LOCUS       sequence-315365-        4437 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses LifeAct-redFAST in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4437)
  AUTHORS   Tebo AG, Moeyaert B, Thauvin M, Carlon-Andres I, Boken D, Volovitch 
            M, Padilla-Parra S, Dedecker P, Vriz S, Gautier A
  TITLE     Orthogonal fluorescent chemogenetic reporters for multicolor 
            imaging.
  JOURNAL   Nat Chem Biol. 2020 Aug 10. pii: 10.1038/s41589-020-0611-0. doi: 
            10.1038/s41589-020-0611-0.
  PUBMED    32778846
REFERENCE   2  (bases 1 to 4437)
  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: "Nat Chem 
            Biol. 2020 Aug 10. pii: 10.1038/s41589-020-0611-0. doi: 
            10.1038/s41589-020-0611-0."
COMMENT     Sequence Label: pAG469-LifeAct-redFAST
FEATURES             Location/Qualifiers
     source          1..4437
                     /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             561..611
                     /codon_start=1
                     /product="peptide that binds filamentous actin (Riedl et 
                     al., 2008)"
                     /label=Lifeact
                     /note="first 17 amino acids of budding yeast Abp140"
                     /translation="MGVADLIKKFESISKEE"
     CDS             627..1001
                     /codon_start=1
                     /product="small protein tag, derived from photoactive 
                     yellow protein, that allows in vivo fluorescent labeling by
                     addition of a cell-permeant fluorogen (Plamont et al., 
                     2016)"
                     /label=Y-FAST
                     /translation="MEHVAFGSEDIENTLAKMDDGQLDGLALGAIQLDGDGNILQYNAA
                     QGDITGADPKQVIGKNFFKDVAPGTDSPEFYGKFKVGVASGNLNTMFEWMIPTNRGPTK
                     VKVHMKKALSGDSYWVFVKRV"
     CDS             1008..1037
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     polyA_signal    1165..1286
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1202..1221)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1256..1275
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1293..1748)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1430..1451)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1642..1661
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        1775..1879
                     /gene="bla"
                     /label=AmpR promoter
     promoter        1881..2238
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        1881..2072
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(1904..1924)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2089..2224
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2273..3067
                     /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             2273..3067
                     /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(2327..2346)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2937..2956
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3255..3274)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3299..3346
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      3675..4263
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4164..4183
                     /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 atgggcgtgg ccgacttgat caagaagttc gagtccatct
      601 ccaaggagga gggggatcca ccggtcatgg agcatgttgc ctttggcagt gaggacatcg
      661 agaacactct ggccaaaatg gacgacggac aactggatgg gttggcctta ggcgcaattc
      721 agctcgatgg tgacgggaat atcctgcagt acaatgctgc tcagggagac atcacaggcg
      781 cagatcccaa acaggtgatt gggaagaact tcttcaagga tgttgcacct ggaacggatt
      841 ctcccgagtt ttacggcaaa ttcaaggtag gcgtagcgtc agggaatctg aacaccatgt
      901 tcgaatggat gataccgaca aacaggggac caaccaaggt caaggtgcac atgaagaaag
      961 ccctttccgg tgacagctat tgggtctttg tgaaacgggt gggatccgaa caaaagctta
     1021 tttctgaaga ggacttgtaa taggcggccg cgactctaga tcataatcag ccataccaca
     1081 tttgtagagg ttttacttgc tttaaaaaac ctcccacacc tccccctgaa cctgaaacat
     1141 aaaatgaatg caattgttgt tgttaacttg tttattgcag cttataatgg ttacaaataa
     1201 agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt
     1261 ttgtccaaac tcatcaatgt atcttaaggc gtaaattgta agcgttaata ttttgttaaa
     1321 attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa
     1381 aatcccttat aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa
     1441 caagagtcca ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca
     1501 gggcgatggc ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg
     1561 taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc
     1621 ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc
     1681 aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca
     1741 gggcgcgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt
     1801 ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata
     1861 atattgaaaa aggaagagtc ctgaggcgga aagaaccagc tgtggaatgt gtgtcagtta
     1921 gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat
     1981 tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     2041 atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta
     2101 actccgccca gttccgccca ttctccgccc catggctgac taattttttt tatttatgca
     2161 gaggccgagg ccgcctcggc ctctgagcta ttccagaagt agtgaggagg cttttttgga
     2221 ggcctaggct tttgcaaaga tcgatcaaga gacaggatga ggatcgtttc gcatgattga
     2281 acaagatgga ttgcacgcag gttctccggc cgcttgggtg gagaggctat tcggctatga
     2341 ctgggcacaa cagacaatcg gctgctctga tgccgccgtg ttccggctgt cagcgcaggg
     2401 gcgcccggtt ctttttgtca agaccgacct gtccggtgcc ctgaatgaac tgcaagacga
     2461 ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct tgcgcagctg tgctcgacgt
     2521 tgtcactgaa gcgggaaggg actggctgct attgggcgaa gtgccggggc aggatctcct
     2581 gtcatctcac cttgctcctg ccgagaaagt atccatcatg gctgatgcaa tgcggcggct
     2641 gcatacgctt gatccggcta cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg
     2701 agcacgtact cggatggaag ccggtcttgt cgatcaggat gatctggacg aagagcatca
     2761 ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg agcatgcccg acggcgagga
     2821 tctcgtcgtg acccatggcg atgcctgctt gccgaatatc atggtggaaa atggccgctt
     2881 ttctggattc atcgactgtg gccggctggg tgtggcggac cgctatcagg acatagcgtt
     2941 ggctacccgt gatattgctg aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct
     3001 ttacggtatc gccgctcccg attcgcagcg catcgccttc tatcgccttc ttgacgagtt
     3061 cttctgagcg ggactctggg gttcgaaatg accgaccaag cgacgcccaa cctgccatca
     3121 cgagatttcg attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg
     3181 gacgccggct ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccacact
     3241 agggggaggc taactgaaac acggaaggag acaataccgg aaggaacccg cgctatgacg
     3301 gcaataaaaa gacagaataa aacgcacggt gttgggtcgt ttgttcataa acgcggggtt
     3361 cggtcccagg gctggcactc tgtcgatacc ccaccgagac cccattgggg ccaatacgcc
     3421 cgcgtttctt ccttttcccc accccacccc ccaagttcgg gtgaaggccc agggctcgca
     3481 gccaacgtcg gggcggcagg ccctgccata gcctcaggtt actcatatat actttagatt
     3541 gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc
     3601 atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag
     3661 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
     3721 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
     3781 aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag
     3841 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
     3901 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
     3961 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
     4021 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc
     4081 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga
     4141 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
     4201 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
     4261 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac
     4321 atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc catgcattag
     4381 ttattaatag taatcaatta cggggtcatt agttcatagc ccatatatgg agttccg
//
