LOCUS       sequence-64368-e        9547 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9547)
  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 9547)
  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"; journalName: "Mol Syst 
            Biol."; date: "2011-05-10"; volume: "7:487. doi"; pages: " 
            10.1038/msb.2011.21"
COMMENT     Sequence Label: pBbA5k-EPL102
FEATURES             Location/Qualifiers
     source          1..9547
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             5890..5907
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      5943..6014
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      6030..6057
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     primer_bind     complement(6085..6102)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6219..6764)
                     /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(6299..6318)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(6878..6972)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(7003..7797)
                     /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(7114..7133)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     7724..7743
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     CDS             complement(7982..9064)
                     /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     9026..9045
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(9065..9142)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     9348..9367
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        join(9453..9458,9459..9476,9477..9483)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    9491..9507
                     /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 atgccaagtc atctcagcag cacagcattc aaattaacac tgcttttttc atccgtggca
       61 ttcattactg gctgtgaaaa cgagcctgtg aaaatcaccc ctaaattagt cgttgttgaa
      121 agagtatcga cagcaacagt gccgctctat ggcaattatg tcggtgtaac aaaggcctcg
      181 ttggatgttg aagtaagagc gagagtcgat ggctttgttg aaaataaaac gtttattgaa
      241 ggtagtgcgg ttaaagaagg ggtgctactt tatcgcatcg ataatcgtcc ttatgtggca
      301 gtggttaacc gtttaagtgc taatgttgaa tcccagcaat cagcgctcga aaaggcacaa
      361 cgtgatgtcg aacgtttaaa gccgttgtat gagcaagatg cagccagtca gttagatttc
      421 gataatgcgc tgtcggttct gtcgcaagca aaatccagtg tcgccgcaag taaagccgag
      481 cttgaagaag caaaacttga actcagttat accgagatta agtcacccat tagtggatta
      541 gtcagccgct cagaagttga tatcggtgcg ttagtcggca gcagtggcca atcgttatta
      601 actcgggtta agcaggtcga tcctatctat gtcaccttta atatgtcggc gttagattat
      661 cttaatgcgc gtcgccgcat gaccagtgat aaagagcaac aagccgctga atcagaaggt
      721 aaagctgttg aaggtttcgt aacaattaca ttgccagata acagtgagta ccggtatttg
      781 ggtgatgtcg gctttaccga tccttctgtc aatcctgaaa caggtacttt tcaggtaaga
      841 gctgaactac ctaaccccga taaagagtta ctgccgggcc aatataccaa tgtcagaatt
      901 aagctcagtg aagtgagtga tgcaatcgtt atcccacaaa aagccactca agttgagcaa
      961 ggtggcgtct atgtgatgat tgtgctgccc aataataaag ttgaacgccg ctttattgtg
     1021 ataaaacatc aaggtgaaat gggcgtagta gtacaaagtg gtttaaaagc gggagaatta
     1081 gtcattgtcg aaggcatgca tcgtgtccgt catggacaac tggctgagcc gttaactgcc
     1141 gagcagtatg ataaaaaaga ggatgtaaaa cagaaagagc aagcattgaa gcaacaaaag
     1201 cttgagcagg aacaggagga taactaatgg cacagtattt tgttaatcgt cccgtttttg
     1261 ccagtgtaat ctctattgtc attgtgctat tggggttaat tgcgatgttt caattaccca
