LOCUS       sequence-174899-        9173 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses efflux pump in E. coli to provide resistance to biofuel 
            (low copy).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9173)
  AUTHORS   Turner WJ, Dunlop MJ
  TITLE     Trade-Offs in Improving Biofuel Tolerance Using Combinations of 
            Efflux Pumps.
  JOURNAL   ACS Synth Biol. 2015 Oct 16;4(10):1056-63. doi: 10.1021/sb500307w. 
            Epub 2014 Dec 12.
  PUBMED    25496359
REFERENCE   2  (bases 1 to 9173)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: "10.1021/sb500307w";
            journalName: "ACS Synth Biol"; date: "2015-10-16- 16"; volume: "4"; 
            issue: "10"; pages: "1056-63"
COMMENT     Sequence Label: pBbS5c-Maqu_3494
FEATURES             Location/Qualifiers
     source          1..9173
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(49..1131)
                     /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     1093..1112
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(1132..1209)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1415..1434
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        join(1520..1525,1526..1543,1544..1550)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    1558..1574
                     /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)."
     regulatory      5500..5509
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     terminator      5916..5987
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      6003..6030
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     CDS             complement(6561..7511)
                     /codon_start=1
                     /gene="rep101"
                     /product="RepA protein needed for replication with the 
                     pSC101 origin"
                     /label=Rep101
                     /translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE
                     RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFEIFQWTNYAKFS
                     SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
                     EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
                     ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLHNGLRKTLHDALTAKIQLTSFEAKF
                     LSDMQSKYDLNGSFSWLTQKQRTTLENILAKYGRI"
     terminator      complement(8273..8367)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(8388..9047)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     8962..8981
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(9048..9150)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
ORIGIN
        1 gacgtcggtg cctaatgagt gagctaactt acattaattg cgttgcgctc actgcccgct
       61 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga
      121 ggcggtttgc gtattgggcg ccagggtggt ttttcttttc accagtgaga cgggcaacag
