LOCUS       sequence-250343-        4765 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses td-FAST in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4765)
  AUTHORS   Tebo AG, Pimenta FM, Zhang Y, Gautier A
  TITLE     Improved Chemical-Genetic Fluorescent Markers for Live Cell 
            Microscopy.
  JOURNAL   Biochemistry. 2018 Oct 2;57(39):5648-5653. doi: 
            10.1021/acs.biochem.8b00649. Epub 2018 Sep 17.
  PUBMED    30204425
REFERENCE   2  (bases 1 to 4765)
  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"; doi: 
            "10.1021/acs.biochem.8b00649"; journalName: "Biochemistry"; date: 
            "2018-10-2- 2"; volume: "57"; issue: "39"; pages: "5648-5653"
COMMENT     Sequence Label: pAG330 td-FAST
FEATURES             Location/Qualifiers
     source          1..4765
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      615..624
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             621..995
                     /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="MEHVAFGSEDIENTLAKMDDGQLDGLAFGAIQLDGDGNILQYNAA
                     EGDITGRDPKQVIGKNFFKDVAPGTDSPEFYGKFKEGVASGNLNTMFEWMIPTSRGPTK
                     VKVHMKKALSGDSYWVFVKRV"
     CDS             1017..1388
                     /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="EHVAFGSEDIENTLAKMDDGQLDGLAFGAIQLDGDGNILQYNAAE
                     GDITGRDPKQVIGKNFFKDVAPGTDSPEFYGKFKEGVASGNLNTMFEWMIPTSRGPTKV
                     KVHMKKALSGDSYWVFVKRV"
     CDS             1395..1424
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     polyA_signal    1553..1674
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1590..1609)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1644..1663
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1681..2136)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1818..1839)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2030..2049
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2163..2267
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2269..2626
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2269..2460
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2292..2312)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2477..2612
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2661..3455
                     /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             2661..3455
                     /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(2715..2734)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3325..3344
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3643..3662)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3687..3734
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4063..4651
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4552..4571
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggactcag atctgccacc atggaacatg tcgccttcgg cagcgaggat atcgagaaca
      661 ctctggctaa gatggatgac gggcaactgg acggcttggc attcggggcc attcaactgg
      721 acggggacgg taatattctc cagtacaatg cagctgaagg tgacataact ggacgtgatc
      781 ctaaacaagt gattggcaag aatttcttca aagatgttgc tcccggcacc gattcacctg
      841 agttttacgg gaagttcaaa gagggcgttg cctccggcaa cctcaacact atgttcgaat
      901 ggatgatacc gacaagcaga ggcccaacca aggtaaaagt tcatatgaag aaagctctgt
      961 ctggagactc ttattgggtg tttgtgaaga gagtcggcgg cggctccgga ggaggagaac
     1021 acgttgcctt tggctctgag gatatcgaaa atacgctggc caaaatggat gacggtcagc
     1081 tggatggcct ggcattcgga gccatacagc ttgacggcga tggaaacatc ctccagtaca
     1141 acgccgcgga gggtgacatt accggtcgcg atccaaaaca ggttattggc aagaacttct
     1201 ttaaggacgt tgcacccggg actgactctc ccgaatttta cggaaagttc aaggaagggg
     1261 tggcatctgg caatcttaat acaatgtttg aatggatgat tcctactagt agaggaccaa
     1321 caaaggtgaa ggtgcacatg aagaaagcct tgtccggcga ctcctactgg gtctttgtga
     1381 aacgggtggg atccgaacaa aagcttattt ctgaagagga cttgtaaata ggcggccgcg
     1441 actctagatc ataatcagcc ataccacatt tgtagaggtt ttacttgctt taaaaaacct
     1501 cccacacctc cccctgaacc tgaaacataa aatgaatgca attgttgttg ttaacttgtt
     1561 tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc
     1621 atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttaaggcgt
     1681 aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat
     1741 tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga
     1801 tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca
     1861 acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccct
     1921 aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc
     1981 cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag
     2041 cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca
     2101 cacccgccgc gcttaatgcg ccgctacagg gcgcgtcagg tggcactttt cggggaaatg
     2161 tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga
     2221 gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtcct gaggcggaaa
     2281 gaaccagctg tggaatgtgt gtcagttagg gtgtggaaag tccccaggct ccccagcagg
     2341 cagaagtatg caaagcatgc atctcaatta gtcagcaacc aggtgtggaa agtccccagg
     2401 ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc
     2461 gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca
     2521 tggctgacta atttttttta tttatgcaga ggccgaggcc gcctcggcct ctgagctatt
     2581 ccagaagtag tgaggaggct tttttggagg cctaggcttt tgcaaagatc gatcaagaga
     2641 caggatgagg atcgtttcgc atgattgaac aagatggatt gcacgcaggt tctccggccg
     2701 cttgggtgga gaggctattc ggctatgact gggcacaaca gacaatcggc tgctctgatg
     2761 ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct ttttgtcaag accgacctgt
     2821 ccggtgccct gaatgaactg caagacgagg cagcgcggct atcgtggctg gccacgacgg
     2881 gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc gggaagggac tggctgctat
     2941 tgggcgaagt gccggggcag gatctcctgt catctcacct tgctcctgcc gagaaagtat
     3001 ccatcatggc tgatgcaatg cggcggctgc atacgcttga tccggctacc tgcccattcg
     3061 accaccaagc gaaacatcgc atcgagcgag cacgtactcg gatggaagcc ggtcttgtcg
     3121 atcaggatga tctggacgaa gagcatcagg ggctcgcgcc agccgaactg ttcgccaggc
     3181 tcaaggcgag catgcccgac ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc
     3241 cgaatatcat ggtggaaaat ggccgctttt ctggattcat cgactgtggc cggctgggtg
     3301 tggcggaccg ctatcaggac atagcgttgg ctacccgtga tattgctgaa gagcttggcg
     3361 gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca
     3421 tcgccttcta tcgccttctt gacgagttct tctgagcggg actctggggt tcgaaatgac
     3481 cgaccaagcg acgcccaacc tgccatcacg agatttcgat tccaccgccg ccttctatga
     3541 aaggttgggc ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga
     3601 tctcatgctg gagttcttcg cccaccctag ggggaggcta actgaaacac ggaaggagac
     3661 aataccggaa ggaacccgcg ctatgacggc aataaaaaga cagaataaaa cgcacggtgt
     3721 tgggtcgttt gttcataaac gcggggttcg gtcccagggc tggcactctg tcgatacccc
     3781 accgagaccc cattggggcc aatacgcccg cgtttcttcc ttttccccac cccacccccc
     3841 aagttcgggt gaaggcccag ggctcgcagc caacgtcggg gcggcaggcc ctgccatagc
     3901 ctcaggttac tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat
     3961 ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt
     4021 ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct
     4081 gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc
     4141 ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc
     4201 aaatactgtc cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc
     4261 gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc
     4321 gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg
     4381 aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata
     4441 cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta
     4501 tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc
     4561 ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg
     4621 atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt
     4681 cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt
     4741 ggataaccgt attaccgcca tgcat
//
