cgaggaagcg 5461 gtcagcccat tcgccgccaa gctcttcagc aatatcacgg gtagccaacg ctatgtcctg 5521 atagcggtcc gccacaccca gccggccaca gtcgatgaat ccagaaaagc ggccattttc 5581 caccatgata ttcggcaagc aggcatcgcc atgggtcacg acgagatcct cgccgtcggg 5641 catgcgcgcc ttgagcctgg cgaacagttc ggctggcgcg agcccctgat gctcttcgtc 5701 cagatcatcc tgatcgacaa gaccggcttc catccgagta cgtgctcgct cgatgcgatg 5761 tttcgcttgg tggtcgaatg ggcaggtagc cggatcaagc gtatgcagcc gccgcattgc 5821 atcagccatg atggatactt tctcggcagg agcaaggtga gatgacagga gatcctgccc 5881 cggcacttcg cccaatagca gccagtccct tcccgcttca gtgacaacgt cgagcacagc 5941 tgcgcaagga acgcccgtcg tggccagcca cgatagccgc gctgcctcgt cctgcagttc 6001 attcagggca ccggacaggt cggtcttgac aaaaagaacc gggcgcccct gcgctgacag 6061 ccggaacacg gcggcatcag agcagccgat tgtctgttgt gcccagtcat agccgaatag 6121 cctctccacc caagcggccg gagaacctgc gtgcaatcca tcttgttcaa tcatgcgaaa 6181 cgatcctcat cctgtctctt gatcagatca tgatcccctg cgccatcaga tccttggcgg 6241 caagaaagcc atccagttta ctttgcaggg cttcccaacc ttaccagagg gcgccccagc 6301 tggcaattcc gacgtcggtg cctaatgagt gagctaactt acattaattg cgttgcgctc 6361 actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg 6421 cgcggggaga ggcggtttgc gtattgggcg ccagggtggt ttttcttttc accagtgaga 6481 cgggcaacag ctgattgccc ttcaccgcct ggccctgaga gagttgcagc aagcggtcca 6541 cgctggtttg ccccagcagg cgaaaatcct gtttgatggt ggttaacggc gggatataac 6601 atgagctgtc ttcggtatcg tcgtatccca ctaccgagat gtccgcacca acgcgcagcc 6661 cggactcggt aatggcgcgc attgcgccca gcgccatctg atcgttggca accagcatcg 6721 cagtgggaac gatgccctca ttcagcattt gcatggtttg ttgaaaaccg gacatggcac 6781 tccagtcgcc ttcccgttcc gctatcggct gaatttgatt gcgagtgaga tatttatgcc 6841 agccagccag acgcagacgc gccgagacag aacttaatgg gcccgctaac agcgcgattt 6901 gctggtgacc caatgcgacc agatgctcca cgcccagtcg cgtaccgtct tcatgggaga 6961 aaataatact gttgatgggt gtctggtcag agacatcaag aaataacgcc ggaacattag 7021 tgcaggcagc ttccacagca atggcatcct ggtcatccag cggatagtta atgatcagcc 7081 cactgacgcg ttgcgcgaga agattgtgca ccgccgcttt acaggcttcg acgccgcttc 7141 gttctaccat cgacaccacc acgctggcac ccagttgatc ggcgcgagat ttaatcgccg 7201 cgacaatttg cgacggcgcg tgcagggcca gactggaggt ggcaacgcca atcagcaacg 7261 actgtttgcc cgccagttgt tgtgccacgc ggttgggaat gtaattcagc tccgccatcg 7321 ccgcttccac tttttcccgc gttttcgcag aaacgtggct ggcctggttc accacgcggg 7381 aaacggtctg ataagagaca ccggcatact ctgcgacatc gtataacgtt actggtttca 7441 cattcaccac cctgaattga ctctcttccg ggcgctatca tgccataccg cgaaaggttt 7501 tgcgccattc gatggtgtcc gggatctcga cgctctccct tatgcgactc ctgcattagg 7561 aagcagccca gtagtaggtt gaggccgttg agcaccgccg ccgcaaggaa tggtgcatgc 7621 aaggagatgg cgcccaacag tcccccggcc acggggcctg ccaccatacc cacgccgaaa 7681 caagcgctca tgagcccgaa gtggcgagcc cgatcttccc catcggtgat gtcggcgata 7741 taggcgccag caaccgcacc tgtggcgccg gtgatgccgg ccacgatgcg tccggcgtag 7801 aggatcgaga tcgtttaggc accccaggct ttacacttta tgcttccggc tcgtataatg 7861 tgtggaattg tgagcggata acaatttcag aattcaaaag atcttttaag aaggagatat 7921 acat //