LOCUS       sequence-414383-        5433 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Robust FRET biosensor enabling sensitive, accurate l-arginine 
            measurement at physiological temp.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5433)
  AUTHORS   Whitfield JH, Zhang W, Herde MK, Clifton BE, Radziejewski J, 
            Janovjak H, Henneberger C, Jackson CJ
  TITLE     Construction of a robust and sensitive arginine biosensor through 
            ancestral protein reconstruction.
  JOURNAL   Protein Sci. 2015 Jun 9. doi: 10.1002/pro.2721.
  PUBMED    26061224
REFERENCE   2  (bases 1 to 5433)
  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. 2015 Jun 9. doi: 10.1002/pro.2721."
COMMENT     Sequence Label: arginine biosensor cpFLIPR
FEATURES             Location/Qualifiers
     source          1..5433
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      208..796
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     697..716
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     950..967
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     1052..1071
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1052..1070
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             1102..1124
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1144..1161
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1165..1197
                     /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(1201..1209,1210..1224)
                     /codon_start=1
                     /product="Xpress(TM) epitope tag, including an enterokinase
                     recognition and cleavage site"
                     /label=Xpress(TM) tag
                     /translation="DLYDDDDK"
     CDS             join(1639..1641,1642..1644,1645..2355)
                     /codon_start=1
                     /product="enhanced CFP"
                     /label=ECFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(3091..3093,3094..3096,3097..3810)
                     /codon_start=1
                     /product="yellow fluorescent protein (YFP) with fast and 
                     efficient maturation (Nagai et al., 2002)"
                     /label=Venus
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KLICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK
                     ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3866..3884)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      3880..3927
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      4024..4479
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4111..4130)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4321..4342
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4505..4609
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4610..4678,4679..5433,1..37)
                     /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(4828..4847)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata
       61 tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct
      121 ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga
      181 ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg
      241 cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc
      301 aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct
      361 agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc
      421 tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt
      481 ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg
      541 cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct
      601 atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag
      661 ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag
      721 tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg
      781 gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg
      841 gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac
      901 cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt
      961 gagcgaggaa gcggatgagc gcccaatacg caaaccgcct ctccccgcgc gttggccgat
     1021 tcattaatgc aggatctcga tcccgcgaaa ttaatacgac tcactatagg gagaccacaa
     1081 cggtttccct ctagaaataa ttttgtttaa ctttaagaag gagatataca tatgcggggt
     1141 tctcatcatc atcatcatca tggtatggct agcatgactg gtggacagca aatgggtcgg
     1201 gatctgtacg acgatgacga taaggatccg aaactgaagg taacagtcaa cggcactgcg
     1261 tatgacgttg acgttgacgt cgacaagtca cacgaaaacc cgatgggcac catcctgttc
     1321 ggcggcggca ccggcggcgc gccggcaccg gcagcaggtg gcgcaggcgc cggtaaggcc
     1381 ggagagggcg agattcccgc tccgctggcc ggcaccgtct ccaagatcct cgtgaaggag
     1441 ggtgacacgg tcaaggctgg tcagaccgtg ctcgttctcg aggccatgaa gatggagacc
     1501 gagatcaacg ctcccaccga cggcaaggtc gagaaggtcc tggtcaagga gcgtgacgcg
