LOCUS       sequence-181402-        5987 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 5987)
  AUTHORS   Kaper T, Lager I, Looger LL, Chermak D, Frommer WB
  TITLE     Fluorescence Resonance Energy Transfer sensors for quantitative 
            monitoring of pentose and disaccharide accumulation in bacteria.
  JOURNAL   Biotechnol Biofuels. 2008 Jun 3. 1(1):11.
  PUBMED    18522753
REFERENCE   2  (bases 1 to 5987)
  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: "Biotechnol 
            Biofuels. 2008 Jun 3. 1(1):11."
COMMENT     Sequence Label: pGWF1
FEATURES             Location/Qualifiers
     source          1..5987
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     84..101
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     186..205
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        186..204
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             236..258
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             278..295
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             299..331
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="MASMTGGQQMG"
     CDS             join(335..343,344..358)
                     /codon_start=1
                     /product="Xpress(TM) epitope tag, including an enterokinase
                     recognition and cleavage site"
                     /label=Xpress(TM) tag
                     /translation="DLYDDDDK"
     CDS             join(371..373,374..376,377..889)
                     /codon_start=1
                     /product="N-terminal fragment of mVenus for use in 
                     bimolecular fluorescence complementation (BiFC) (Kodama and
                     Hu, 2010)"
                     /label=VN173
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIK
                     ANFKIRHNIE"
     protein_bind    1070..1194
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(1219..1224,1225..1242,1243..1249)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             1303..1962
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(1369..1388)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             2304..2609
                     /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     2488..2507
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(2650..2774)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             join(2782..2784,2785..3498)
                     /codon_start=1
                     /product="yellow fluorescent protein (YFP) with fast and 
                     efficient maturation (Nagai et al., 2002)"
                     /label=Venus
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     LICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIKA
                     NFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(3554..3572)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      3568..3615
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      3712..4167
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3799..3818)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4009..4030
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4193..4297
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4298..4366,4367..5158)
                     /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     complement(4516..4535)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5329..5917
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5818..5837
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc
       61 gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa gagcgcccaa
      121 tacgcaaacc gcctctcccc gcgcgttggc cgattcatta atgcaggatc tcgatcccgc
      181 gaaattaata cgactcacta tagggagacc acaacggttt ccctctagaa ataattttgt
      241 ttaactttaa gaaggagata tacatatgcg gggttctcat catcatcatc atcatggtat
      301 ggctagcatg actggtggac agcaaatggg tcgggatctg tacgacgatg acgataagga
      361 tccgggccgc atggtgagca agggcgagga gctgttcacc ggggtggtgc ccatcctggt
      421 cgagctggac ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg gcgagggcga
      481 tgccacctac ggcaagctga ccctgaagtt catctgcacc accggcaagc tgcccgtgcc
      541 ctggcccacc ctcgtgacca ccctgacctg gggcgtgcag tgcttcagcc gctaccccga
      601 ccacatgaag cagcacgact tcttcaagtc cgccatgccc gaaggctacg tccaggagcg
      661 caccatcttc ttcaaggacg acggcaacta caagacccgc gccgaggtga agttcgaggg
      721 cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg acggcaacat
      781 cctggggcac aagctggagt acaactacat cagccacaac gtctatatca ccgccgacaa
      841 gcagaagaac ggcatcaagg ccaacttcaa gatccgccac aacatcgagg acggcagcgt
      901 gcagctcgcc gaccactacc agcagaacac ccccatcggc gacggccccg tgctgctgcc
      961 cgacaaccac tacctgagca cccagtccgc cctgagcaaa gaccccaacg agaagcgcga
     1021 tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc actgatatca caagtttgta
     1081 caaaaaagct gaacgagaaa cgtaaaatga tataaatatc aatatattaa attagatttt
     1141 gcataaaaaa cagactacat aatactgtaa aacacaacat atccagtcac tatggcggcc
     1201 gcattaggca ccccaggctt tacactttat gcttccggct cgtataatgt gtggattttg
     1261 agttaggatc cgtcgagatt ttcaggagct aaggaagcta aaatggagaa aaaaatcact
     1321 ggatatacca ccgttgatat atcccaatgg catcgtaaag aacattttga ggcatttcag
     1381 tcagttgctc aatgtaccta taaccagacc gttcagctgg atattacggc ctttttaaag
     1441 accgtaaaga aaaataagca caagttttat ccggccttta ttcacattct tgcccgcctg
     1501 atgaatgctc atccggaatt ccgtatggca atgaaagacg gtgagctggt gatatgggat