     1321 ttgaccaata tccttacatt accccgcctc aggtaaaaat atcggcatcc tatcccggtg
     1381 caacatccac taccgcggca gaatctgtgg ccacaccgtt agagcaagag cttaatggcc
     1441 tgccaaatat gatttatatg agctctaaga gtaccaactc tggtagctcc aacatcacga
     1501 ttacgtttga tgtcggtaca aaccctgatc ttgccgctgt tgatgcacaa aactcaacgc
     1561 agcaagcaac cggtagcttg cctattgatg tgcaaacaga aggggtgtca gtttctaaag
     1621 aagcctctgt tgagctactc aaattagcat taacctctga agatgagcgt tatgatgaaa
     1681 tttatcttag taactacgcc actattaata ttcagtcagc attaaaacgg atccctggtg
     1741 tgggacgagt cagaaataca ggtgcgcgta gttattcaat gagagtgtgg ttaaatcccg
     1801 acacaatggc gggttatggg ctcaccacgt cagatgttat cgatgcgata aaagcgcaaa
     1861 acaaggaatc gccagcgggc agtatcggct cacaacctaa tgcagacacc ttgagcatga
     1921 ctctgccgat caccgcagcc ggtcgaatga gcagtgttcc gcagtttaac gagattattg
     1981 ttcgtgcaag cgccgatggt tcgattattc gcttgcgtga cattgccaac attgaactcg
     2041 gttcatcctc ttatacactc cagtcacaac tcaatggtaa caatgccacc atcttgcagg
     2101 tttacttatt gcccggtgct aatgcgctag aagtcactaa aaaagttaaa gctgagatgg
     2161 caaagctcgc gcaaaaattc ccacaaggga tgaattggga agtctttttt gatgcctccg
     2221 tgtttatcga aaactccata gatgaggtgg taaaaacctt agttgaagcg cttattttgg
     2281 ttattcttgt ggtgttcatg tttttacaga atattcgcgc aacactcata cccgccattg
     2341 cggtgcctgt ttcattaatt ggcacattag ccgcaatgct tgcctttggt tttaccatca
     2401 atactgtgag tttgttagcg ctagtgctgg caattggtat tgtggtcgat gatgccattg
     2461 ttgtggttga aaatgttgaa cgtttaatgg atgagaaagg gctgagttca tcgcaggcga
     2521 ctaaggttgc tatgaaggaa ctgtctggtg cgcttatcgc taccagccta gtgcttgcgg
     2581 cggtatttgt cccagtgtct ttcttgtctg ggatcaccgg tattatgtat cgagagtttg
     2641 ctgtcgcgat aacggttgcg gtactgatct caaccgtagt tgcactgaca ctttcgccag
     2701 cactttgtgc gttattactc aagcccggtg ataaagcgac ctcaggcttt ttcaagtgga
     2761 tgaatgatcg tttagacaca gccacaacta agtatgtcag attggtggtg ttaaccaaca
     2821 agcacgctaa acgcagttac ctgttgtttg ccttaatggt gggcggtgtg tacttaacaa
     2881 tgagttcgct gccatcgagc tttatgccag atgaagatca gggtcgattt tttatcgatg
     2941 tgtctctgcc taatggggca acggttaatc gtactcaaga tgtccttaag aaagccgaag
     3001 cgacagtgct tgctcatcct gcgatagctt actcgtttac cctagcgggt gaaaacagac
     3061 gctcaggttc gaatcaagca aacggtcagt ttgagatcat tctaaaacct tggtcagatc
     3121 gcgttgataa tgatgcaacc gtacagaagg tgatgaacga gattaaacaa tcactgcatg
     3181 atgtgctcga ggctgaattc agaatttatt taccctcggc agtcccaggt cttggtaatg
     3241 gctccggtgt tgaaatggag ctgcaagata cttcagggag taactttaaa ggcttaatgg
     3301 agaccgctga tgagctggtt gaagccttaa aactgcaacc tgagatagcc acggcaggat
     3361 tatcgctgca atctgcaatt ccgcagttac accttaatgt tgatgaagct aaagccatgg
     3421 ccattggggt taaggtgtct gatatttatg gcaccattaa aacatttaca gactcatcga
     3481 cggttaatga cttcaattta tttgggcgag tgtaccgagt gaaagtacaa gcagaagagc
     3541 agtatcgtca atttccggat caaatagagg attaccatgt tcgctcctca tcgggcgcta
     3601 tggtaccaat tggtgtatta gctgaatcta attactcggt agggccagca gcagtcacgc
     3661 actacaacat gtttactagc gcttcgatta atgcatctcc tgcaccgggt tatgcctcag
     3721 gcgatgtgat ccgcgctatt gagcgggtgg caaagcccat gctacctgac gagtttagct
     3781 atgagtggac cggaataacc tatcaagagg tgcagtctgc caatcaaacg gctattgcgg
     3841 ttacgttagc gatggtattt gtgttcctgt ttctcgctgc gctttatgag agctggactc
     3901 tgccgatagc ggtattgctc atcgccccca ttgcgatgtt aggtgcatct gtggggacct
     3961 tagtgagcgg tatggaaagc aatctctttt ttcaggtggc gtttatcgca ctgatcggaa
     4021 tggcggcaaa aaactcgata ttgattgtcg agtttgccaa ccaattacat aaagcaggta
