LOCUS Exported 7975 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION synthetic circular DNA ACCESSION . VERSION . KEYWORDS pBbA5k-EPL100 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7975) 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 7975) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7975 /organism="synthetic DNA construct" /mol_type="other DNA" CDS 4318..4335 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" terminator 4371..4442 /label=rrnB T1 terminator /note="transcription terminator T1 from the E. coli rrnB gene" terminator 4458..4485 /label=T7Te terminator /note="phage T7 early transcription terminator" primer_bind complement(4513..4530) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(4647..5192) /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(4727..4746) /label=pBR322ori-F /note="pBR322 origin, forward primer" terminator 5306..5400 /label=lambda t0 terminator /note="transcription terminator from phage lambda" CDS complement(5431..6225) /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(5542..5561) /label=Neo-F /note="Neomycin resistance gene, forward primer" primer_bind 6152..6171 /label=Neo-R /note="Neomycin resistance gene, reverse primer" CDS complement(6410..7492) /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 7454..7473 /label=LacI-R /note="LacI, reverse primer" promoter complement(7493..7570) /gene="lacI" /label=lacI promoter primer_bind 7776..7795 /label=pBRrevBam /note="pBR322 vectors, tet region, downstream of BamHI, reverse primer" promoter 7881..7911 /label=lac UV5 promoter /note="E. coli lac promoter with an ""up"" mutation" protein_bind 7919..7935 /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 atgcggcaaa tagtgaaaat tgcctctgtt ttaagtgtgg cgttatggat aagtggctgt 61 gagcaaaaac ctgaacaggg gccgcaacag cgtggaccaa tggaagtggg ggttataact 121 attgctgcta agcctcaatc aatcatggtt gaattaccgg gtcgtagtaa gtcttattta 181 gaggctgaag tgcgtccaca ggtaagcggt attattatgg aacgtggttt tgttgaaggt 241 agggatgtcg aaaaaggaca atctttatat cagatcgagc catcaaccta cgaagcagca 301 ctcgttagtg ctaaagccga tcttaagagt gcgcaagcgg gtctcgtttc ggctaaagca 361 aaggcactgc gttttgaaaa actagtcaag cttaactcaa tcagtgagca agattatgat 421 gaagcggatg cggcttacaa agaagctcta gcacgtgtaa cggttgctga agcgaatatt 481 aatactgctc aaattaatct agagtacacc aaggtaacgg cgccaatctc tggccgtatc 541 ggtaagtctg atgtgactgc tggtgcactt gttaccgcaa atcagagcca aaccttggcg 601 acgattcagc aacttgaccc aatgaatatc gatatcgctc agtcaagcac gcagatgctg 661 cgtcttaagt ctcagctaaa gcgtggtcaa attgaggctg cggcaaatgc caatgttgag 721 ctgattttag aagacggtac tccgtaccat catactggtg tgttgcagtt tgctgaggtg 781 acggtcaacg aaaataccgg ttcggtgatt atccgtgcag aatttccaaa cccagacggt 841 gtgttgatgc ctggcatgta tgtacgtgca gtattgaata cgggtaccga tcctgaagcg 901 attcttgtcc cacaaaaagc