LOCUS       sequence-229314-        7216 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bimolecular Fluorescence Complimentation (BiFC) assay. Gateway 
            compatible N-terminal vector with BiFC-tag (GFP11-Flag) to screen 
            interaction partners..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7216)
  AUTHORS   Viita T, Kyheroinen S, Prajapati B, Virtanen J, Frilander MJ, 
            Varjosalo M, Vartiainen MK
  TITLE     Nuclear actin interactome analysis links actin to KAT14 histone 
            acetyl transferase and mRNA splicing.
  JOURNAL   J Cell Sci. 2019 Apr 17;132(8). pii: jcs.226852. doi: 
            10.1242/jcs.226852.
  PUBMED    30890647
REFERENCE   2  (bases 1 to 7216)
  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: "J Cell 
            Sci."; date: "2019-04-17"; volume: "132(8). pii: jcs.226852. doi"; 
            pages: " 10.1242/jcs.226852"
COMMENT     Sequence Label: Flag-GFP11-pDEST-N
FEATURES             Location/Qualifiers
     source          1..7216
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        15..318
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        319..522
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     473..493
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      561..570
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             576..623
                     /codon_start=1
                     /product="11th beta-strand of superfolder GFP"
                     /label=GFP11
                     /note="associates with GFP(1-10), consisting of the first 
                     10 beta-strands, to reconstitute fluorescent GFP (Cabantous
                     et al., 2005; Kamiyama et al., 2016)"
                     /translation="RDHMVLHEYVNAAGIT"
     CDS             630..653
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     protein_bind    684..808
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(833..838,839..856,857..863)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             917..1576
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(983..1002)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             1918..2223
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     2102..2121
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(2264..2388)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(2420..2439)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2421..2439)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     polyA_signal    2596..2730
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2633..2652)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2687..2706
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      2856..3311
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2943..2962)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3153..3174
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3428..3785
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3428..3619
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3451..3471)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3636..3771
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3836..4630
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3890..3909)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4500..4519
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4694..4742
                     /note="synthetic polyadenylation signal"
     primer_bind     complement(4783..4802)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4902..4924
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     promoter        5048..5152
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5153..5221,5222..6013)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(5371..5390)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6184..6772
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6673..6692
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6926..6943
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        complement(7017..7121)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cattagatgc atgtcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac
       61 gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact
      121 ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa
      181 gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg
      241 cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta
      301 gtcatcgcta ttaccatggt gatgcggttt tggcagtaca tcaatgggcg tggatagcgg
      361 tttgactcac ggggatttcc aagtctccac cccattgacg tcaatgggag tttgttttgg
      421 caccaaaatc aacgggactt tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg
      481 ggcggtaggc gtgtacggtg ggaggtctat ataagcagag ctggtttagt gaaccgtcag
      541 atccgctagc gctaccggtc gccaccatgg gcagccggga ccacatggtg ctgcacgagt
      601 acgtgaacgc cgccggcatc acaggcagcg attacaagga tgacgacgat aagtccggac
      661 tcagatctcg agcttctaga tcaacaagtt tgtacaaaaa agctgaacga gaaacgtaaa
      721 atgatataaa tatcaatata ttaaattaga ttttgcataa aaaacagact acataatact
      781 gtaaaacaca acatatccag tcactatggc ggccgcatta ggcaccccag gctttacact
      841 ttatgcttcc ggctcgtata atgtgtggat tttgagttag gatccgtcga gattttcagg
      901 agctaaggaa gctaaaatgg agaaaaaaat cactggatat accaccgttg atatatccca
      961 atggcatcgt aaagaacatt ttgaggcatt tcagtcagtt gctcaatgta cctataacca
     1021 gaccgttcag ctggatatta cggccttttt aaagaccgta aagaaaaata agcacaagtt
     1081 ttatccggcc tttattcaca ttcttgcccg cctgatgaat gctcatccgg aattccgtat
     1141 ggcaatgaaa gacggtgagc tggtgatatg ggatagtgtt cacccttgtt acaccgtttt
     1201 ccatgagcaa actgaaacgt tttcatcgct ctggagtgaa taccacgacg atttccggca
     1261 gtttctacac atatattcgc aagatgtggc gtgttacggt gaaaacctgg cctatttccc
     1321 taaagggttt attgagaata tgtttttcgt ctcagccaat ccctgggtga gtttcaccag
     1381 ttttgattta aacgtggcca atatggacaa cttcttcgcc cccgttttca ccatgggcaa
     1441 atattatacg caaggcgaca aggtgctgat gccgctggcg attcaggttc atcatgccgt
     1501 ttgtgatggc ttccatgtcg gcagaatgct taatgaatta caacagtact gcgatgagtg
     1561 gcagggcggg gcgtaaagat ctggatccgg cttactaaaa gccagataac agtatgcgta
     1621 tttgcgcgct gatttttgcg gtataagaat atatactgat atgtataccc gaagtatgtc
     1681 aaaaagaggt atgctatgaa gcagcgtatt acagtgacag ttgacagcga cagctatcag
     1741 ttgctcaagg catatatgat gtcaatatct ccggtctggt aagcacaacc atgcagaatg
     1801 aagcccgtcg tctgcgtgcc gaacgctgga aagcggaaaa tcaggaaggg atggctgagg
