LOCUS       sequence-315225-        4647 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses FRB-N-frFAST in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4647)
  AUTHORS   Li C, Tebo AG, Thauvin M, Plamont MA, Volovitch M, Morin X, Vriz S, 
            Gautier A
  TITLE     A Far-Red Emitting Fluorescent Chemogenetic Reporter for In Vivo 
            Molecular Imaging.
  JOURNAL   Angew Chem Int Ed Engl. 2020 Jun 22. doi: 10.1002/anie.202006576.
  PUBMED    32568417
REFERENCE   2  (bases 1 to 4647)
  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: "Angew Chem 
            Int Ed Engl. 2020 Jun 22. doi: 10.1002/anie.202006576."
COMMENT     Sequence Label: pAG499-FRB-N-frFAST
FEATURES             Location/Qualifiers
     source          1..4647
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..304
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        305..508
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     459..479
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      555..564
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             564..593
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             600..869
                     /codon_start=1
                     /product="FKBP-rapamycin binding domain of human FRAP"
                     /label=FRB
                     /translation="EMWHEGLEEASRLYFGERNVKGMFEVLEPLHAMMERGPQTLKETS
                     FNQAYGRDLMEAQEWCRKYMKSGNVKDLTQAWDLYYHVFRRISKQ"
     polyA_signal    1375..1496
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1412..1431)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1466..1485
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1503..1958)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1640..1661)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1852..1871
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        1985..2089
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2091..2448
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2091..2282
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2114..2134)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2299..2434
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2483..3277
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             2483..3277
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2537..2556)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3147..3166
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3465..3484)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3509..3556
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      3885..4473
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4374..4393
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
       61 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      121 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      181 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      241 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      301 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      361 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      421 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      481 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      541 ccggactcag atctgccacc atggaacaaa agcttatttc tgaagaggac ttggaattcg
      601 agatgtggca tgaaggcctg gaagaggcat ctcgtttgta ctttggggaa aggaacgtga
      661 aaggcatgtt tgaggtgctg gagcccttgc atgctatgat ggaacggggc ccccagactc
      721 tgaaggaaac atcctttaat caggcctatg gtcgagattt aatggaggcc caagagtggt
      781 gcaggaagta catgaaatca gggaatgtca aggacctcac ccaagcctgg gacctctatt
      841 atcatgtgtt ccgacgaatc tcaaagcagg tctccggagg aggcggcagc ggcggagggg
      901 gatccatgga gcatgttgcc tttggcagtg aggacatcga gaacactttg gccaaaatgg
      961 acgacggaca actggatggg ttggcctttg gcgcaattca gctcgatggt gacgggaata
     1021 tcctgcagta caatgctgct gaaggagaca tcacaggcag agatcccaaa caggtgattg
     1081 ggaagaactt attcaaggat gttgcacctg gaacggtttc ctccgggttt tacggcaaat
     1141 tcaaggaagg cgtagcgtca gggaatctga acaccatgtt cgaatggatg ataccgacaa
     1201 gcaggggacc aaccaaggtc aagatacaca tgaagaaagc cctttcctaa taggcggccg
     1261 cgactctaga tcataatcag ccataccaca tttgtagagg ttttacttgc tttaaaaaac
     1321 ctcccacacc tccccctgaa cctgaaacat aaaatgaatg caattgttgt tgttaacttg
     1381 tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa
     1441 gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttaaggc
     1501 gtaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
     1561 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
     1621 gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc
     1681 caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc
     1741 ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag
     1801 cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa
     1861 agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac
     1921 cacacccgcc gcgcttaatg cgccgctaca gggcgcgtca ggtggcactt ttcggggaaa
     1981 tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat
     2041 gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagtc ctgaggcgga
     2101 aagaaccagc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca
     2161 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca
     2221 ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc
     2281 ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc
     2341 catggctgac taattttttt tatttatgca gaggccgagg ccgcctcggc ctctgagcta
     2401 ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaga tcgatcaaga
     2461 gacaggatga ggatcgtttc gcatgattga acaagatgga ttgcacgcag gttctccggc
     2521 cgcttgggtg gagaggctat tcggctatga ctgggcacaa cagacaatcg gctgctctga
     2581 tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca agaccgacct
     2641 gtccggtgcc ctgaatgaac tgcaagacga ggcagcgcgg ctatcgtggc tggccacgac
     2701 gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg actggctgct
     2761 attgggcgaa gtgccggggc aggatctcct gtcatctcac cttgctcctg ccgagaaagt
     2821 atccatcatg gctgatgcaa tgcggcggct gcatacgctt gatccggcta cctgcccatt
     2881 cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact cggatggaag ccggtcttgt
     2941 cgatcaggat gatctggacg aagagcatca ggggctcgcg ccagccgaac tgttcgccag
     3001 gctcaaggcg agcatgcccg acggcgagga tctcgtcgtg acccatggcg atgcctgctt
     3061 gccgaatatc atggtggaaa atggccgctt ttctggattc atcgactgtg gccggctggg
     3121 tgtggcggac cgctatcagg acatagcgtt ggctacccgt gatattgctg aagagcttgg
     3181 cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc gccgctcccg attcgcagcg
     3241 catcgccttc tatcgccttc ttgacgagtt cttctgagcg ggactctggg gttcgaaatg
     3301 accgaccaag cgacgcccaa cctgccatca cgagatttcg attccaccgc cgccttctat
     3361 gaaaggttgg gcttcggaat cgttttccgg gacgccggct ggatgatcct ccagcgcggg
     3421 gatctcatgc tggagttctt cgcccaccct agggggaggc taactgaaac acggaaggag
     3481 acaataccgg aaggaacccg cgctatgacg gcaataaaaa gacagaataa aacgcacggt
     3541 gttgggtcgt ttgttcataa acgcggggtt cggtcccagg gctggcactc tgtcgatacc
     3601 ccaccgagac cccattgggg ccaatacgcc cgcgtttctt ccttttcccc accccacccc
     3661 ccaagttcgg gtgaaggccc agggctcgca gccaacgtcg gggcggcagg ccctgccata
     3721 gcctcaggtt actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg
     3781 atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg
     3841 ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt
     3901 ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg
     3961 ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata
     4021 ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca
     4081 ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag
     4141 tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc
     4201 tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga
     4261 tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg
     4321 tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac
     4381 gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg
     4441 tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg
     4501 ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct
     4561 gtggataacc gtattaccgc catgcattag ttattaatag taatcaatta cggggtcatt
     4621 agttcatagc ccatatatgg agttccg
//
