LOCUS       sequence-64011-e        7918 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 7918)
  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 7918)
  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-EPL43
FEATURES             Location/Qualifiers
     source          1..7918
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             4261..4278
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      4314..4385
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      4401..4428
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     primer_bind     complement(4456..4473)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4590..5135)
                     /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(4670..4689)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(5249..5343)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(5374..6168)
                     /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(5485..5504)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     6095..6114
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     CDS             complement(6353..7435)
                     /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"
     promoter        complement(7436..7513)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     7719..7738
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        join(7824..7829,7830..7847,7848..7854)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    7862..7878
                     /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 atgtcgaata aactctttgc gccattctgc ctggtcgttc tggcagcggc gctgagcgcg
       61 tgtggcaaga ccgaaggcga gcagacgccg ccaccggcca aggtacgaat cgaaaccgta
      121 cagagccagg ccctgtccat caccagcgaa ctgagcgggc gcatcgccgc gccacggatc
      181 gccgaagtcc gcgcccgggt ggccggggtg gtgctgcaac ggaccttccg cgaaggctcc
      241 gatgtcaagc agggcgaggt cttgttccgc atcgacccgg ccccgttcaa ggctgacctg
      301 gacagcgccc aggccagcct gcgcaaggcc gaggccaccg ccttccaggc gcagttgcag
      361 gagcagcgct attccaagct ggtcgccgac caggccatca gcggccagga atacgacaat
      421 gcccgggctg cggcacgcca ggccagcgcc gacgtcgccg ccaaccaggc cgcggtgcag
      481 cgggccaagc tcaacctcgg ctatgccacc gtcaccgcgc cgatttccgg ccgggtcggc
      541 cgcgccctgg tcaccgaagg cgccctggtg ggccagaacg agaccacgcc gctggcgctg
      601 atccagcaac tggacccgat ccacgccgac ctgacccagt ccacccgtga actcaacgac
      661 ctgcgccggg ccttccgcgc cggccagttg cagcaggtgg gcaaggacca ggccaaggcg
      721 accctgatcc aggacgacgg cagcctctac ccgctgccgg gcaagctgtt gttcgccgat
      781 atcagcgttg acccgagtac cgggcagatc atcctgcgca gcgagttccc caacccggac
      841 ctcgacctgc tgcccggcag tttcgtgcgc gtgcgcctgg aacaggcggt caaccagcaa
      901 ggcatcagcg tgccgcagcg ggccatccag cgcgacagcg ccggcatcgc ccaggtgctg
      961 ctggtggatg cgcaaaacaa cgtggtgcag cacccggtgc aactgggtgg agtgcagaac
     1021 gatcgctgga tcgtcaccga gggcctcaag cccggtgacc gggtgattgt cgaaggcctg
