LOCUS       sequence-12931-e        6401 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 6401)
  AUTHORS   Zhao Y, Terry D, Shi L, Weinstein H, Blanchard SC, Javitch JA
  TITLE     Single-molecule dynamics of gating in a neurotransmitter transporter
            homologue.
  JOURNAL   Nature. 2010 May 13. 465(7295):188-93.
  PUBMED    20463731
REFERENCE   2  (bases 1 to 6401)
  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: "Nature. 
            2010 May 13. 465(7295):188-93."
COMMENT     Sequence Label: pQO18-TEV
FEATURES             Location/Qualifiers
     source          1..6401
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(10..24,25..30,31..47,48..54)
                     /label=T5 promoter
                     /note="bacteriophage T5 promoter for E. coli RNA 
                     polymerase, with embedded lac operator"
     protein_bind    30..46
                     /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)."
     protein_bind    62..78
                     /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)."
     CDS             138..167
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             177..197
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     terminator      1854..1948
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     primer_bind     complement(2058..2077)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      2720..2806
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(2899..3981)
                     /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     3943..3962
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(3982..4059)
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     primer_bind     complement(4143..4165)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4251..4391
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4306..4325
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4406..4423)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4577..5165)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4657..4676)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5336..6127,6128..6196))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5959..5978
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6197..6301)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6369..6387
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 ctcgagaaat cataaaaaat ttatttgctt tgtgagcgga taacaattat aatagattca
       61 attgtgagcg gataacaatt tcacacagaa ttcgaaggaa ctacaaaata aattataagg
      121 aggcactcac catgtcgcat caccatcacc atcaccatca ccatcacgat gacgatgaaa
      181 acctgtattt tcagtccatg gaagtgaaac gcgaacattg ggcgacccgc ctgggcctga
      241 ttctggcgat ggcgggcaac gcggtgggcc tgggcaactt tctgcgcttt cctgtacagg
      301 cggcggaaaa cggcggcggc gcgtttatga ttccgtatat tattgcgttt ctgctggtgg
      361 gcattccgct gatgtggatt gaatgggcga tgggccgcta tggcggcgcg cagggccacg
      421 gcaccacacc tgcgattttt tatctgctgt ggcgcaaccg ctttgcgaaa attctgggcg
      481 tgtttggcct gtggattccg ctggtggtgg cgatttatta tgtgtatatt gaaagctgga
      541 ccctgggctt tgcgattaaa tttctggtgg gcctggtgcc ggaaccgccg ccgaacgcga
      601 cggatccgga tagcattctg cgcccgttta aagaatttct gtatagctat attggcgtgc
      661 cgaaaggcga tgaaccgatt ctgaaaccga gcctgtttgc gtatattgtg tttctgatta
      721 ccatgtttat taacgtgagc attctgattc gcggcattag caaaggcatt gagcgctttg
      781 cgaaaattgc gatgccgacc ctgtttattc tggcggtgtt tctggtgatt cgcgtgtttc
      841 tgctggaaac cccgaacggc accgctgcag atggcctgaa ctttctgtgg accccggatt
      901 ttgaaaaact gaaagatccg ggcgtgtgga ttgcggcggt gggccagatt ttttttaccc
      961 tgagcctggg ctttggcgcg attattacct atgcgagcta tgtacgtaaa gatcaggata
     1021 ttgtgctgag cggcctgacc gcggcgaccc tgaacgaaaa agcggaagtg attctgggcg
     1081 gcagcattag cattccggcg gcggtggcgt tttttggcgt ggcgaacgcg gtggcgattg
     1141 cgaaagcggg cgcgtttaac ctaggcttta ttaccctgcc ggcgattttt agccagaccg
     1201 cgggcggcac ctttctgggc tttctgtggt tttttctgct gttttttgcg ggcctgacca
     1261 gcagcattgc gattatgcag ccgatgattg cgtttctgga agatgaactt aagctgagcc
     1321 gcaaacatgc ggtgctgtgg accgcggcga ttgtgttttt tagcgcgcat ctggtgatgt
     1381 ttctgaacaa aagcctggat gaaatggatt tttgggcggg taccattggc gtggtgtttt
     1441 ttggcctgac cgaactgatt attttttttt ggatttttgg cgcggataaa gcgtgggagg
     1501 agattaaccg cggcggcatt attaaagtgc cgcgcattta ttattatgtg atgcgctata
     1561 ttaccccggc gtttctggcg gtgctgctgg tggtgtgggc ccgggaatat attccgaaaa
     1621 ttatggaaga aacccattgg accgtgtgga ttacccgctt ttatattatt ggcctgtttc
     1681 tgtttctgac ctttctggtg tttctggcgg aacgccgccg caaccatgaa agcgcgtaag
     1741 ctttctttga accaaaatta gaaaaccaag gcttgaaacg ttcaattgaa atggcaatta
     1801 aacaaattac agcacgtggt acaaagtggt gatagcttaa ttagctgagc ttggactcct
     1861 gttgatagat ccagtaatga cctcagaact ccatctggat ttgttcagaa cgctcggttg
     1921 ccgccgggcg ttttttattg gtgagaatcc aagctagctt ggcgagattt tcaggagcta
     1981 aggaagctaa aatggagaaa aaaatcactg gatataccac cgttgatata tcccaatggc
     2041 atcgtaaaga acattttgag gcatttcagt cagttgctca atgtacctat aaccagaccg
     2101 ttcagctgga tattacggcc tttttaaaga ccgtaaagaa aaataagcac aagttttatc