      181 ctgattgccc ttcaccgcct ggccctgaga gagttgcagc aagcggtcca cgctggtttg
      241 ccccagcagg cgaaaatcct gtttgatggt ggttaacggc gggatataac atgagctgtc
      301 ttcggtatcg tcgtatccca ctaccgagat gtccgcacca acgcgcagcc cggactcggt
      361 aatggcgcgc attgcgccca gcgccatctg atcgttggca accagcatcg cagtgggaac
      421 gatgccctca ttcagcattt gcatggtttg ttgaaaaccg gacatggcac tccagtcgcc
      481 ttcccgttcc gctatcggct gaatttgatt gcgagtgaga tatttatgcc agccagccag
      541 acgcagacgc gccgagacag aacttaatgg gcccgctaac agcgcgattt gctggtgacc
      601 caatgcgacc agatgctcca cgcccagtcg cgtaccgtct tcatgggaga aaataatact
      661 gttgatgggt gtctggtcag agacatcaag aaataacgcc ggaacattag tgcaggcagc
      721 ttccacagca atggcatcct ggtcatccag cggatagtta atgatcagcc cactgacgcg
      781 ttgcgcgaga agattgtgca ccgccgcttt acaggcttcg acgccgcttc gttctaccat
      841 cgacaccacc acgctggcac ccagttgatc ggcgcgagat ttaatcgccg cgacaatttg
      901 cgacggcgcg tgcagggcca gactggaggt ggcaacgcca atcagcaacg actgtttgcc
      961 cgccagttgt tgtgccacgc ggttgggaat gtaattcagc tccgccatcg ccgcttccac
     1021 tttttcccgc gttttcgcag aaacgtggct ggcctggttc accacgcggg aaacggtctg
     1081 ataagagaca ccggcatact ctgcgacatc gtataacgtt actggtttca cattcaccac
     1141 cctgaattga ctctcttccg ggcgctatca tgccataccg cgaaaggttt tgcgccattc
     1201 gatggtgtcc gggatctcga cgctctccct tatgcgactc ctgcattagg aagcagccca
     1261 gtagtaggtt gaggccgttg agcaccgccg ccgcaaggaa tggtgcatgc aaggagatgg
     1321 cgcccaacag tcccccggcc acggggcctg ccaccatacc cacgccgaaa caagcgctca
     1381 tgagcccgaa gtggcgagcc cgatcttccc catcggtgat gtcggcgata taggcgccag
     1441 caaccgcacc tgtggcgccg gtgatgccgg ccacgatgcg tccggcgtag aggatcgaga
     1501 tcgtttaggc accccaggct ttacacttta tgcttccggc tcgtataatg tgtggaattg
     1561 tgagcggata acaatttcag aattcaaaag atcttttaag aaggagatat acatgtgatt
     1621 tcaaacaaac acctgcccga cagccgcctg ctggcgctgc tgctaaccag tatttttgta
     1681 ctttccggat gcagcgatcc ggcaccgcaa caacaaaacc agcgcccggc gcctcaggtg
     1741 ggggtgtatg aagtggaaga acaaccactg gccctgtctt ccgccctgcc tggccgcacc
     1801 cgcgcctggc ggactgccga ggttcgccca caggtgaatg gcattattga agagcgactg
     1861 ttcgttgaag gcagcgtagt ggatgagggg tcagtgctgt accagatcga tgacgctccc
     1921 taccgggccg ccctggcccg ggccaatgcc agcctgacca atgcccgcaa cctggccaag
     1981 cgctacgaaa acctggtggg cacccgttcc atcagccagc agcagtatga cgacgccctg
     2041 gctgccctgc gcagcgccga ggcggaatac gaagccgcca gcatcaacat gaactacacc
     2101 cgggtgaaag ccccgatcac cggcaagatc agccgttcgc gagtcagcga aggcgccctg
     2161 gtgaccagcg gccagcagca ggaaatggcc accatcaccc aactggaccc gatctacgtg
     2221 gacatcgtgc aaccggtcac ccgccttctg gacctgcaga gcgctatcga agccgggctg
     2281 attgaagccg atgagaacga cgtggttgaa gtctcgctgg tgctggagaa cggccaggac
     2341 tatccgcaca ccggtttgct gaagttctcc gaagtggtgg tcgatgaggg caccggctcg
