LOCUS       sequence-7735-ex        7062 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 7062)
  AUTHORS   Scheich C, Kummel D, Soumailakakis D, Heinemann U, Bussow K
  TITLE     Vectors for co-expression of an unrestricted number of proteins.
  JOURNAL   Nucleic Acids Res. 2007;35(6):e43. doi: 10.1093/nar/gkm067. Epub 
            2007 Feb 20.
  PUBMED    17311810
REFERENCE   2  (bases 1 to 7062)
  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: "Nucleic 
            Acids Res."; date: "2007"; volume: "35(6):e43. doi"; pages: " 
            10.1093/nar/gkm067. Epub 2007 Feb 20"
COMMENT     Sequence Label: pQLinkHD
FEATURES             Location/Qualifiers
     source          1..7062
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(36..50,51..56,57..73,74..80)
                     /label=T5 promoter
                     /note="bacteriophage T5 promoter for E. coli RNA 
                     polymerase, with embedded lac operator"
     protein_bind    56..72
                     /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    88..104
                     /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)."
     RBS             123..134
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             147..167
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=7xHis
                     /translation="HHHHHHH"
     protein_bind    180..304
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             complement(733..1038)
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     complement(835..854)
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             complement(1383..2042)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     promoter        complement(2043..2145)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     protein_bind    complement(2323..2447)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     terminator      2469..2563
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             2657..3316
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     terminator      3381..3467
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(3560..4642)
                     /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     4604..4623
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(4643..4720)
                     /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(4804..4826)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4912..5052
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4967..4986
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(5067..5084)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5238..5826)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5318..5337)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5997..6788,6789..6857))
                     /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     6620..6639
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6858..6962)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7030..7048
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 ctcgagctta attaacaaca ccatttgtcg agaaatcata aaaaatttat ttgctttgtg
       61 agcggataac aattataata gattcaattg tgagcggata acaatttcac acagaattca
      121 ttaaagagga gaaattaact atgaaacatc accatcacca tcaccatggt gctgcaggta
      181 caagtttgta caaaaaagct gaacgagaaa cgtaaaatga tataaatatc aatatattaa
      241 attagatttt gcataaaaaa cagactacat aatactgtaa aacacaacat atccagtcac
      301 tatgaatcaa ctacttagat ggtattagtg acctgtagtc gaccgacagc cttccaaatg
      361 ttcttcgggt gatgctgcca acttagtcga ccgacagcct tccaaatgtt cttctcaaac
      421 ggaatcgtcg tatccagcct actcgctatt gtcctcaatg ccgtattaaa tcataaaaag
      481 aaataagaaa aagaggtgcg agcctctttt ttgtgtgaca aaataaaaac atctacctat
      541 tcatatacgc tagtgtcata gtcctgaaaa tcatctgcat caagaacaat ttcacaactc
      601 ttatactttt ctcttacaag tcgttcggct tcatctggat tttcagcctc tatacttact