     2161 cggcctttat tcacattctt gcccgcctga tgaatgctca tccggaattt cgtatggcaa
     2221 tgaaagacgg tgagctggtg atatgggata gtgttcaccc ttgttacacc gttttccatg
     2281 agcaaactga aacgttttca tcgctctgga gtgaatacca cgacgatttc cggcagtttc
     2341 tacacatata ttcgcaagat gtggcgtgtt acggtgaaaa cctggcctat ttccctaaag
     2401 ggtttattga gaatatgttt ttcgtctcag ccaatccctg ggtgagtttc accagttttg
     2461 atttaaacgt ggccaatatg gacaacttct tcgcccccgt tttcaccatg catgggcaaa
     2521 tattatacgc aaggcgacaa ggtgctgatg ccgctggcga ttcaggttca tcatgccgtc
     2581 tgtgatggct tccatgtcgg cagaatgctt aatgaattac aacagtactg cgatgagtgg
     2641 cagggcgggg cgtaattttt ttaaggcagt tattggtgcc cttaaacgcc tggggtaatg
     2701 actctctagc ttgaggcatc aaataaaacg aaaggctcag tcgaaagact gggcctttcg
     2761 ttttatctgt tgtttgtcgg tgaacgctct cctgagtagg acaaatccgc cctctagatt
     2821 acgtgcagtc gatgataagc tgtcaaacat gagaattgtg cctaatgagt gagctaactt
     2881 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg
     2941 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ccagggtggt
     3001 ttttcttttc accagtgaga cgggcaacag ctgattgccc ttcaccgcct ggccctgaga
     3061 gagttgcagc aagcggtcca cgctggtttg ccccagcagg cgaaaatcct gtttgatggt
     3121 ggttaacggc gggatataac atgagctgtc ttcggtatcg tcgtatccca ctaccgagat
     3181 atccgcacca acgcgcagcc cggactcggt aatggcgcgc attgcgccca gcgccatctg
     3241 atcgttggca accagcatcg cagtgggaac gatgccctca ttcagcattt gcatggtttg
     3301 ttgaaaaccg gacatggcac tccagtcgcc ttcccgttcc gctatcggct gaatttgatt
     3361 gcgagtgaga tatttatgcc agccagccag acgcagacgc gccgagacag aacttaatgg
     3421 gcccgctaac agcgcgattt gctggtgacc caatgcgacc agatgctcca cgcccagtcg
     3481 cgtaccgtct tcatgggaga aaataatact gttgatgggt gtctggtcag agacatcaag
     3541 aaataacgcc ggaacattag tgcaggcagc ttccacagca atggcatcct ggtcatccag
     3601 cggatagtta atgatcagcc cactgacgcg ttgcgcgaga agattgtgca ccgccgcttt
     3661 acaggcttcg acgccgcttc gttctaccat cgacaccacc acgctggcac ccagttgatc
     3721 ggcgcgagat ttaatcgccg cgacaatttg cgacggcgcg tgcagggcca gactggaggt
     3781 ggcaacgcca atcagcaacg actgtttgcc cgccagttgt tgtgccacgc ggttgggaat
     3841 gtaattcagc tccgccatcg ccgcttccac tttttcccgc gttttcgcag aaacgtggct
     3901 ggcctggttc accacgcggg aaacggtctg ataagagaca ccggcatact ctgcgacatc
     3961 gtataacgtt actggtttca cattcaccac cctgaattga ctctcttccg ggcgctatca
     4021 tgccataccg cgaaaggttt tgcaccattc gatggtgtcg gaatttcggg cagcgttggg
     4081 tcctggccac gggtgcgcat gatctagagc tgcctcgcgc gtttcggtga tgacggtgaa
     4141 aacctctgac acatgcagct cccggagacg gtcacagctt gtctgtaagc ggatgccggg
     4201 agcagacaag cccgtcaggg cgcgtcagcg ggtgttggcg ggtgtcgggg cgcagccatg
     4261 acccagtcac gtagcgatag cggagtgtat actggcttaa ctatgcggca tcagagcaga
     4321 ttgtactgag agtgcaccat atgcggtgtg aaataccgca cagatgcgta aggagaaaat
     4381 accgcatcag gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc
     4441 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
     4501 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
     4561 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
     4621 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
     4681 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
     4741 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
     4801 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
     4861 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
     4921 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
     4981 tcttgaagtg gtggcctaac tacggctaca ctagaaggac agtatttggt atctgcgctc
     5041 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
     5101 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
     5161 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
     5221 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt
     5281 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc
     5341 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
     5401 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg
     5461 ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc
     5521 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta
     5581 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg
     5641 ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct
     5701 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta
     5761 gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg
     5821 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga
     5881 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt
     5941 gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca
     6001 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt
     6061 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt
     6121 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga
     6181 aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat cagggttatt
     6241 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc
     6301 gcacatttcc ccgaaaagtg ccacctgacg tctaagaaac cattattatc atgacattaa
     6361 cctataaaaa taggcgtatc acgaggccct ttcgtcttca c
//