     4081 agagccgact cgattcagcc attgaggcgg ctaatatgcg ttttagaccg attttaatga
     4141 cctcgttagc cttcattcta ggcgtacttc cactggtgtt ttctgtcggg cctggcgctg
     4201 tcagtcggca aagtatctct atccctattt tatgtggcat gattttcgcg acaaccattg
     4261 gcatcataat ggtaccgctg ttttttgtga caacggcggg ctgggttcgc tctaaaatca
     4321 ccggcaaagc cgatacagag gaggtatcga ctcatgccta atcaacttaa catcactcaa
     4381 gcctatacca atcataagat gacaatcggt ggtgttaact cgtttcgaat tagcaccaca
     4441 gcattttgtg tcgcctgtgg gattttgcta ctttctggtt gtgccatggg gccagattac
     4501 cagagaccag atcttgaaat accggaacaa tttcaccacg agttgcagca aaccgacgag
     4561 cagaacgttg gcctcaagca ttggcgtgag ttttatctcg atcctgagct acaggtattg
     4621 atagaacatg ctcttgatca aaacctcgat ttagagggag tgcgctcaag actgttagcg
     4681 gcaagagcgg gggtaacagt taccgacgct gcactatttc ccgtagtagg acttgatgca
     4741 aatggcgagc gagcattaga cagcgcgttg accagtaccg atccggttaa agaagatact
     4801 ttttatgttg cgggtactgt ttcgtgggaa ctggatcttt ggggagcgaa tcgtcgcagg
     4861 agtgaagcag aatttgccaa tttcatctct tcacaagaag cacttaattt agccaccatc
     4921 agtttaatca gtgatgtcgc cagtcgttat tatgaatggc tcgatattga gcaacgttat
     4981 caaatatcgc ttaatacggt cgagcttcgt aaaaaggagc gagatcttgc cagactgcga
     5041 aaacaaaatg gtgtgatatc gggcctgcaa gttaggcagg ctgaggtgga gtatcaaagt
     5101 gccagagtca cattgccaga tcttgattat gagcgaagtg aaaaggccaa tcagttgcgg
     5161 attttactgg gggaatttaa atatcccctt atccctaaaa ctgaagcgca gttaaaagct
     5221 gaaggagcgt tgtttcctga ctcattcgca gtgggggttc catcacagat gttgtcacag
     5281 cgcccagatg taaaatccgc agagcaagct cttgttgcag ccaatgccaa tgtcggtgtc
     5341 gctaaaacag cattctttcc taaatttacc attagcggca gctatggttc tgaatctgaa
     5401 agcctcagtg acatatttga tagccaaggg atcacttggt cgcttttagg tggtatcacc
     5461 gcccccattt tcaatgcagg taagatcact gctcaatatg atattgccca ggaagatgca
     5521 aaacaggcga tgctcagcta tagaagtaca gtgctcggtg catactttga agtgaatgat
     5581 gcgatgaaca gtttcagacg ctctgaattg gcgattgacg ctcagcagga attagttgtg
     5641 gcatcaactg aatatacgcg cttagcaacg ttaagatacc gaaatggggt ttcgagttca
     5701 ttggatttga tggatgctca gcgaagtctg tttagcgccg aactgactct gagtcaggtg
     5761 aaacgtgacc agctactgtc gatgattaca ctttatcgtg cgctgggtgg tggagtactg
     5821 accagtgatg agatgaagac tgaacttgaa aaaacgactg gcgagcaaca agtcgaagaa
     5881 cttactaaac atcatcacca tcaccactga ggatccaaac tcgagtaagg atctccaggc
     5941 atcaaataaa acgaaaggct cagtcgaaag actgggcctt tcgttttatc tgttgtttgt
     6001 cggtgaacgc tctctactag agtcacactg gctcaccttc gggtgggcct ttctgcgttt
     6061 atacctaggg atatattccg cttcctcgct cactgactcg ctacgctcgg tcgttcgact
     6121 gcggcgagcg gaaatggctt acgaacgggg cggagatttc ctggaagatg ccaggaagat
     6181 acttaacagg gaagtgagag ggccgcggca aagccgtttt tccataggct ccgcccccct
     6241 gacaagcatc acgaaatctg acgctcaaat cagtggtggc gaaacccgac aggactataa
     6301 agataccagg cgtttccccc tggcggctcc ctcgtgcgct ctcctgttcc tgcctttcgg
     6361 tttaccggtg tcattccgct gttatggccg cgtttgtctc attccacgcc tgacactcag
     6421 ttccgggtag gcagttcgct ccaagctgga ctgtatgcac gaaccccccg ttcagtccga
     6481 ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggaaagac atgcaaaagc
     6541 accactggca gcagccactg gtaattgatt tagaggagtt agtcttgaag tcatgcgccg
     6601 gttaaggcta aactgaaagg acaagttttg gtgactgcgc tcctccaagc cagttacctc
     6661 ggttcaaaga gttggtagct cagagaacct tcgaaaaacc gccctgcaag gcggtttttt
     6721 cgttttcaga gcaagagatt acgcgcagac caaaacgatc tcaagaagat catcttatta
     6781 atcagataaa atatttctag atttcagtgc aatttatctc ttcaaatgta gcacctgaag