     2401 gtcaccctgc gggctgagtt cccgaatccg gatggcaagc tgctgccggg tatgtttgtg
     2461 cgcgcgctgg tggaagaagg cacccgcccg aacgccatcc tggtgcccca gcaagcggtg
     2521 tcacgcgata cccgcggcgt accaaccgtc tgggtgctgg ccgaggacaa cacggtaagc
     2581 gtccgccaga ttgaaaccga gcgcaccgtg ggcaacgcct ggctggtgca aaaggggctg
     2641 caggccggtg aacaggtggt gaccgagggc gttcagcgcc tggcgccggg cattcccgtg
     2701 cagccggcac cggcagccaa tgtcgacctg aaaaccgaat tcgcaaccca ggccgaataa
     2761 ggagctccgg acatgccccg attttttatc gaacgcccca tatttgcctg ggtcgtcgcc
     2821 ctgatgctga tgctcggagg cggactggca gtaaagaacc tcgccgttaa ccagtttccg
     2881 gatgtcgccc cgccggcgat tgccttgtcg gtgaattacc ccggcgcttc cgcccagacc
     2941 gtgcaggaca cggtggtcca ggtgatcgag caacagctca acggcctgga cggcttgcgc
     3001 tacatctcct ccgagagcaa ctcggatggc agcatgacca tcattgccac cttcgaacag
     3061 ggcaccgacc cagacatcgc ccaggtgcag gtgcaaaaca aactgcagct ggccaacccg
     3121 ctgctgcccg aagaagtgca gcgtcaaggc atccgggtgt ctaagtacaa ggtgaacttc
     3181 ttcaccgtat ttgcgctgac cagccctgat ggcaagtaca cccagggcga cctggccgat
     3241 tacatcgtgt ccaatatcca ggacccggtt gcgcgaaccc agggcgtcgg tgactttctg
     3301 ctgttcgtct cccagtacgc catgcgcctg tggctcgacc cggaaaaact caacagctac
     3361 cagctgaccc cgcaggacgt gatcaattcg gtgcgcgccc agaacgtgca ggtctctgcc
     3421 ggccagctcg gtggcctgcc caccgcagaa ggcgtgcagc tgcaggccac cgttatcggc
     3481 aagcagcgca tgaagaccgc cgaagaattc gaaaacatcc tgctgaaagt gaacccggac
     3541 ggctcccagg tgcgcctggc cgatgtcgcc gaggtgaatc tgggcaacga gaactacgcc
     3601 accaccggta aatacaacgg cgcgccggca gccggtatgg ccctgcgcct ggccaccggc
     3661 gccaaccagc tggaaaccgc tggccgggta aaagaaaccc tggcagaact ggaacgcttt
     3721 ctgcctgaag gcgtagagat cgtcttcccc tacgacacca caccggtcgt gtcggcgtct
     3781 atcgagaccg tcgccatgac cctgatcgaa gccgtggtgc tggtgttcct ggtgatgttc
     3841 ctgttcctgc aaagctggcg cgccaccatc attcccacac tggcggtgcc ggtggttctg
     3901 ctggctacct ttggcgtact gtacgccttt ggcttcaccg tcaacgtgat gaccatgttt
     3961 gccatggtcc tggccatcgg tttgctggtg gacgacgcca tcgtagtggt ggaaaacgtc
     4021 gagcgactga tggaagaaga ggggctgtcg cccaaggaag ccgccaagaa atccatggac
     4081 cagatctccg gcgcactgct gggcatcggc ctggtgattt cggcggtgtt cctgcccatg
     4141 gccttttttg gcggctccac cggggtgatc taccggcagt tctcggtcac catcatctcg
     4201 gcaatgtcgt tctctgtgct ggtggcattt attttcaccc cggccctgtg cgccaccttg
     4261 ctgaaacccg gcgaccagca cgtcaggaaa ggctttttcg gctggtttaa ccgcaccttt
     4321 gaccgcagcg cagaccgcta caaatcaggc gtcagttacc tgatcaaacg taagggccgc
     4381 tttatgggcg tgtacctgtt gctggtcgtg gctgtgggct tcctgttcaa gggcctgccc
     4441 accgcattcc tgccggacga agatcagggc gtgatgattg tgatggtgca gctgcccacc
     4501 aatgccacgg gcgagcgcac cgaagccgta ctggcagaag ctggaaatta tttgctggaa
     4561 gaagagtccg aggtggtcaa gagcgtgatg tctgtgcggg gcttcaactt cgccggccgt
     4621 ggccagaact ccggtattct gttcgtggac ctgaagccct ttgctgaccg ggaaagcttt
