LOCUS       sequence-254933-        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        96..399
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        400..603
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     554..574
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      650..659
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             656..1030
                     /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             1052..1423
                     /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             1430..1459
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     polyA_signal    1588..1709
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1625..1644)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1679..1698
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1716..2171)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1853..1874)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2065..2084
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2198..2302
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2304..2661
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2304..2495
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2327..2347)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2512..2647
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2696..3490
                     /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             2696..3490
                     /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(2750..2769)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3360..3379
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3678..3697)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3722..3769
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4098..4686
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4587..4606
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ctgattctgt ggataaccgt attaccgcca tgcattagtt attaatagta atcaattacg
       61 gggtcattag ttcatagccc atatatggag ttccgcgtta cataacttac ggtaaatggc
      121 ccgcctggct gaccgcccaa cgacccccgc ccattgacgt caataatgac gtatgttccc
      181 atagtaacgc caatagggac tttccattga cgtcaatggg tggagtattt acggtaaact
      241 gcccacttgg cagtacatca agtgtatcat atgccaagta cgccccctat tgacgtcaat
      301 gacggtaaat ggcccgcctg gcattatgcc cagtacatga ccttatggga ctttcctact
      361 tggcagtaca tctacgtatt agtcatcgct attaccatgg tgatgcggtt ttggcagtac
      421 atcaatgggc gtggatagcg gtttgactca cggggatttc caagtctcca ccccattgac
      481 gtcaatggga gtttgttttg gcaccaaaat caacgggact ttccaaaatg tcgtaacaac
      541 tccgccccat tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta tataagcaga
      601 gctggtttag tgaaccgtca gatccgctag cgctaccgga ctcagatctg ccaccatgga
      661 acatgtcgcc ttcggcagcg aggatatcga gaacactctg gctaagatgg atgacgggca
      721 actggacggc ttggcattcg gggccattca actggacggg gacggtaata ttctccagta
      781 caatgcagct gaaggtgaca taactggacg tgatcctaaa caagtgattg gcaagaattt
      841 cttcaaagat gttgctcccg gcaccgattc acctgagttt tacgggaagt tcaaagaggg
      901 cgttgcctcc ggcaacctca acactatgtt cgaatggatg ataccgacaa gcagaggccc
      961 aaccaaggta aaagttcata tgaagaaagc tctgtctgga gactcttatt gggtgtttgt
     1021 gaagagagtc ggcggcggct ccggaggagg agaacacgtt gcctttggct ctgaggatat
     1081 cgaaaatacg ctggccaaaa tggatgacgg tcagctggat ggcctggcat tcggagccat
     1141 acagcttgac ggcgatggaa acatcctcca gtacaacgcc gcggagggtg acattaccgg
     1201 tcgcgatcca aaacaggtta ttggcaagaa cttctttaag gacgttgcac ccgggactga
     1261 ctctcccgaa ttttacggaa agttcaagga aggggtggca tctggcaatc ttaatacaat
     1321 gtttgaatgg atgattccta ctagtagagg accaacaaag gtgaaggtgc acatgaagaa
     1381 agccttgtcc ggcgactcct actgggtctt tgtgaaacgg gtgggatccg aacaaaagct
     1441 tatttctgaa gaggacttgt aaataggcgg ccgcgactct agatcataat cagccatacc
     1501 acatttgtag aggttttact tgctttaaaa aacctcccac acctccccct gaacctgaaa
     1561 cataaaatga atgcaattgt tgttgttaac ttgtttattg cagcttataa tggttacaaa
     1621 taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt
     1681 ggtttgtcca aactcatcaa tgtatcttaa ggcgtaaatt gtaagcgtta atattttgtt
     1741 aaaattcgcg ttaaattttt gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg
     1801 caaaatccct tataaatcaa aagaatagac cgagataggg ttgagtgttg ttccagtttg
     1861 gaacaagagt ccactattaa agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta
     1921 tcagggcgat ggcccactac gtgaaccatc accctaatca agttttttgg ggtcgaggtg
     1981 ccgtaaagca ctaaatcgga accctaaagg gagcccccga tttagagctt gacggggaaa
     2041 gccggcgaac gtggcgagaa aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct
     2101 ggcaagtgta gcggtcacgc tgcgcgtaac caccacaccc gccgcgctta atgcgccgct
     2161 acagggcgcg tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt
     2221 tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca
     2281 ataatattga aaaaggaaga gtcctgaggc ggaaagaacc agctgtggaa tgtgtgtcag
     2341 ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc
     2401 aattagtcag caaccaggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa
     2461 agcatgcatc tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc
     2521 ctaactccgc ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat
     2581 gcagaggccg aggccgcctc ggcctctgag ctattccaga agtagtgagg aggctttttt
     2641 ggaggcctag gcttttgcaa agatcgatca agagacagga tgaggatcgt ttcgcatgat
     2701 tgaacaagat ggattgcacg caggttctcc ggccgcttgg gtggagaggc tattcggcta
     2761 tgactgggca caacagacaa tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca
     2821 ggggcgcccg gttctttttg tcaagaccga cctgtccggt gccctgaatg aactgcaaga
     2881 cgaggcagcg cggctatcgt ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga
     2941 cgttgtcact gaagcgggaa gggactggct gctattgggc gaagtgccgg ggcaggatct
     3001 cctgtcatct caccttgctc ctgccgagaa agtatccatc atggctgatg caatgcggcg
     3061 gctgcatacg cttgatccgg ctacctgccc attcgaccac caagcgaaac atcgcatcga
     3121 gcgagcacgt actcggatgg aagccggtct tgtcgatcag gatgatctgg acgaagagca
     3181 tcaggggctc gcgccagccg aactgttcgc caggctcaag gcgagcatgc ccgacggcga
     3241 ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat atcatggtgg aaaatggccg
     3301 cttttctgga ttcatcgact gtggccggct gggtgtggcg gaccgctatc aggacatagc
     3361 gttggctacc cgtgatattg ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt
     3421 gctttacggt atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga
     3481 gttcttctga gcgggactct ggggttcgaa atgaccgacc aagcgacgcc caacctgcca
     3541 tcacgagatt tcgattccac cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc
     3601 cgggacgccg gctggatgat cctccagcgc ggggatctca tgctggagtt cttcgcccac
     3661 cctaggggga ggctaactga aacacggaag gagacaatac cggaaggaac ccgcgctatg
     3721 acggcaataa aaagacagaa taaaacgcac ggtgttgggt cgtttgttca taaacgcggg
     3781 gttcggtccc agggctggca ctctgtcgat accccaccga gaccccattg gggccaatac
     3841 gcccgcgttt cttccttttc cccaccccac cccccaagtt cgggtgaagg cccagggctc
     3901 gcagccaacg tcggggcggc aggccctgcc atagcctcag gttactcata tatactttag
     3961 attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat
     4021 ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa
     4081 aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca
     4141 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt
     4201 ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg
     4261 tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc
     4321 ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga
     4381 cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc
     4441 agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc
     4501 gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca
     4561 ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg
     4621 tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta
     4681 tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct
     4741 cacatgttct ttcctgcgtt atccc
//