     1561 agtgttcacc cttgttacac cgttttccat gagcaaactg aaacgttttc atcgctctgg
     1621 agtgaatacc acgacgattt ccggcagttt ctacacatat attcgcaaga tgtggcgtgt
     1681 tacggtgaaa acctggccta tttccctaaa gggtttattg agaatatgtt tttcgtctca
     1741 gccaatccct gggtgagttt caccagtttt gatttaaacg tggccaatat ggacaacttc
     1801 ttcgcccccg ttttcaccat gggcaaatat tatacgcaag gcgacaaggt gctgatgccg
     1861 ctggcgattc aggttcatca tgccgtttgt gatggcttcc atgtcggcag aatgcttaat
     1921 gaattacaac agtactgcga tgagtggcag ggcggggcgt aaacgcgtgg atccggctta
     1981 ctaaaagcca gataacagta tgcgtatttg cgcgctgatt tttgcggtat aagaatatat
     2041 actgatatgt atacccgaag tatgtcaaaa agaggtatgc tatgaagcag cgtattacag
     2101 tgacagttga cagcgacagc tatcagttgc tcaaggcata tatgatgtca atatctccgg
     2161 tctggtaagc acaaccatgc agaatgaagc ccgtcgtctg cgtgccgaac gctggaaagc
     2221 ggaaaatcag gaagggatgg ctgaggtcgc ccggtttatt gaaatgaacg gctcttttgc
     2281 tgacgagaac aggggctggt gaaatgcagt ttaaggttta cacctataaa agagagagcc
     2341 gttatcgtct gtttgtggat gtacagagtg atattattga cacgcccggg cgacggatgg
     2401 tgatccccct ggccagtgca cgtctgctgt cagataaagt ctcccgtgaa ctttacccgg
     2461 tggtgcatat cggggatgaa agctggcgca tgatgaccac cgatatggcc agtgtgccgg
     2521 tctccgttat cggggaagaa gtggctgatc tcagccaccg cgaaaatgac atcaaaaacg
     2581 ccattaacct gatgttctgg ggaatataaa tgtcaggctc ccttatacac agccagtctg
     2641 caggtcgacc atagtgactg gatatgttgt gttttacagt attatgtagt ctgtttttta
     2701 tgcaaaatct aatttaatat attgatattt atatcatttt acgtttctcg ttcagctttc
     2761 ttgtacaaag tggtgatatc ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc
     2821 atcctggtcg agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc
     2881 gagggcgatg ccacctacgg caagctgacc ctgaagctga tctgcaccac cggcaagctg
     2941 cccgtgccct ggcccaccct cgtgaccacc ctgggctacg gcctgcagtg cttcgcccgc
     3001 taccccgacc acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc
     3061 caggagcgca ccatcttctt caaggacgac ggcaactaca agacccgcgc cgaggtgaag
     3121 ttcgagggcg acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac
     3181 ggcaacatcc tggggcacaa gctggagtac aactacaaca gccacaacgt ctatatcacc
     3241 gccgacaagc agaagaacgg catcaaggcc aacttcaaga tccgccacaa catcgaggac
     3301 ggcggcgtgc agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg
     3361 ctgctgcccg acaaccacta cctgagctac cagtccgccc tgagcaaaga ccccaacgag
     3421 aagcgcgatc acatggtcct gctggagttc gtgaccgccg ccgggatcac tctcggcatg
     3481 gacgagctgt acaagtaaaa gcttgatccg gctgctaaca aagcccgaaa ggaagctgag
     3541 ttggctgctg ccaccgctga gcaataacta gcataacccc ttggggcctc taaacgggtc
     3601 ttgaggggtt ttttgctgaa aggaggaact atatccggat ctggcgtaat agcgaagagg
     3661 cccgcaccga tcgcccttcc caacagttgc gcagcctgaa tggcgaatgg gacgcgccct
     3721 gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg
     3781 ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg
     3841 gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac
     3901 ggcacctcga ccccaaaaaa cttgattagg gtgttggttc acgtagtggg ccatcgccct
     3961 gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt
     4021 tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta taagggattt
     4081 tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt
     4141 ttaacaaaat attaacgctt acaatttagg tggcactttt cggggaaatg tgcgcggaac
     4201 ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc
     4261 ctgataaatg cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt
     4321 cgcccttatt cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct
     4381 ggtgaaagta aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga
     4441 tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag
     4501 cacttttaaa gttctgctat gtggcgcggt attatcccgt attgacgccg ggcaagagca
     4561 actcggtcgc cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga
     4621 aaagcatctt acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag
     4681 tgataacact gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc
     4741 ttttttgcac aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa
     4801 tgaagccata ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg caacaacgtt
     4861 gcgcaaacta ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg
     4921 gatggaggcg gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt
     4981 tattgctgat aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg
     5041 gccagatggt aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat
     5101 ggatgaacga aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact
     5161 gtcagaccaa gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa
     5221 aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt
     5281 ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt
     5341 ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg
     5401 tttgccggat caagagctac caactctttt tccgaaggta actggcttca gcagagcgca
     5461 gataccaaat actgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt
     5521 agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga
     5581 taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc
     5641 gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact
     5701 gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga
     5761 caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg
     5821 aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt
     5881 tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt
     5941 acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctg
//