     4681 gcccagagcg tgttcgccct cgcgggccgc tccggggcaa ggttcgccca gatcaaagac
     4741 gccatcgtgt tcccgattgt gccaccggcc attctggagc tcggcaacgc caccggcttt
     4801 gacctctacc tgaaagacaa cggagctatc ggccaccacg ccctgatggc cgccaccaat
     4861 gagtttatca gcagagccaa tgccgcgccg gagctgaaca tggtgcgcca caacggcctg
     4921 ccggacgagc cccaatacca ggtgatcatc gatgacgaaa aagcccggct gttgcaggtg
     4981 tccattgccg acatcaatgc caccatgtct gccgcctggg gttcaagcta tgtgaacgac
     5041 ttcctgcaca acggccgggt gaagaaagtc tatgtgcagg gcaaaccgga ttcacgactg
     5101 gcaccggaag acttcgacaa atggtttgtg cgcaacgccc agggtgagat ggtgcccttt
     5161 gccgcctttg ccactggcga gtgggtgttc ggttcgccgc gcctgcagcg gtatcagggc
     5221 ttacccgcta cgcaaatcca gggtgctccg gctaacggtt acagtaccgg tgatgccatg
     5281 gccgcactgg aacgcattgc cgccgacctg ccccagggcc tgggcctgga atacaccggc
     5341 ctgtcgttcg aggaaaaaca ggccggcaac caggcgatga tgctgtatct gctgtcgatc
     5401 ctggtggtgt tcctgtgcct ggcagccctg tatgaaagct ggtcgattcc gtttgcggtg
     5461 atcatgctgg tgccactggg cgtgcttggc gcggtcctgg ccaccatggc cagagggctg
     5521 tccaacgatg tgttcttcca ggtgggtatg ctgactacca tggggctggc cgccaagaac
     5581 gccatcctga tcgtggagtt cgcccgccag ctctatgagc aggaaggcaa gccgctgctg
     5641 caggccaccg ccgaggccgc ccggctgcgc ctgaggccga tcatcatgac ctcgctggcg
     5701 ttcatcttcg gggtactgcc catggccatc gccagcggtg cctccagcgc cagccagcat
     5761 gccatcggca ccgcggtggt cggcggcaca ctggccgcga ccatcctggc gatcttcttt
     5821 gtaccgctgt tctacgtgtt tgtggtgggc ctgaccggaa agcggaaatc cgctgacgat
     5881 taaggatcca aactcgagta aggatctcca ggcatcaaat aaaacgaaag gctcagtcga
     5941 aagactgggc ctttcgtttt atctgttgtt tgtcggtgaa cgctctctac tagagtcaca
     6001 ctggctcacc ttcgggtggg cctttctgcg tttataccta gggtacgggt tttgctgccc
     6061 gcaaacgggc tgttctggtg ttgctagttt gttatcagaa tcgcagatcc ggcttcagcc
     6121 ggtttgccgg ctgaaagcgc tatttcttcc agaattgcca tgattttttc cccacgggag
     6181 gcgtcactgg ctcccgtgtt gtcggcagct ttgattcgat aagcagcatc gcctgtttca
     6241 ggctgtctat gtgtgactgt tgagctgtaa caagttgtct caggtgttca atttcatgtt
     6301 ctagttgctt tgttttactg gtttcacctg ttctattagg tgttacatgc tgttcatctg
     6361 ttacattgtc gatctgttca tggtgaacag ctttgaatgc accaaaaact cgtaaaagct
     6421 ctgatgtatc tatctttttt acaccgtttt catctgtgca tatggacagt tttccctttg
     6481 atatgtaacg gtgaacagtt gttctacttt tgtttgttag tcttgatgct tcactgatag
     6541 atacaagagc cataagaacc tcagatcctt ccgtatttag ccagtatgtt ctctagtgtg
     6601 gttcgttgtt tttgcgtgag ccatgagaac gaaccattga gatcatactt actttgcatg
     6661 tcactcaaaa attttgcctc aaaactggtg agctgaattt ttgcagttaa agcatcgtgt
     6721 agtgtttttc ttagtccgtt atgtaggtag gaatctgatg taatggttgt tggtattttg
     6781 tcaccattca tttttatctg gttgttctca agttcggtta cgagatccat ttgtctatct
     6841 agttcaactt ggaaaatcaa cgtatcagtc gggcggcctc gcttatcaac caccaatttc