catcactcgt aatactaagg gacaggctgt ggctatggtt 961 attgaagatg gcaaggttgc agcgcgcatc gtgactacag ctgaagtgat tgacaatcaa 1021 tggcgtatta ccagcggcct taaagtaggc gaacagctaa tcgttgaagg ccttcaaaaa 1081 attcgtcctg gagcccctgt taaagcggga cctttggccg ctgataagaa accagaaaca 1141 caacagaaat aggatagggt tatggcacgt ttctttatcg atcgccctat ttttgcatgg 1201 gtgatcgcta ttattgtgat gctggcaggt gtgctctcta ttgtgggcct acccgtatct 1261 cagtacccga gtattgcacc gccgtcggtt gtgattagcg ccgtttatcc tggtgcatcg 1321 gctaagacga tggaagactc ggtaactcag gttattgagc agcgaatgac aggtcttgat 1381 aacctgcgct acatttcatc gaccagcgat agctttggta atgcgcaaat taccttaacc 1441 tttaacgccg aagccgatcc tgatatcgcg caggtacagg tgcagaacaa gctacagctt 1501 gctatgccac tattaccgca agaagttcag gcacagggtg ttaacgtaac taagtcgagt 1561 gctggtttct tgatggtagc tgccttcgta tctgaagacg gttcactgga taaaaatgat 1621 attgccgatt atgtgggttc aaacattcag gatccgctaa gccgtgttcc tggtgtgggc 1681 actattgagc tatttggtgc gcagtatgca atgcgtatct ggcttgaccc actaaaattg 1741 acgcaattta acctaacaag ccaagatgtt attagctcga ttcgagagca aaatgctcag 1801 gtttcagcgg ggcagttagg tggtgcacca tcgctaccgg gccaagagct aaacgcgaca 1861 gtatcggctc agagccgact acaaacggcc gaacagttcc aaaaaatcat cattaaagcc 1921 gatacttcgg gcgctaaggt gttcctagaa gatgtggcgc gtgttgagct gggtgctgaa 1981 agctacgcag ttgagtcatt ttataacggc aagcctgcag ctggtattgg tattcagtta 2041 gcaacgggtg caaacgcatt ggctacggct gaaggtgtac gtgcaaagat tgccgaactg 2101 aagcctttct tccctgaagg gatggaagtg gtttatccat acgacaccac accattcgtt 2161 gagaaatcaa ttgaaggtgt tgtatataca ctgctcgaag ctgtggtgtt agtatttttg 2221 atcatgtacc tgttcttgca gaacttccgt gcaacgctaa ttccaacgat tgcagtgcca 2281 gttgttctat tgggtacatt tgcaattttg gctgtagcag gtttctctat taacacctta 2341 accatgttcg ctatggtact ggcaattggc cttctggttg atgacgcgat tgtggtggtg 2401 gagaacgttg agcgtgtgat gcaggaggag gggctcagtc caattgaggc caccaagaag 2461 tcgatggatc agatcaccgg cgcgctggtg ggtattggtc taacgctgtc tgctgtattt 2521 gttccgatgg catttatgtc gggctcaaca ggtgtgattt accgtcagtt ctcaatcacg 2581 attgtatcgg cgatggcact gtcggtaatg gttgctatca tcctgacacc agcactgtgt 2641 gccactatgc ttaagccaat tgctaagggc gaaagccatg caaaaacggg cttctttggt 2701 tggtttaacc gtaagtttga tgcattaacg tctcgttatg aagctaacgt tgcggcaatg 2761 atcaagcgta ctggtcgcac tatgatggtc taccttgcta tcacggttgc cgtgggttgg 2821 atcttcatgc gtatgcctac agcattcctg ccagacgaag accaagggat cttgtttact 2881 caggctattt tgcctgtcaa ctcgactcag caagaaacca aacaagtact tgagcaagtg 2941 tctgactact tcctaaacga tgaaagtgcc aacgttaaat cagtatttag cgtatccggt 3001 tacagcttcg cgggtatggg tcaaaacatg ggtctcgcgt tcgtcggtct taaagactgg 3061 tctgaacgtt cgaatccagg tcaagacgta cagtcagttg ctaaccgtgc catggccaag 3121 ttctcgcaga tgaaagaagc cttcgtgttt gccttcgtac cgccagcggt aatcgaattg 3181 ggcaccgcta acggtttcga tttctacctg caggataaga atggccaagg tcatgaaaag 3241 ttggttgaag cgcgtaacat gttgctgggt