     1861 tcgcccggtt tattgaaatg aacggctctt ttgctgacga gaacaggggc tggtgaaatg
     1921 cagtttaagg tttacaccta taaaagagag agccgttatc gtctgtttgt ggatgtacag
     1981 agtgatatta ttgacacgcc cgggcgacgg atggtgatcc ccctggccag tgcacgtctg
     2041 ctgtcagata aagtctcccg tgaactttac ccggtggtgc atatcgggga tgaaagctgg
     2101 cgcatgatga ccaccgatat ggccagtgtg ccggtctccg ttatcgggga agaagtggct
     2161 gatctcagcc accgcgaaaa tgacatcaaa aacgccatta acctgatgtt ctggggaata
     2221 taaatgtcag gctcccttat acacagccag tctgcaggtc gaccatagtg actggatatg
     2281 ttgtgtttta cagtattatg tagtctgttt tttatgcaaa atctaattta atatattgat
     2341 atttatatca ttttacgttt ctcgttcagc tttcttgtac aaagtggttg atcgcgtgca
     2401 tgcgacgtca tagctctctc cctatagtga gtcgtattat aagctaggca ctggccgtcg
     2461 ttgatgggat ccaccggatc tagataactg atcataatca gccataccac atttgtagag
     2521 gttttacttg ctttaaaaaa cctcccacac ctccccctga acctgaaaca taaaatgaat
     2581 gcaattgttg ttgttaactt gtttattgca gcttataatg gttacaaata aagcaatagc
     2641 atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa
     2701 ctcatcaatg tatcttatca tgtctggatc gatcctgcat taatgaatcg gccaacgcgc
     2761 ggggagaggc ggtttgcgta ttggctggcg taatagcgaa gaggcccgca ccgatcgccc
     2821 ttcccaacag ttgcgcagcc tgaatggcga atgggacgcg ccctgtagcg gcgcattaag
     2881 cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc
     2941 cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc
     3001 tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa
     3061 aaaacttgat tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg
     3121 ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac
     3181 actcaaccct atctcggtct attcttttga tttataaggg attttgccga tttcggccta
     3241 ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattttaaca aaatattaac
     3301 gcttacaatt tcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc
     3361 atacgcggat ctgcgcagca ccatggcctg aaataacctc tgaaagagga acttggttag
     3421 gtaccttctg aggcggaaag aaccagctgt ggaatgtgtg tcagttaggg tgtggaaagt
     3481 ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca
     3541 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt
     3601 agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt
     3661 ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg
     3721 cctcggcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt
     3781 gcaaaaagct tgattcttct gacacaacag tctcgaactt aaggctagag ccaccatgat
     3841 tgaacaagat ggattgcacg caggttctcc ggccgcttgg gtggagaggc tattcggcta
     3901 tgactgggca caacagacaa tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca
     3961 ggggcgcccg gttctttttg tcaagaccga cctgtccggt gccctgaatg aactgcagga
     4021 cgaggcagcg cggctatcgt ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga
     4081 cgttgtcact gaagcgggaa gggactggct gctattgggc gaagtgccgg ggcaggatct
     4141 cctgtcatct caccttgctc ctgccgagaa agtatccatc atggctgatg caatgcggcg
     4201 gctgcatacg cttgatccgg ctacctgccc attcgaccac caagcgaaac atcgcatcga
     4261 gcgagcacgt actcggatgg aagccggtct tgtcgatcag gatgatctgg acgaagagca
     4321 tcaggggctc gcgccagccg aactgttcgc caggctcaag gcgcgcatgc ccgacggcga
     4381 ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat atcatggtgg aaaatggccg
     4441 cttttctgga ttcatcgact gtggccggct gggtgtggcg gaccgctatc aggacatagc
     4501 gttggctacc cgtgatattg ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt
     4561 gctttacggt atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga
     4621 gttcttctga gcgggactct ggggttcgaa atgaccgacc aagcgacgcc caacctgcca
     4681 tcacgatggc cgcaataaaa tatctttatt ttcattacat ctgtgtgttg gttttttgtg
     4741 tgaatcgata gcgataagga tccgcgtatg gtgcactctc agtacaatct gctctgatgc
     4801 cgcatagtta agccagcccc gacacccgcc aacacccgct gacgcgccct gacgggcttg
     4861 tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct gcatgtgtca
     4921 gaggttttca ccgtcatcac cgaaacgcgc gagacgaaag ggcctcgtga tacgcctatt
     4981 tttataggtt aatgtcatga taataatggt ttcttagacg tcaggtggca cttttcgggg
     5041 aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct
     5101 catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat
     5161 tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc
     5221 tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg
     5281 ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg
     5341 ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga
     5401 cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta
     5461 ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc
     5521 tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc
     5581 gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg
     5641 ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc
     5701 aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca
     5761 acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct
     5821 tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat
     5881 cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg
     5941 gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat
     6001 taagcattgg taactgtcag accaagttta ctcatatata ctttagattg atttaaaact
     6061 tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat
     6121 cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc
     6181 ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct
     6241 accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg
     6301 cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca
     6361 cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc
     6421 tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga
     6481 taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac
     6541 gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga
     6601 agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag
     6661 ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg
     6721 acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag
     6781 caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc
     6841 tgcgttatcc cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc
     6901 tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg aagagcgccc
     6961 aatacgcaaa ccgcctctcc ccgcgcgttg gccgattcat taatgcagag cttgcaattc
     7021 gcgctttttc aatattattg aagcatttat cagggttatt gtctcatgag cggatacata
     7081 tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg
     7141 ccacctgacg tctaagaaac cattattatc atgacattaa cctataaaaa taggcgtatt
     7201 acgaggccct ttcacg
//