     6901 atattgctgt aagtgtttaa atctttactt attggtttca aaacccattg gttaagcctt
     6961 ttaaactcat ggtagttatt ttcaagcatt aacatgaact taaattcatc aaggctaatc
     7021 tctatatttg ccttgtgagt tttcttttgt gttagttctt ttaataacca ctcataaatc
     7081 ctcatagagt atttgttttc aaaagactta acatgttcca gattatattt tatgaatttt
     7141 tttaactgga aaagataagg caatatctct tcactaaaaa ctaattctaa tttttcgctt
     7201 gagaacttgg catagtttgt ccactggaaa atctcaaagc ctttaaccaa aggattcctg
     7261 atttccacag ttctcgtcat cagctctctg gttgctttag ctaatacacc ataagcattt
     7321 tccctactga tgttcatcat ctgagcgtat tggttataag tgaacgatac cgtccgttct
     7381 ttccttgtag ggttttcaat cgtggggttg agtagtgcca cacagcataa aattagcttg
     7441 gtttcatgct ccgttaagtc atagcgacta atcgctagtt catttgcttt gaaaacaact
     7501 aattcagaca tacatctcaa ttggtctagg tgattttaat cactatacca attgagatgg
     7561 gctagtcaat gataattact agtccttttc ccgggtgatc tgggtatctg taaattctgc
     7621 tagacctttg ctggaaaact tgtaaattct gctagaccct ctgtaaattc cgctagacct
     7681 ttgtgtgttt tttttgttta tattcaagtg gttataattt atagaataaa gaaagaataa
     7741 aaaaagataa aaagaataga tcccagccct gtgtataact cactacttta gtcagttccg
     7801 cagtattaca aaaggatgtc gcaaacgctg tttgctcctc tacaaaacag accttaaaac
     7861 cctaaaggct taagtagcac cctcgcaagc tcgggcaaat cgctgaatat tccttttgtc
     7921 tccgaccatc aggcacctga gtcgctgtct ttttcgtgac attcagttcg ctgcgctcac
     7981 ggctctggca gtgaatgggg gtaaatggca ctacaggcgc cttttatgga ttcatgcaag
     8041 gaaactaccc ataatacaag aaaagcccgt cacgggcttc tcagggcgtt ttatggcggg
     8101 tctgctatgt ggtgctatct gactttttgc tgttcagcag ttcctgccct ctgattttcc
     8161 agtctgacca cttcggatta tcccgtgaca ggtcattcag actggctaat gcacccagta
     8221 aggcagcggt atcatcaaca ggcttacccg tcttactgtc cctagtgctt ggattctcac
     8281 caataaaaaa cgcccggcgg caaccgagcg ttctgaacaa atccagatgg agttctgagg
     8341 tcattactgg atctatcaac aggagtccaa gcgagctcga tatcaaatta cgccccgccc
     8401 tgccactcat cgcagtactg ttgtaattca ttaagcattc tgccgacatg gaagccatca
     8461 caaacggcat gatgaacctg aatcgccagc ggcatcagca ccttgtcgcc ttgcgtataa
     8521 tatttgccca tggtgaaaac gggggcgaag aagttgtcca tattggccac gtttaaatca
     8581 aaactggtga aactcaccca gggattggct gagacgaaaa acatattctc aataaaccct
     8641 ttagggaaat aggccaggtt ttcaccgtaa cacgccacat cttgcgaata tatgtgtaga
     8701 aactgccgga aatcgtcgtg gtattcactc cagagcgatg aaaacgtttc agtttgctca
     8761 tggaaaacgg tgtaacaagg gtgaacacta tcccatatca ccagctcacc gtctttcatt
     8821 gccatacgaa attccggatg agcattcatc aggcgggcaa gaatgtgaat aaaggccgga
     8881 taaaacttgt gcttattttt ctttacggtc tttaaaaagg ccgtaatatc cagctgaacg
     8941 gtctggttat aggtacattg agcaactgac tgaaatgcct caaaatgttc tttacgatgc
     9001 cattgggata tatcaacggt ggtatatcca gtgatttttt tctccatttt agcttcctta
     9061 gctcctgaaa atctcgataa ctcaaaaaat acgcccggta gtgatcttat ttcattatgg
     9121 tgaaagttgg aacctcttac gtgccgatca acgtctcatt ttcgccagat atc
//