atggcagcgc agaatccaaa cctagtgggc 3301 gtacgtccaa atggtcaaga agatgcacca ctttaccaag ttcatatcga tcatggtaag 3361 ttgcgagcct taggtatcga tattgaatcg gttaacagcg tattaggtac tgcctggggt 3421 ggtaactacg ttaatgactt tatcgaccgt ggccgtatta agaaagtata cgttcaaggt 3481 gaagctgagt accgtatgca gccagaagat cttgattctt ggtatgtacg taactctaaa 3541 ggtgagatgg ttcctttctc tgcctttgct tcaggcactt ggggatatgg ttctccacgt 3601 ctagagcgtt tcaacggctt acctgcgatg aacattcaag gtggtacagc accaggttac 3661 agtactggtg cggcaatgga tgatattgaa gctatggtta agaagctgcc acctggattt 3721 ggtatcgagt ggaacggtct ttcttatgaa gagcgtctgt cgggtaacca agcgccaatg 3781 ctttacgcct tgtctattgt ggtggtattc cttgtactag ctgcactata cgagagctgg 3841 tcggtaccat ttgcggtagt actcgttgta cctctgggta ttatcggtgc cttgattgcg 3901 atgaatggtc gtggtttacc taacgacgtg ttcttccagg tcggtctact aaccaccgta 3961 ggtctagcga ctaagaatgc tatcttgatt gtagaattcg ccaaagaata ctatgagaaa 4021 ggtgctggac tcgttgaggc aaccttacac gcggttcgag tgcgtttacg tccaatcttg 4081 atgacgtcgc tagcattcgg actaggtgtt gtgccgttag caatcagtag tggtgtgggt 4141 tcagggagcc agaacgccat tggtactgcg gtacttggcg gtatgatgag ctcgaccttt 4201 atgggtattt tctttatccc aatcttcttc gtgattgttg agcgtatgtt cagtaagcgc 4261 gagcgagatg cagctagagc agtaaaagcg gaagccaaag cggcgaaagc taacaaacat 4321 catcaccatc accactgagg atccaaactc gagtaaggat ctccaggcat caaataaaac 4381 gaaaggctca gtcgaaagac tgggcctttc gttttatctg ttgtttgtcg gtgaacgctc 4441 tctactagag tcacactggc tcaccttcgg gtgggccttt ctgcgtttat acctagggat 4501 atattccgct tcctcgctca ctgactcgct acgctcggtc gttcgactgc ggcgagcgga 4561 aatggcttac gaacggggcg gagatttcct ggaagatgcc aggaagatac ttaacaggga 4621 agtgagaggg ccgcggcaaa gccgtttttc cataggctcc gcccccctga caagcatcac 4681 gaaatctgac gctcaaatca gtggtggcga aacccgacag gactataaag ataccaggcg 4741 tttccccctg gcggctccct cgtgcgctct cctgttcctg cctttcggtt taccggtgtc 4801 attccgctgt tatggccgcg tttgtctcat tccacgcctg acactcagtt ccgggtaggc 4861 agttcgctcc aagctggact gtatgcacga accccccgtt cagtccgacc gctgcgcctt 4921 atccggtaac tatcgtcttg agtccaaccc ggaaagacat gcaaaagcac cactggcagc 4981 agccactggt aattgattta gaggagttag tcttgaagtc atgcgccggt taaggctaaa 5041 ctgaaaggac aagttttggt gactgcgctc ctccaagcca gttacctcgg ttcaaagagt 5101 tggtagctca gagaaccttc gaaaaaccgc cctgcaaggc ggttttttcg ttttcagagc 5161 aagagattac gcgcagacca aaacgatctc aagaagatca tcttattaat cagataaaat 5221 atttctagat ttcagtgcaa tttatctctt caaatgtagc acctgaagtc agccccatac 5281 gatataagtt gttactagtg cttggattct caccaataaa aaacgcccgg cggcaaccga 5341 gcgttctgaa caaatccaga tggagttctg aggtcattac tggatctatc aacaggagtc 5401 caagcgagct ctcgaacccc agagtcccgc tcagaagaac tcgtcaagaa ggcgatagaa 5461 ggcgatgcgc tgcgaatcgg gagcggcgat accgtaaagc acgaggaagc ggtcagccca 5521 ttcgccgcca agctcttcag caatatcacg ggtagccaac gctatgtcct gatagcggtc 5581 cgccacaccc agccggccac agtcgatgaa tccagaaaag