LOCUS       sequence-254956-        4785 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses H2B-iFAST in mammalian cells (nuclear labeling).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4785)
  TITLE     iFAST-containing plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 4785)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pAG472 H2B-iFAST
FEATURES             Location/Qualifiers
     source          1..4785
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        50..353
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        354..557
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     508..528
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      1018..1027
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1024..1398
                     /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
                     VKIHMKKALSGDSYWVFVKRV"
     CDS             1405..1434
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     polyA_signal    1562..1683
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1599..1618)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1653..1672
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1690..2145)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1827..1848)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2039..2058
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2172..2276
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2278..2635
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2278..2469
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2301..2321)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2486..2621
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2670..3464
                     /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             2670..3464
                     /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(2724..2743)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3334..3353
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3652..3671)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3696..3743
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4072..4660
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4561..4580
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 agtaatcaat tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac
       61 ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa
      121 tgacgtatgt tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt
      181 atttacggta aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc
      241 ctattgacgt caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttat
      301 gggactttcc tacttggcag tacatctacg tattagtcat cgctattacc atggtgatgc
      361 ggttttggca gtacatcaat gggcgtggat agcggtttga ctcacgggga tttccaagtc
      421 tccaccccat tgacgtcaat gggagtttgt tttggcacca aaatcaacgg gactttccaa
      481 aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg taggcgtgta cggtgggagg
      541 tctatataag cagagctggt ttagtgaacc gtcagatccg ctagcgctac cggactcaga
      601 tctgaattcg ccaccatgcc cgaacctgcg aagtcagcgc ccgctcccaa aaaaggctct
      661 aaaaaagctg tcgccaagac ccagaagaag ggggataaga aaaggcgtaa gaccaggaaa
      721 gagagttacg ccatttacgt gtacaaagta ctaaaacaag tccacccgga cactggcatc
      781 tcctcaaagg cgatgggcat tatgaactca tttgtaaacg acatcttcga gcgcatcgcc
      841 ggagaagcgt cgcgcctggc gcattacaac aagcgctcca ctatcacatc ccgggagatc
      901 cagacggccg tgcgcctgct cctgcccgga gaactggcca aacacgctgt gtctgagggc
      961 acaaaggccg tgaccaagta caccagctcc aagggcggag gctccggagg cggatctgcc
     1021 accatggagc atgttgcctt tggcagtgag gacatcgaga acactctggc caaaatggac
     1081 gacggacaac tggatgggtt ggcctttggc gcaattcagc tcgatggtga cgggaatatc
     1141 ctgcagtaca atgctgctga aggagacatc acaggcagag atcccaaaca ggtgattggg
     1201 aagaacttct tcaaggatgt tgcacctgga acggattctc ccgagtttta cggcaaattc
     1261 aaggaaggcg tagcgtcagg gaatctgaac accatgttcg aatggatgat accgacaagc
     1321 aggggaccaa ccaaggtcaa gatacacatg aagaaagccc tttccggtga cagctattgg
     1381 gtctttgtga aacgggtggg atccgaacaa aagcttattt ctgaagagga cttgtaatag
     1441 gcggccgcga ctctagatca taatcagcca taccacattt gtagaggttt tacttgcttt
     1501 aaaaaacctc ccacacctcc ccctgaacct gaaacataaa atgaatgcaa ttgttgttgt
     1561 taacttgttt attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac
     1621 aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc
     1681 ttaaggcgta aattgtaagc gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa
     1741 tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat cccttataaa tcaaaagaat
     1801 agaccgagat agggttgagt gttgttccag tttggaacaa gagtccacta ttaaagaacg
     1861 tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac
     1921 catcacccta atcaagtttt ttggggtcga ggtgccgtaa agcactaaat cggaacccta
     1981 aagggagccc ccgatttaga gcttgacggg gaaagccggc gaacgtggcg agaaaggaag
     2041 ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg
     2101 taaccaccac acccgccgcg cttaatgcgc cgctacaggg cgcgtcaggt ggcacttttc
     2161 ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc
     2221 cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtcctg
     2281 aggcggaaag aaccagctgt ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc
     2341 cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca ggtgtggaaa
     2401 gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac
     2461 catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc
     2521 tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg cctcggcctc
     2581 tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt gcaaagatcg
     2641 atcaagagac aggatgagga tcgtttcgca tgattgaaca agatggattg cacgcaggtt
     2701 ctccggccgc ttgggtggag aggctattcg gctatgactg ggcacaacag acaatcggct
     2761 gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg cccggttctt tttgtcaaga
     2821 ccgacctgtc cggtgccctg aatgaactgc aagacgaggc agcgcggcta tcgtggctgg
     2881 ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg ggaagggact
     2941 ggctgctatt gggcgaagtg ccggggcagg atctcctgtc atctcacctt gctcctgccg
     3001 agaaagtatc catcatggct gatgcaatgc ggcggctgca tacgcttgat ccggctacct
     3061 gcccattcga ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg atggaagccg
     3121 gtcttgtcga tcaggatgat ctggacgaag agcatcaggg gctcgcgcca gccgaactgt
     3181 tcgccaggct caaggcgagc atgcccgacg gcgaggatct cgtcgtgacc catggcgatg
     3241 cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc tggattcatc gactgtggcc
     3301 ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc tacccgtgat attgctgaag
     3361 agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt
     3421 cgcagcgcat cgccttctat cgccttcttg acgagttctt ctgagcggga ctctggggtt
     3481 cgaaatgacc gaccaagcga cgcccaacct gccatcacga gatttcgatt ccaccgccgc
     3541 cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca
     3601 gcgcggggat ctcatgctgg agttcttcgc ccaccctagg gggaggctaa ctgaaacacg
     3661 gaaggagaca ataccggaag gaacccgcgc tatgacggca ataaaaagac agaataaaac
     3721 gcacggtgtt gggtcgtttg ttcataaacg cggggttcgg tcccagggct ggcactctgt
     3781 cgatacccca ccgagacccc attggggcca atacgcccgc gtttcttcct tttccccacc
     3841 ccacccccca agttcgggtg aaggcccagg gctcgcagcc aacgtcgggg cggcaggccc
     3901 tgccatagcc tcaggttact catatatact ttagattgat ttaaaacttc atttttaatt
     3961 taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga
     4021 gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc
     4081 tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt
     4141 ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc
     4201 gcagatacca aatactgttc ttctagtgta gccgtagtta ggccaccact tcaagaactc
     4261 tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg
     4321 cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg
     4381 gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga
     4441 actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc
     4501 ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg
     4561 gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg
     4621 atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt
     4681 tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc
     4741 tgattctgtg gataaccgta ttaccgccat gcattagtta ttaat
//