      661 aaacgtgata aagtttctgt aatttctact gtatcgacct gcagactggc tgtgtataag
      721 ggagcctgac atttatattc cccagaacat caggttaatg gcgtttttga tgtcattttc
      781 gcggtggctg agatcagcca cttcttcccc gataacggag accggcacac tggccatatc
      841 ggtggtcatc atgcgccagc tttcatcccc gatatgcacc accgggtaaa gttcacggga
      901 gactttatct gacagcagac gtgcactggc cagggggatc accatccgtc gcccgggcgt
      961 gtcaataata tcactctgta catccacaaa cagacgataa cggctctctc ttttataggt
     1021 gtaaacctta aactgcattt caccagcccc tgttctcgtc agcaaaagag ccgttcattt
     1081 caataaaccg ggcgacctca gccatccctt cctgattttc cgctttccag cgttcggcac
     1141 gcagacgacg ggcttcattc tgcatggttg tgcttaccag accggagata ttgacatcat
     1201 atatgccttg agcaactgat agctgtcgct gtcaactgtc actgtaatac gctgcttcat
     1261 agcatacctc tttttgacat acttcgggta tacatatcag tatatattct tataccgcaa
     1321 aaatcagcgc gcaaatacgc atactgttat ctggctttta gtaagccgga tccacgcggc
     1381 gtttacgccc cgccctgcca ctcatcgcag tactgttgta attcattaag cattctgccg
     1441 acatggaagc catcacagac ggcatgatga acctgaatcg ccagcggcat cagcaccttg
     1501 tcgccttgcg tataatattt gcccatggtg aaaacggggg cgaagaagtt gtccatattg
     1561 gccacgttta aatcaaaact ggtgaaactc acccagggat tggctgagac gaaaaacata
     1621 ttctcaataa accctttagg gaaataggcc aggttttcac cgtaacacgc cacatcttgc
     1681 gaatatatgt gtagaaactg ccggaaatcg tcgtggtatt cactccagag cgatgaaaac
     1741 gtttcagttt gctcatggaa aacggtgtaa caagggtgaa cactatccca tatcaccagc
     1801 tcaccgtctt tcattgccat acggaattcc ggatgagcat tcatcaggcg ggcaagaatg
     1861 tgaataaagg ccggataaaa cttgtgctta tttttcttta cggtctttaa aaaggccgta
     1921 atatccagct gaacggtctg gttataggta cattgagcaa ctgactgaaa tgcctcaaaa
     1981 tgttctttac gatgccattg ggatatatca acggtggtat atccagtgat ttttttctcc
     2041 attttagctt ccttagctcc tgaaaatctc gataactcaa aaaatacgcc cggtagtgat
     2101 cttatttcat tatggtgaaa gttggaacct cttacgtgcc gatcaacgtc tcattttcgc
     2161 caaaagttgg cccagggctt cccggtatca acagggacac caggatttat ttattctgcg
     2221 aagtgatctt ccgtcacagg tatttattcg gcgcaaagtg cgtcgggtga tgctgccaac
     2281 ttagtcgact acaggtcact aataccatct aagtagttga ttcatagtga ctggatatgt
     2341 tgtgttttac agtattatgt agtctgtttt ttatgcaaaa tctaatttaa tatattgata
     2401 tttatatcat tttacgtttc tcgttcagct ttcttgtaca aagtggtaag cttaattagc
     2461 tgagcttgga ctcctgttga tagatccagt aatgacctca gaactccatc tggatttgtt
     2521 cagaacgctc ggttgccgcc gggcgttttt tattggtgag aatccaagta acaacaccat
     2581 ttaaatggag tggttacaaa tggagtggtt aattaaggct agcttggcga gattttcagg
     2641 agctaaggaa gctaaaatgg agaaaaaaat cactggatat accaccgttg atatatccca
     2701 atggcatcgt aaagaacatt ttgaggcatt tcagtcagtt gctcaatgta cctataacca
     2761 gaccgttcag ctggatatta cggccttttt aaagaccgta aagaaaaata agcacaagtt
     2821 ttatccggcc tttattcaca ttcttgcccg cctgatgaat gctcatccgg aatttcgtat
     2881 ggcaatgaaa gacggtgagc tggtgatatg ggatagtgtt cacccttgtt acaccgtttt
     2941 ccatgagcaa actgaaacgt tttcatcgct ctggagtgaa taccacgacg atttccggca
     3001 gtttctacac atatattcgc aagatgtggc gtgttacggt gaaaacctgg cctatttccc
     3061 taaagggttt attgagaata tgtttttcgt ctcagccaat ccctgggtga gtttcaccag
     3121 ttttgattta aacgtggcca atatggacaa cttcttcgcc cccgttttca ccatgggcaa
     3181 atattatacg caaggcgaca aggtgctgat gccgctggcg attcaggttc atcatgccgt
     3241 ttgtgatggc ttccatgtcg gcagaatgct taatgaatta caacagtact gcgatgagtg
     3301 gcagggcggg gcgtaatttt tttaaggcag ttattggtgc ccttaaacgc ctggggtaat
     3361 gactctctag cttgaggcat caaataaaac gaaaggctca gtcgaaagac tgggcctttc
     3421 gttttatctg ttgtttgtcg gtgaacgctc tcctgagtag gacaaatccg ccctctagat
     3481 tacgtgcagt cgatgataag ctgtcaaaca tgagaattgt gcctaatgag tgagctaact
     3541 tacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct
     3601 gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc gccagggtgg
     3661 tttttctttt caccagtgag acgggcaaca gctgattgcc cttcaccgcc tggccctgag
     3721 agagttgcag caagcggtcc acgctggttt gccccagcag gcgaaaatcc tgtttgatgg