     6841 tcagccccat acgatataag ttgttactag tgcttggatt ctcaccaata aaaaacgccc
     6901 ggcggcaacc gagcgttctg aacaaatcca gatggagttc tgaggtcatt actggatcta
     6961 tcaacaggag tccaagcgag ctctcgaacc ccagagtccc gctcagaaga actcgtcaag
     7021 aaggcgatag aaggcgatgc gctgcgaatc gggagcggcg ataccgtaaa gcacgaggaa
     7081 gcggtcagcc cattcgccgc caagctcttc agcaatatca cgggtagcca acgctatgtc
     7141 ctgatagcgg tccgccacac ccagccggcc acagtcgatg aatccagaaa agcggccatt
     7201 ttccaccatg atattcggca agcaggcatc gccatgggtc acgacgagat cctcgccgtc
     7261 gggcatgcgc gccttgagcc tggcgaacag ttcggctggc gcgagcccct gatgctcttc
     7321 gtccagatca tcctgatcga caagaccggc ttccatccga gtacgtgctc gctcgatgcg
     7381 atgtttcgct tggtggtcga atgggcaggt agccggatca agcgtatgca gccgccgcat
     7441 tgcatcagcc atgatggata ctttctcggc aggagcaagg tgagatgaca ggagatcctg
     7501 ccccggcact tcgcccaata gcagccagtc ccttcccgct tcagtgacaa cgtcgagcac
     7561 agctgcgcaa ggaacgcccg tcgtggccag ccacgatagc cgcgctgcct cgtcctgcag
     7621 ttcattcagg gcaccggaca ggtcggtctt gacaaaaaga accgggcgcc cctgcgctga
     7681 cagccggaac acggcggcat cagagcagcc gattgtctgt tgtgcccagt catagccgaa
     7741 tagcctctcc acccaagcgg ccggagaacc tgcgtgcaat ccatcttgtt caatcatgcg
     7801 aaacgatcct catcctgtct cttgatcaga tcatgatccc ctgcgccatc agatccttgg
     7861 cggcaagaaa gccatccagt ttactttgca gggcttccca accttaccag agggcgcccc
     7921 agctggcaat tccgacgtcg gtgcctaatg agtgagctaa cttacattaa ttgcgttgcg
     7981 ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca
     8041 acgcgcgggg agaggcggtt tgcgtattgg gcgccagggt ggtttttctt ttcaccagtg
     8101 agacgggcaa cagctgattg cccttcaccg cctggccctg agagagttgc agcaagcggt
     8161 ccacgctggt ttgccccagc aggcgaaaat cctgtttgat ggtggttaac ggcgggatat
     8221 aacatgagct gtcttcggta tcgtcgtatc ccactaccga gatgtccgca ccaacgcgca
     8281 gcccggactc ggtaatggcg cgcattgcgc ccagcgccat ctgatcgttg gcaaccagca
     8341 tcgcagtggg aacgatgccc tcattcagca tttgcatggt ttgttgaaaa ccggacatgg
     8401 cactccagtc gccttcccgt tccgctatcg gctgaatttg attgcgagtg agatatttat
     8461 gccagccagc cagacgcaga cgcgccgaga cagaacttaa tgggcccgct aacagcgcga
     8521 tttgctggtg acccaatgcg accagatgct ccacgcccag tcgcgtaccg tcttcatggg
     8581 agaaaataat actgttgatg ggtgtctggt cagagacatc aagaaataac gccggaacat
     8641 tagtgcaggc agcttccaca gcaatggcat cctggtcatc cagcggatag ttaatgatca
     8701 gcccactgac gcgttgcgcg agaagattgt gcaccgccgc tttacaggct tcgacgccgc
     8761 ttcgttctac catcgacacc accacgctgg cacccagttg atcggcgcga gatttaatcg
     8821 ccgcgacaat ttgcgacggc gcgtgcaggg ccagactgga ggtggcaacg ccaatcagca
     8881 acgactgttt gcccgccagt tgttgtgcca cgcggttggg aatgtaattc agctccgcca
     8941 tcgccgcttc cactttttcc cgcgttttcg cagaaacgtg gctggcctgg ttcaccacgc
     9001 gggaaacggt ctgataagag acaccggcat actctgcgac atcgtataac gttactggtt
     9061 tcacattcac caccctgaat tgactctctt ccgggcgcta tcatgccata ccgcgaaagg
     9121 ttttgcgcca ttcgatggtg tccgggatct cgacgctctc ccttatgcga ctcctgcatt
     9181 aggaagcagc ccagtagtag gttgaggccg ttgagcaccg ccgccgcaag gaatggtgca
     9241 tgcaaggaga tggcgcccaa cagtcccccg gccacggggc ctgccaccat acccacgccg
     9301 aaacaagcgc tcatgagccc gaagtggcga gcccgatctt ccccatcggt gatgtcggcg
     9361 atataggcgc cagcaaccgc acctgtggcg ccggtgatgc cggccacgat gcgtccggcg
     9421 tagaggatcg agatcgttta ggcaccccag gctttacact ttatgcttcc ggctcgtata
     9481 atgtgtggaa ttgtgagcgg ataacaattt cagaattcaa aagatctttt aagaaggaga
     9541 tatacat
//