     1081 caacacgccc gccccggcga gccggtggaa atcgacgcca gcccccttgc gcacgtccag
     1141 aaccctggcc agtaggtagt acacatgccg caattcttta ttgaccgccc ggtattcgcc
     1201 tgggtggtcg ccttcttcat cctgttggcc ggcgccctgg cgatccccca actgccggtg
     1261 gcgcaatacc ccaccgtggc cccgccgcag atcgagatct acgccgccta cccgggcgcc
     1321 tcggcgcaga ccctggacga atcggtggtc agcctgatcg agcaggagct caatggtgcc
     1381 gaccacctgc tgtacttctc gtcccagagc agcctcggca gcgccaccat caccgccacc
     1441 ttccagccgg gtaccaaccc ggaactggcc caggtggacg tgcagaaccg tctcaaggca
     1501 gtggaaccgc gcctgcccca ggcggtgacc cagcagggcc tgcaggtgga gaaagtctcc
     1561 gccggcttcc tcttgctgat caccctgacc tccagtgacg gccgcctgaa cgatgtcgcc
     1621 ctcagcgact acctggcgcg caacgtgatg aacgagatcc gccgcctcga tggcgtcggc
     1681 aaggcccagt tgtacggtgc cgagcgggcc atgcggatct ggctcgaccc gcagaaactg
     1741 atcggtttca acctgacccc ggccgacgtc aacgccgcgg tggcggcgca gaacgcccag
     1801 gttccggccg gcagcatcgg cgacctgccc aaccccgcca gccaggaaat caccgcgacc
     1861 atcctggtca agggccagct gtccagccct gaagagtttg ccgatatcgt cctgcgggcc
     1921 aaccccgacg gctccacggt acgcatcggc gatgtggccc gggtggaagt cggcagccag
     1981 gaataccagt acggcacccg cctgaacggc aagccgtcca ccgcggtggg cgtgcagttg
     2041 tcccccgggg ccaacgcgtt gaataccgcg accctgatcc gcgccaagat ggacgagctg
     2101 tcgcgctatt tcccggccgg cgtggaatac aagatcccct acgacacctc gcccttcgtc
     2161 aaggtctcga tcaccaaggt gctctacacc ctggccgaag cgatggtgct ggtgttcctg
     2221 gtgatgttcc tgttcctgca gaacatccgc tacaccctga tccccaccct ggtggtgccc
     2281 gtggccctga tgggcacctt cgccaccatg ctggccctgg ggttctcgat caacgtgctg
     2341 accatgttcg gcatggtgct ggccatcggc atcctggtgg acgatgccat cgtggtggtg
     2401 gagaacgtcg agcggatcat ggccaccgaa ggcctgtcgc ccaaggaagc cacacgcaag
     2461 gcgatgcagc agatcaccgg ggccatcatc ggcatcaccc tggtgctggt agcggtgttc
     2521 atccccatgg ccttcatgcc cggctcggtg ggggtgatct accagcagtt ctcgctgtcc
     2581 atggccacct cgatcctgtt ctcggcgttc ctcgccctga ccctgacccc ggccctgtgc
     2641 gccaccctgc tcaagcccat cgcggcgggc gaacaccatg aacgcacggg gttcttcggc
     2701 tggttcaacc ggcgcttcga acgcctgagc gacagctatc agggcggcgt gacctatgcg
     2761 ctcaagcgta ccggccgcta cctgctgatc tacctggcgc tgctggcgat catggcgctg
     2821 ctgttcagcc gcctgccctc ctccttcctg cccgtggagg accagggcta caccatcacc
     2881 gacatccagc tgccacctgg tgccagccag aaccgcacga tcaaggtggt ggagcagatc
     2941 gaagcgcaca acgccagcga accgggagtc ggcgacagca ccatcattct cggcttcagc
     3001 ttctcgggta acgggcagaa cgccgcgctg gccttcacca cgctcaagga ctggtcgcaa
     3061 cggagcagcg aggattcggc gatgtccatc gccgaccgcg ccaacgccgc cctcggcgcg
     3121 atcaaggatg ccgtggtgta cgccgtgctg ccgccgccga tcgatggcct gggcacctcc
     3181 agcggcttcg agttccgcct gcaggaccgt ggcggcgtcg gccacgcggc cctgatgcag
     3241 gcccgggacg agctgctcaa ggccacggaa aagagcccga tcctggccaa cgtccgggaa
     3301 agtgccctgg ccgaagcgcc gcaggtgcaa ctggaggtcg accgcaagca ggccaacgcc