     3781 tggttaacgg cgggatataa catgagctgt cttcggtatc gtcgtatccc actaccgaga
     3841 tatccgcacc aacgcgcagc ccggactcgg taatggcgcg cattgcgccc agcgccatct
     3901 gatcgttggc aaccagcatc gcagtgggaa cgatgccctc attcagcatt tgcatggttt
     3961 gttgaaaacc ggacatggca ctccagtcgc cttcccgttc cgctatcggc tgaatttgat
     4021 tgcgagtgag atatttatgc cagccagcca gacgcagacg cgccgagaca gaacttaatg
     4081 ggcccgctaa cagcgcgatt tgctggtgac ccaatgcgac cagatgctcc acgcccagtc
     4141 gcgtaccgtc ttcatgggag aaaataatac tgttgatggg tgtctggtca gagacatcaa
     4201 gaaataacgc cggaacatta gtgcaggcag cttccacagc aatggcatcc tggtcatcca
     4261 gcggatagtt aatgatcagc ccactgacgc gttgcgcgag aagattgtgc accgccgctt
     4321 tacaggcttc gacgccgctt cgttctacca tcgacaccac cacgctggca cccagttgat
     4381 cggcgcgaga tttaatcgcc gcgacaattt gcgacggcgc gtgcagggcc agactggagg
     4441 tggcaacgcc aatcagcaac gactgtttgc ccgccagttg ttgtgccacg cggttgggaa
     4501 tgtaattcag ctccgccatc gccgcttcca ctttttcccg cgttttcgca gaaacgtggc
     4561 tggcctggtt caccacgcgg gaaacggtct gataagagac accggcatac tctgcgacat
     4621 cgtataacgt tactggtttc acattcacca ccctgaattg actctcttcc gggcgctatc
     4681 atgccatacc gcgaaaggtt ttgcaccatt cgatggtgtc ggaatttcgg gcagcgttgg
     4741 gtcctggcca cgggtgcgca tgatctagag ctgcctcgcg cgtttcggtg atgacggtga
     4801 aaacctctga cacatgcagc tcccggagac ggtcacagct tgtctgtaag cggatgccgg
     4861 gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc gggtgtcggg gcgcagccat
     4921 gacccagtca cgtagcgata gcggagtgta tactggctta actatgcggc atcagagcag
     4981 attgtactga gagtgcacca catgcggtgt gaaataccgc acagatgcgt aaggagaaaa
     5041 taccgcatca ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg
     5101 ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg
     5161 gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag
     5221 gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga
     5281 cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct
     5341 ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc
     5401 tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg
     5461 gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc
     5521 tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca
     5581 ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag
     5641 ttcttgaagt ggtggcctaa ctacggctac actagaagga cagtatttgg tatctgcgct
     5701 ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc
     5761 accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga
     5821 tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca
     5881 cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat
     5941 taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac
     6001 caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt
     6061 gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt
     6121 gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag
     6181 ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct
     6241 attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt
     6301 gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc
     6361 tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt
     6421 agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg
     6481 gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg
     6541 actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct
     6601 tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc
     6661 attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt
     6721 tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt
     6781 tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg
     6841 aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta tcagggttat
     6901 tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg
     6961 cgcacatttc cccgaaaagt gccacctgac gtctaagaaa ccattattat catgacatta
     7021 acctataaaa ataggcgtat cacgaggccc tttcgtcttc ac
//