     1561 gtgcagggcg gtcagggtct catcaagatc ggggatctcg agctcatcga aggctcgagc
     1621 ggttcggatc cgggccgcat ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc
     1681 atcctggtcg agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc
     1741 gagggcgatg ccacctacgg caagctgacc ctgaagttca tctgcaccac cggcaagctg
     1801 cccgtgccct ggcccaccct cgtgaccacc ctgacctggg gcgtgcagtg cttcagccgc
     1861 taccccgacc acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc
     1921 caggagcgca ccatcttctt caaggacgac ggcaactaca agacccgcgc cgaggtgaag
     1981 ttcgagggcg acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac
     2041 ggcaacatcc tggggcacaa gctggagtac aactacatca gccacaacgt ctatatcacc
     2101 gccgacaagc agaagaacgg catcaaggcc aacttcaaga tccgccacaa catcgaggac
     2161 ggcagcgtgc agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg
     2221 ctgctgcccg acaaccacta cctgagcacc cagtccgccc tgagcaaaga ccccaacgag
     2281 aagcgcgatc acatggtcct gctggagttc gtgaccgccg ccgggatcac tctcggcatg
     2341 gacgagctgt acaagggtgg taccggaatc gaacagtatg gtattgcact gaaaaaaggc
     2401 aatgatgagc tgctggaaaa aatcaacaaa gccctgaaaa aactgaaagc caatggcacc
     2461 tatgacaaaa tctatgagaa atggctgttt ggcaaagaag tttcaggcgg tagcggtggt
     2521 agtggtggtt ctggtggcag cggtgataaa aaactgcgtg ttggcaccga tgcaacctat
     2581 ccgccttttg aatttaaaga tgaaaatggt gaactgaccg gctttgatat tgatctggca
     2641 aaagcaattg ccaaagaaat gggtgtgaaa gtggaattcg tgaacatgga ttttgatggt
     2701 ctgattccgg cactgcagag cggtaaaatt gatattgcaa ttagcgccat taccatcacc
     2761 gatgaacgta aaaaaaccgt ggattttagc gatccgtatt atgatgcagg tctggccatt
     2821 gttgtgaaaa aagataacaa cggcatcaaa agcgtcgaag atctgaaagg taaacgtgtt
     2881 gcagttcaga ccggtacaac cagcgaagaa tatgtgaaag aaaactttcc gggtgccggt
     2941 gttaaaagct ttgaaaatat gagcgaagcc tttatggaac tgaaaaatgg tcgtgttgat
     3001 gcggtgatta ccgataatcc ggttgcagca gattatgtta aaaccagtcc ggatggtaaa
     3061 ctgaaagttg ttctcgccgg taccggtgga atggtgagca agggcgagga gctgttcacc
     3121 ggggtggtgc ccatcctggt cgagctggac ggcgacgtaa acggccacaa gttcagcgtg
     3181 tccggcgagg gcgagggcga tgccacctac ggcaagctga ccctgaagct gatctgcacc
     3241 accggcaagc tgcccgtgcc ctggcccacc ctcgtgacca ccctgggcta cggcctgcag
     3301 tgcttcgccc gctaccccga ccacatgaag cagcacgact tcttcaagtc cgccatgccc
     3361 gaaggctacg tccaggagcg caccatcttc ttcaaggacg acggcaacta caagacccgc
     3421 gccgaggtga agttcgaggg cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac
     3481 ttcaaggagg acggcaacat cctggggcac aagctggagt acaactacaa cagccacaac
     3541 gtctatatca ccgccgacaa gcagaagaac ggcatcaagg ccaacttcaa gatccgccac
     3601 aacatcgagg acggcggcgt gcagctcgcc gaccactacc agcagaacac ccccatcggc
     3661 gacggccccg tgctgctgcc cgacaaccac tacctgagct accagtccgc cctgagcaaa
     3721 gaccccaacg agaagcgcga tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc
     3781 actctcggca tggacgagct gtacaagtaa aagcttgatc cggctgctaa caaagcccga
     3841 aaggaagctg agttggctgc tgccaccgct gagcaataac tagcataacc ccttggggcc
     3901 tctaaacggg tcttgagggg ttttttgctg aaaggaggaa ctatatccgg atctggcgta
     3961 atagcgaaga ggcccgcacc gatcgccctt cccaacagtt gcgcagcctg aatggcgaat
     4021 gggacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga
     4081 ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg
     4141 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat
     4201 ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgttggt tcacgtagtg
     4261 ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata
     4321 gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt
     4381 tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat
     4441 ttaacgcgaa ttttaacaaa atattaacgc ttacaattta ggtggcactt ttcggggaaa
     4501 tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat
     4561 gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca
     4621 acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca
     4681 cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta
     4741 catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt
     4801 tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc
     4861 cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc
     4921 accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc
     4981 cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa
     5041 ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga
     5101 accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat
     5161 ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca
     5221 attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc
     5281 ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat
     5341 tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag
     5401 tcaggcaact atggatgaac gaaatagaca gat
//
