LOCUS       sequence-7688-ex        5302 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 5302)
  AUTHORS   Deuschle K, Okumoto S, Fehr M, Looger LL, Kozhukh L, Frommer WB
  TITLE     Construction and optimization of a family of genetically encoded 
            metabolite sensors by semirational protein engineering.
  JOURNAL   Protein Sci. 2005 Sep . 14(9):2304-14.
  PUBMED    16131659
REFERENCE   2  (bases 1 to 5302)
  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: "Protein 
            Sci. 2005 Sep . 14(9):2304-14."
COMMENT     Sequence Label: pRSET FLII12Pglu-600u
FEATURES             Location/Qualifiers
     source          1..5302
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     19..38
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        19..37
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             67..89
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             109..126
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             130..162
                     /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(166..174,175..189)
                     /codon_start=1
                     /product="Xpress(TM) epitope tag, including an enterokinase
                     recognition and cleavage site"
                     /label=Xpress(TM) tag
                     /translation="DLYDDDDK"
     CDS             join(244..246,247..249,250..957)
                     /codon_start=1
                     /product="enhanced CFP"
                     /label=ECFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELY"
     CDS             join(1873..1875,1876..1878,1879..2589)
                     /codon_start=1
                     /product="enhanced variant of YFP (Heikal et al., 2001)"
                     /label=Citrine
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLMCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2648..2666)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2662..2709
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2806..3261
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2893..2912)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3103..3124
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3342..3446
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3447..3515,3516..4307)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCDTLLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDESDTTMPVAMPTTLRKLLTGELLTLASRQQLID*MEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKRSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      4478..5066
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4967..4986
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5220..5237
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 atctcgatcc cgcgaaatta atacgactca ctatagggag accacaacgg tttccctcta
       61 gataattttg tttaacttta agaaggagat atacatatgc ggggttctca tcatcatcat
      121 catcatggta tggctagcat gactggtgga cagcaaatgg gtcgggatct gtacgacgat
      181 gacgataagg atccgggccg cgctgatact cgcattggtg taacaatcta taagtcggct
      241 ggtatggtga gcaagggcga ggagctgttc accggggtgg tgcccatcct ggtcgagctg
      301 gacggcgacg taaacggcca caagttcagc gtgtccggcg agggcgaggg cgatgccacc
      361 tacggcaagc tgaccctgaa gttcatctgc accaccggca agctgcccgt gccctggccc
      421 accctcgtga ccaccctgac ctggggcgtg cagtgcttca gccgctaccc cgaccacatg
      481 aagcagcacg acttcttcaa gtccgccatg cccgaaggct acgtccagga gcgcaccatc
      541 ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg tgaagttcga gggcgacacc
      601 ctggtgaacc gcatcgagct gaagggcatc gacttcaagg aggacggcaa catcctgggg
      661 cacaagctgg agtacaacta catcagccac aacgtctata tcaccgccga caagcagaag
      721 aacggcatca aggccaactt caagatccgc cacaacatcg aggacggcag cgtgcagctc
      781 gccgaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct gcccgacaac
      841 cactacctga gcacccagtc cgccctgagc aaagacccca acgagaagcg cgatcacatg
      901 gtcctgctgg agttcgtgac cgccgccggg atcactctcg gcatggacga gctgtacggt
      961 agccgagata acgcgatgtc tgtagtgcgc aaggctattg agcaagatgc gaaagccgcg
     1021 ccagatgttc agctgctgat gaatgattct cagaatgacc agtccaagca gaacgatcag
     1081 atcgacgtat tgctggcgaa aggggtgaag gcactggcaa tcaacctggt tgacccggca
     1141 gctgcgggta cggtgattga gaaagcgcgt gggcaaaacg tgccggtggt tttcttcaac
     1201 aaagaaccgt ctcgtaaggc gctggatagc tacgacaaag cctactacgt tggcactgac
     1261 tccaaagagt ccggcattat tcaaggcgat ttgattgcta aacactgggc ggcgaatcag
     1321 ggttgggatc tgaacaaaga cggtcagatt cagttcgtac tgctgaaagg tgaaccgggc
     1381 catccggatg cagaagcacg taccacttac gtgattaaag aattgaacga taaaggcatc
     1441 aaaactgaac agttacagtt agataccgca atgtgggaca ccgctcaggc gaaagataag
     1501 atggacgcct ggctgtctgg cccgaacgcc aacaaaatcg aagtggttat cgccaacaac
     1561 gatgcgatgg caatgggcgc ggttgaagcg ctgaaagcac acaacaagtc cagcattccg
     1621 gtgtttggcg tcgatgcgct gccagaagcg ctggcgctgg tgaaatccgg tgcactggcg
     1681 ggcaccgtac tgaacgatgc taacaaccag gcgaaagcga cctttgatct ggcgaaaaac
     1741 ctggccgatg gtaaaggtgc ggctgatggc accaactgga aaatcgacaa caaagtggtc
     1801 cgcgtacctt atgttggcgt agataaagac aacctggctg aattcagcaa gaaaggcgcc
     1861 ggtaccggtg gaatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg
     1921 gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc
     1981 gatgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg
     2041 ccctggccca ccctcgtgac caccttcggc tacggcctga tgtgcttcgc ccgctacccc
     2101 gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag
     2161 cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag
     2221 ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac
     2281 atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat catggccgac
     2341 aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga ggacggcagc
     2401 gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg
     2461 cccgacaacc actacctgag ctaccagtcc gccctgagca aagaccccaa cgagaagcgc
     2521 gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag
     2581 ctgtacaagt aaaagcttga tccggctgct aacaaagccc gaaaggaagc tgagttggct
     2641 gctgccaccg ctgagcaata actagcataa ccccttgggg cctctaaacg ggtcttgagg
     2701 ggttttttgc tgaaaggagg aactatatcc ggatctggcg taatagcgaa gaggcccgca
     2761 ccgatcgccc ttcccaacag ttgcgcagcc tgaatggcga atgggacgcg ccctgtagcg
     2821 gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg
     2881 ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc
     2941 cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagagct ttacggcacc
     3001 tcgaccgcaa aaaacttgat ttgggtgatg gttcacgtag tgggccatcg ccctgataga
     3061 cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa
     3121 ctggaacaac actcaaccct atcgcggtct attcttttga tttataaggg attttgccga
     3181 tttcggccta ttggttaaaa aatgagctga tttaacaaat atttaacgcg aattttaaca
     3241 aaatattaac gtttacaatt tcgcctgatg cggtattttc tccttacgca tctgtgcggt
     3301 atttcacacc gcatacaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt
     3361 tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc
     3421 ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc
     3481 ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa
     3541 aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg
     3601 gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag
     3661 ttctgctatg tgatacacta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc
     3721 gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta
     3781 cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg
     3841 cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca
     3901 acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac
     3961 caaacgacga gagtgacacc acgatgcctg tagcaatgcc aacaacgttg cgcaaactat
     4021 taactggcga actacttact ctagcttccc ggcaacaatt aatagactga atggaggcgg
     4081 ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata
     4141 aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta
     4201 agcgctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa
     4261 atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag
     4321 tttactcata tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg
     4381 tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact
     4441 gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg
     4501 taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc
     4561 aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata
     4621 ctgtccttct agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta
     4681 catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc
     4741 ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg
     4801 ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac
     4861 agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg
     4921 taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt
     4981 atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct
     5041 cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg
     5101 gcttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata
     5161 accgtattac cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca
     5221 gcgagtcagt gagcgaggaa gcggaagagc gcccaatacg caaaccgcct ctccccgcgc
     5281 gttggccgat tcattaatgc ag
//