     3361 ctgggcgtgt cctttgccga tatcggcagc ctgctgtcca ccgccctggg gtcgtcctac
     3421 gtcaacgact tccccaacca ggggcggatg cagcgggtgg tgatccaggc cgaaggcgac
     3481 cagcgcagcc aggtcgaaga cttgctgcgc ctgaacgtgc gcaacagcgc cggcaagatg
     3541 gtgccgctgt cggccttcgt ccgcgccagc tggagcctgg gcccggcgca actgacccgc
     3601 tacaacggct acccggccat tgccgtctcc ggcgagccgg cccctggcca cagtaccggt
     3661 gaagccatgc aggaaatcga acggctggtg gcgcagttgc ccaccgggct gggcctggaa
     3721 tggaccggcc tgtcactcca ggaacggctg tccggcagcc aggcgccgat gctgctgggc
     3781 ctgtcgctgc tgatcgtgtt cctgtgcctg gcggcgctct acgagagctg gtcgattccc
     3841 acttcggtgc tgctggtggt gcccctgggg gtactgggcg cagtgctggc ggtgagcctg
     3901 cgcggcatgc ccaacgacgt gttcttcaag gtcggcctga tcaccatcat cggcctgtcg
     3961 gcgaagaacg cgatcctgat catcgagttc gccaagagcc tgtacgacga aggccatgat
     4021 ctgatcgatg ccacggtgca ggctgcacgc ctgcgcctgc gccccatcgt catgacctcg
     4081 ctggcgttca tcctcggcgt ggtgcccctg gccatcgcca ccggcgccag ctccgccagc
     4141 cagcaggcga tcggtaccgg ggtgatcggc ggcatgatca ccgcaaccct ggcagtgatc
     4201 ttcgtgccgg tgttcttcgt ggtggtgatg aaactggtga aacgcggttc gtccaaggac
     4261 catcatcacc atcaccactg aggatccaaa ctcgagtaag gatctccagg catcaaataa
     4321 aacgaaaggc tcagtcgaaa gactgggcct ttcgttttat ctgttgtttg tcggtgaacg
     4381 ctctctacta gagtcacact ggctcacctt cgggtgggcc tttctgcgtt tatacctagg
     4441 gatatattcc gcttcctcgc tcactgactc gctacgctcg gtcgttcgac tgcggcgagc
     4501 ggaaatggct tacgaacggg gcggagattt cctggaagat gccaggaaga tacttaacag
     4561 ggaagtgaga gggccgcggc aaagccgttt ttccataggc tccgcccccc tgacaagcat
     4621 cacgaaatct gacgctcaaa tcagtggtgg cgaaacccga caggactata aagataccag
     4681 gcgtttcccc ctggcggctc cctcgtgcgc tctcctgttc ctgcctttcg gtttaccggt
     4741 gtcattccgc tgttatggcc gcgtttgtct cattccacgc ctgacactca gttccgggta
     4801 ggcagttcgc tccaagctgg actgtatgca cgaacccccc gttcagtccg accgctgcgc
     4861 cttatccggt aactatcgtc ttgagtccaa cccggaaaga catgcaaaag caccactggc
     4921 agcagccact ggtaattgat ttagaggagt tagtcttgaa gtcatgcgcc ggttaaggct
     4981 aaactgaaag gacaagtttt ggtgactgcg ctcctccaag ccagttacct cggttcaaag
     5041 agttggtagc tcagagaacc ttcgaaaaac cgccctgcaa ggcggttttt tcgttttcag
     5101 agcaagagat tacgcgcaga ccaaaacgat ctcaagaaga tcatcttatt aatcagataa
     5161 aatatttcta gatttcagtg caatttatct cttcaaatgt agcacctgaa gtcagcccca
     5221 tacgatataa gttgttacta gtgcttggat tctcaccaat aaaaaacgcc cggcggcaac
     5281 cgagcgttct gaacaaatcc agatggagtt ctgaggtcat tactggatct atcaacagga
     5341 gtccaagcga gctctcgaac cccagagtcc cgctcagaag aactcgtcaa gaaggcgata
     5401 gaaggcgatg cgctgcgaat cgggagcggc gataccgtaa agcacgagga agcggtcagc
     5461 ccattcgccg ccaagctctt cagcaatatc acgggtagcc aacgctatgt cctgatagcg
     5521 gtccgccaca cccagccggc cacagtcgat gaatccagaa aagcggccat tttccaccat
     5581 gatattcggc aagcaggcat cgccatgggt cacgacgaga tcctcgccgt cgggcatgcg