cggccatttt ccaccatgat 5641 attcggcaag caggcatcgc catgggtcac gacgagatcc tcgccgtcgg gcatgcgcgc 5701 cttgagcctg gcgaacagtt cggctggcgc gagcccctga tgctcttcgt ccagatcatc 5761 ctgatcgaca agaccggctt ccatccgagt acgtgctcgc tcgatgcgat gtttcgcttg 5821 gtggtcgaat gggcaggtag ccggatcaag cgtatgcagc cgccgcattg catcagccat 5881 gatggatact ttctcggcag gagcaaggtg agatgacagg agatcctgcc ccggcacttc 5941 gcccaatagc agccagtccc ttcccgcttc agtgacaacg tcgagcacag ctgcgcaagg 6001 aacgcccgtc gtggccagcc acgatagccg cgctgcctcg tcctgcagtt cattcagggc 6061 accggacagg tcggtcttga caaaaagaac cgggcgcccc tgcgctgaca gccggaacac 6121 ggcggcatca gagcagccga ttgtctgttg tgcccagtca tagccgaata gcctctccac 6181 ccaagcggcc ggagaacctg cgtgcaatcc atcttgttca atcatgcgaa acgatcctca 6241 tcctgtctct tgatcagatc atgatcccct gcgccatcag atccttggcg gcaagaaagc 6301 catccagttt actttgcagg gcttcccaac cttaccagag ggcgccccag ctggcaattc 6361 cgacgtcggt gcctaatgag tgagctaact tacattaatt gcgttgcgct cactgcccgc 6421 tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag 6481 aggcggtttg cgtattgggc gccagggtgg tttttctttt caccagtgag acgggcaaca 6541 gctgattgcc cttcaccgcc tggccctgag agagttgcag caagcggtcc acgctggttt 6601 gccccagcag gcgaaaatcc tgtttgatgg tggttaacgg cgggatataa catgagctgt 6661 cttcggtatc gtcgtatccc actaccgaga tgtccgcacc aacgcgcagc ccggactcgg 6721 taatggcgcg cattgcgccc agcgccatct gatcgttggc aaccagcatc gcagtgggaa 6781 cgatgccctc attcagcatt tgcatggttt gttgaaaacc ggacatggca ctccagtcgc 6841 cttcccgttc cgctatcggc tgaatttgat tgcgagtgag atatttatgc cagccagcca 6901 gacgcagacg cgccgagaca gaacttaatg ggcccgctaa cagcgcgatt tgctggtgac 6961 ccaatgcgac cagatgctcc acgcccagtc gcgtaccgtc ttcatgggag aaaataatac 7021 tgttgatggg tgtctggtca gagacatcaa gaaataacgc cggaacatta gtgcaggcag 7081 cttccacagc aatggcatcc tggtcatcca gcggatagtt aatgatcagc ccactgacgc 7141 gttgcgcgag aagattgtgc accgccgctt tacaggcttc gacgccgctt cgttctacca 7201 tcgacaccac cacgctggca cccagttgat cggcgcgaga tttaatcgcc gcgacaattt 7261 gcgacggcgc gtgcagggcc agactggagg tggcaacgcc aatcagcaac gactgtttgc 7321 ccgccagttg ttgtgccacg cggttgggaa tgtaattcag ctccgccatc gccgcttcca 7381 ctttttcccg cgttttcgca gaaacgtggc tggcctggtt caccacgcgg gaaacggtct 7441 gataagagac accggcatac tctgcgacat cgtataacgt tactggtttc acattcacca 7501 ccctgaattg actctcttcc gggcgctatc atgccatacc gcgaaaggtt ttgcgccatt 7561 cgatggtgtc cgggatctcg acgctctccc ttatgcgact cctgcattag gaagcagccc 7621 agtagtaggt tgaggccgtt gagcaccgcc gccgcaagga atggtgcatg caaggagatg 7681 gcgcccaaca gtcccccggc cacggggcct gccaccatac ccacgccgaa acaagcgctc 7741 atgagcccga agtggcgagc ccgatcttcc ccatcggtga tgtcggcgat ataggcgcca 7801 gcaaccgcac ctgtggcgcc ggtgatgccg gccacgatgc gtccggcgta gaggatcgag 7861 atcgtttagg caccccaggc tttacacttt atgcttccgg ctcgtataat gtgtggaatt 7921 gtgagcggat aacaatttca gaattcaaaa gatcttttaa gaaggagata tacat //