LOCUS       sequence-251606-        4644 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  K92A.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4644)
  AUTHORS   Ewald JC, Reich S, Baumann S, Frommer WB, Zamboni N
  TITLE     Engineering genetically encoded nanosensors for real-time in vivo 
            measurements of citrate concentrations.
  JOURNAL   PLoS One. 2011;6(12):e28245. doi: 10.1371/journal.pone.0028245. Epub
            2011 Dec 2.
  PUBMED    22164251
REFERENCE   2  (bases 1 to 4644)
  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: "PLoS One.";
            date: "2011"; volume: "6(12):e28245. doi"; pages: " 
            10.1371/journal.pone.0028245. Epub 2011 Dec 2"
COMMENT     Sequence Label: CIT32u
FEATURES             Location/Qualifiers
     source          1..4644
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     94..111
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     196..215
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        196..214
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             246..268
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             288..305
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             309..341
                     /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(345..353,354..368)
                     /codon_start=1
                     /product="Xpress(TM) epitope tag, including an enterokinase
                     recognition and cleavage site"
                     /label=Xpress(TM) tag
                     /translation="DLYDDDDK"
     CDS             join(384..386,387..899)
                     /codon_start=1
                     /product="N-terminal fragment of mVenus for use in 
                     bimolecular fluorescence complementation (BiFC) (Kodama and
                     Hu, 2010)"
                     /label=VN173
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIKA
                     NFKIRHNIE"
     CDS             1911..2162
                     /codon_start=1
                     /product="C-terminal fragment of mVenus for use in 
                     bimolecular fluorescence complementation (BiFC) (Kodama and
                     Hu, 2010)"
                     /label=VC155
                     /translation="DKQKNGIKANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNH
                     YLSYQSALSKDPNEKRDHMVLLEFVTAAGITLGMDELYK"
     primer_bind     complement(2221..2239)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2235..2282
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2379..2834
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2466..2485)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2676..2697
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2860..2964
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2965..3033,3034..3825)
                     /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(3183..3202)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3996..4584
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4485..4504
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct
       61 gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa
      121 gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc cgattcatta atgcaggatc
      181 tcgatcccgc gaaattaata cgactcacta tagggagacc acaacggttt ccctctagaa
      241 ataattttgt ttaactttaa gaaggagata tacatatgcg gggttctcat catcatcatc
      301 atcatggtat ggctagcatg actggtggac agcaaatggg tcgggatctg tacgacgatg
      361 acgataagga tccgggcctc gaggtgagca agggcgagga gctgttcacc ggggtggtgc
      421 ccatcctggt cgagctggac ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg
      481 gcgagggcga tgccacctac ggcaagctga ccctgaagtt catctgcacc accggcaagc
      541 tgcccgtgcc ctggcccacc ctcgtgacca ccctgacctg gggcgtgcag tgcttcagcc
      601 gctaccccga ccacatgaag cagcacgact tcttcaagtc cgccatgccc gaaggctacg
      661 tccaggagcg caccatcttc ttcaaggacg acggcaacta caagacccgc gccgaggtga
      721 agttcgaggg cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg
      781 acggcaacat cctggggcac aagctggagt acaactacat cagccacaac gtctatatca
      841 ccgccgacaa gcagaagaac ggcatcaagg ccaacttcaa gatccgccac aacatcgagg
      901 acggcagcgt gcagctcgcc gaccactacc agcagaacac ccccatcggc gacggccccg
      961 tgctgctgcc cgacaaccac tacctgagca cccagtccgc cctgagcaaa gaccccaacg
     1021 agaagcgcga tcacatggtc ctgctggagt tcgtgaccgc cgccgggggt acccgtctgc
     1081 attatcaggt cgggcagcgg gcgctgattc aggcaatgca gatttcggcg atgccggagc
     1141 tggtcgaagc ggttcagaaa cgcgatctcg ccagaatcaa agcgcttatc gatcccatgc
     1201 gttcgttctc cgacgccacc tatatcaccg tcggtgatgc caacggccag cgcctctatc
     1261 atgttaatcc tgatgaaatc ggcaaatcga tggagggcgg cgatagcgat gaggcgttga
     1321 ttaatgctgc aagctacgtg tcagtgcgca aaggctcgct gggctcctcg ctgcgcggta
     1381 aatcaccgat tcaggacgag accggcaagg tgattggcat tgtgtcggta ggctatacca
     1441 tcgagcaaac tagtaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc
     1501 tggacggcga cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca
     1561 cctacggcaa gctgaccctg aagctgatct gcaccaccgg caagctgccc gtgccctggc
     1621 ccaccctcgt gaccaccctg ggctacggcc tgcagtgctt cgcccgctac cccgaccaca
     1681 tgaagcagca cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca
     1741 tcttcttcaa ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca
     1801 ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg
     1861 ggcacaagct ggagtacaac tacaacagcc acaacgtcta tatcaccgcc gacaagcaga
     1921 agaacggcat caaggccaac ttcaagatcc gccacaacat cgaggacggc ggcgtgcagc
     1981 tcgccgacca ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca
     2041 accactacct gagctaccag tccgccctga gcaaagaccc caacgagaag cgcgatcaca
     2101 tggtcctgct ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtaca
     2161 agtaagggcc cgatccggct gctaacaaag cccgaaagga agctgagttg gctgctgcca
     2221 ccgctgagca ataactagca taaccccttg gggcctctaa acgggtcttg aggggttttt
     2281 tgctgaaagg aggaactata tccggatctg gcgtaatagc gaagaggccc gcaccgatcg
     2341 cccttcccaa cagttgcgca gcctgaatgg cgaatgggac gcgccctgta gcggcgcatt
     2401 aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc
     2461 gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca
     2521 agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc
     2581 caaaaaactt gattagggtg ttggttcacg tagtgggcca tcgccctgat agacggtttt
     2641 tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac
     2701 aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc
     2761 ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaatttta acaaaatatt
     2821 aacgcttaca atttaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta
     2881 tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt
     2941 caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc
     3001 ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa
     3061 gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt
     3121 aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt
     3181 ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc
     3241 atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg
     3301 gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg
     3361 gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac
     3421 atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca
     3481 aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta
     3541 actggcgaac tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat
     3601 aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa
     3661 tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag
     3721 ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat
     3781 agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt
     3841 tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg
     3901 aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga
     3961 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
     4021 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     4081 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
     4141 gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
     4201 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
     4261 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg
     4321 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
     4381 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta
     4441 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat
     4501 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg
     4561 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc
     4621 ttttgctggc cttttgctca catg
//