     5641 cgccttgagc ctggcgaaca gttcggctgg cgcgagcccc tgatgctctt cgtccagatc
     5701 atcctgatcg acaagaccgg cttccatccg agtacgtgct cgctcgatgc gatgtttcgc
     5761 ttggtggtcg aatgggcagg tagccggatc aagcgtatgc agccgccgca ttgcatcagc
     5821 catgatggat actttctcgg caggagcaag gtgagatgac aggagatcct gccccggcac
     5881 ttcgcccaat agcagccagt cccttcccgc ttcagtgaca acgtcgagca cagctgcgca
     5941 aggaacgccc gtcgtggcca gccacgatag ccgcgctgcc tcgtcctgca gttcattcag
     6001 ggcaccggac aggtcggtct tgacaaaaag aaccgggcgc ccctgcgctg acagccggaa
     6061 cacggcggca tcagagcagc cgattgtctg ttgtgcccag tcatagccga atagcctctc
     6121 cacccaagcg gccggagaac ctgcgtgcaa tccatcttgt tcaatcatgc gaaacgatcc
     6181 tcatcctgtc tcttgatcag atcatgatcc cctgcgccat cagatccttg gcggcaagaa
     6241 agccatccag tttactttgc agggcttccc aaccttacca gagggcgccc cagctggcaa
     6301 ttccgacgtc ggtgcctaat gagtgagcta acttacatta attgcgttgc gctcactgcc
     6361 cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg
     6421 gagaggcggt ttgcgtattg ggcgccaggg tggtttttct tttcaccagt gagacgggca
     6481 acagctgatt gcccttcacc gcctggccct gagagagttg cagcaagcgg tccacgctgg
     6541 tttgccccag caggcgaaaa tcctgtttga tggtggttaa cggcgggata taacatgagc
     6601 tgtcttcggt atcgtcgtat cccactaccg agatgtccgc accaacgcgc agcccggact
     6661 cggtaatggc gcgcattgcg cccagcgcca tctgatcgtt ggcaaccagc atcgcagtgg
     6721 gaacgatgcc ctcattcagc atttgcatgg tttgttgaaa accggacatg gcactccagt
     6781 cgccttcccg ttccgctatc ggctgaattt gattgcgagt gagatattta tgccagccag
     6841 ccagacgcag acgcgccgag acagaactta atgggcccgc taacagcgcg atttgctggt
     6901 gacccaatgc gaccagatgc tccacgccca gtcgcgtacc gtcttcatgg gagaaaataa
     6961 tactgttgat gggtgtctgg tcagagacat caagaaataa cgccggaaca ttagtgcagg
     7021 cagcttccac agcaatggca tcctggtcat ccagcggata gttaatgatc agcccactga
     7081 cgcgttgcgc gagaagattg tgcaccgccg ctttacaggc ttcgacgccg cttcgttcta
     7141 ccatcgacac caccacgctg gcacccagtt gatcggcgcg agatttaatc gccgcgacaa
     7201 tttgcgacgg cgcgtgcagg gccagactgg aggtggcaac gccaatcagc aacgactgtt
     7261 tgcccgccag ttgttgtgcc acgcggttgg gaatgtaatt cagctccgcc atcgccgctt
     7321 ccactttttc ccgcgttttc gcagaaacgt ggctggcctg gttcaccacg cgggaaacgg
     7381 tctgataaga gacaccggca tactctgcga catcgtataa cgttactggt ttcacattca
     7441 ccaccctgaa ttgactctct tccgggcgct atcatgccat accgcgaaag gttttgcgcc
     7501 attcgatggt gtccgggatc tcgacgctct cccttatgcg actcctgcat taggaagcag
     7561 cccagtagta ggttgaggcc gttgagcacc gccgccgcaa ggaatggtgc atgcaaggag
     7621 atggcgccca acagtccccc ggccacgggg cctgccacca tacccacgcc gaaacaagcg
     7681 ctcatgagcc cgaagtggcg agcccgatct tccccatcgg tgatgtcggc gatataggcg
     7741 ccagcaaccg cacctgtggc gccggtgatg ccggccacga tgcgtccggc gtagaggatc
     7801 gagatcgttt aggcacccca ggctttacac tttatgcttc cggctcgtat aatgtgtgga
     7861 attgtgagcg gataacaatt tcagaattca aaagatcttt taagaaggag atatacat
//
