LOCUS       sequence-229457-        7177 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Gateway compatible mammalian expression vector with N-terminal 2Flag
            to tag proteins of interest..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7177)
  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 7177)
  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: 2Flag-pDEST-N
FEATURES             Location/Qualifiers
     source          1..7177
                     /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             570..593
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             600..623
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     protein_bind    645..769
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(794..799,800..817,818..824)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             878..1537
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(944..963)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             1879..2184
                     /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     2063..2082
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(2225..2349)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(2381..2400)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2382..2400)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     polyA_signal    2557..2691
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2594..2613)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2648..2667
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      2817..3272
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2904..2923)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3114..3135
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3389..3746
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3389..3580
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3412..3432)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3597..3732
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3797..4591
                     /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(3851..3870)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4461..4480
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4655..4703
                     /note="synthetic polyadenylation signal"
     primer_bind     complement(4744..4763)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4863..4885
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     promoter        5009..5113
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5114..5182,5183..5974)
                     /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(5332..5351)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6145..6733
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6634..6653
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6887..6904
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        complement(6978..7082)
                     /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 attacaagga tgacgacgat aagggcgcag
      601 attacaagga tgacgacgat aagagatctc gagcttctag atcaacaagt ttgtacaaaa
      661 aagctgaacg agaaacgtaa aatgatataa atatcaatat attaaattag attttgcata
      721 aaaaacagac tacataatac tgtaaaacac aacatatcca gtcactatgg cggccgcatt
      781 aggcacccca ggctttacac tttatgcttc cggctcgtat aatgtgtgga ttttgagtta
      841 ggatccgtcg agattttcag gagctaagga agctaaaatg gagaaaaaaa tcactggata
      901 taccaccgtt gatatatccc aatggcatcg taaagaacat tttgaggcat ttcagtcagt
      961 tgctcaatgt acctataacc agaccgttca gctggatatt acggcctttt taaagaccgt
     1021 aaagaaaaat aagcacaagt tttatccggc ctttattcac attcttgccc gcctgatgaa
     1081 tgctcatccg gaattccgta tggcaatgaa agacggtgag ctggtgatat gggatagtgt
     1141 tcacccttgt tacaccgttt tccatgagca aactgaaacg ttttcatcgc tctggagtga
     1201 ataccacgac gatttccggc agtttctaca catatattcg caagatgtgg cgtgttacgg
     1261 tgaaaacctg gcctatttcc ctaaagggtt tattgagaat atgtttttcg tctcagccaa
     1321 tccctgggtg agtttcacca gttttgattt aaacgtggcc aatatggaca acttcttcgc
     1381 ccccgttttc accatgggca aatattatac gcaaggcgac aaggtgctga tgccgctggc
     1441 gattcaggtt catcatgccg tttgtgatgg cttccatgtc ggcagaatgc ttaatgaatt
     1501 acaacagtac tgcgatgagt ggcagggcgg ggcgtaaaga tctggatccg gcttactaaa
     1561 agccagataa cagtatgcgt atttgcgcgc tgatttttgc ggtataagaa tatatactga
     1621 tatgtatacc cgaagtatgt caaaaagagg tatgctatga agcagcgtat tacagtgaca
     1681 gttgacagcg acagctatca gttgctcaag gcatatatga tgtcaatatc tccggtctgg
     1741 taagcacaac catgcagaat gaagcccgtc gtctgcgtgc cgaacgctgg aaagcggaaa
     1801 atcaggaagg gatggctgag gtcgcccggt ttattgaaat gaacggctct tttgctgacg
     1861 agaacagggg ctggtgaaat gcagtttaag gtttacacct ataaaagaga gagccgttat
     1921 cgtctgtttg tggatgtaca gagtgatatt attgacacgc ccgggcgacg gatggtgatc
     1981 cccctggcca gtgcacgtct gctgtcagat aaagtctccc gtgaacttta cccggtggtg
     2041 catatcgggg atgaaagctg gcgcatgatg accaccgata tggccagtgt gccggtctcc
     2101 gttatcgggg aagaagtggc tgatctcagc caccgcgaaa atgacatcaa aaacgccatt
     2161 aacctgatgt tctggggaat ataaatgtca ggctccctta tacacagcca gtctgcaggt
     2221 cgaccatagt gactggatat gttgtgtttt acagtattat gtagtctgtt ttttatgcaa
     2281 aatctaattt aatatattga tatttatatc attttacgtt tctcgttcag ctttcttgta
     2341 caaagtggtt gatcgcgtgc atgcgacgtc atagctctct ccctatagtg agtcgtatta
     2401 taagctaggc actggccgtc gttgatggga tccaccggat ctagataact gatcataatc
     2461 agccatacca catttgtaga ggttttactt gctttaaaaa acctcccaca cctccccctg
     2521 aacctgaaac ataaaatgaa tgcaattgtt gttgttaact tgtttattgc agcttataat
     2581 ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat
     2641 tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctggat cgatcctgca
     2701 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attggctggc gtaatagcga
     2761 agaggcccgc accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg aatgggacgc
     2821 gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac
     2881 acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt
     2941 cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc
     3001 tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc
     3061 gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact
     3121 cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg
     3181 gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc
     3241 gaattttaac aaaatattaa cgcttacaat ttcctgatgc ggtattttct ccttacgcat
     3301 ctgtgcggta tttcacaccg catacgcgga tctgcgcagc accatggcct gaaataacct
     3361 ctgaaagagg aacttggtta ggtaccttct gaggcggaaa gaaccagctg tggaatgtgt
     3421 gtcagttagg gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc
     3481 atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca ggcagaagta
     3541 tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact ccgcccatcc
     3601 cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta atttttttta
     3661 tttatgcaga ggccgaggcc gcctcggcct ctgagctatt ccagaagtag tgaggaggct
     3721 tttttggagg cctaggcttt tgcaaaaagc ttgattcttc tgacacaaca gtctcgaact
     3781 taaggctaga gccaccatga ttgaacaaga tggattgcac gcaggttctc cggccgcttg
     3841 ggtggagagg ctattcggct atgactgggc acaacagaca atcggctgct ctgatgccgc
     3901 cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg acctgtccgg
     3961 tgccctgaat gaactgcagg acgaggcagc gcggctatcg tggctggcca cgacgggcgt
     4021 tccttgcgca gctgtgctcg acgttgtcac tgaagcggga agggactggc tgctattggg
     4081 cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct cctgccgaga aagtatccat
     4141 catggctgat gcaatgcggc ggctgcatac gcttgatccg gctacctgcc cattcgacca
     4201 ccaagcgaaa catcgcatcg agcgagcacg tactcggatg gaagccggtc ttgtcgatca
     4261 ggatgatctg gacgaagagc atcaggggct cgcgccagcc gaactgttcg ccaggctcaa
     4321 ggcgcgcatg cccgacggcg aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa
     4381 tatcatggtg gaaaatggcc gcttttctgg attcatcgac tgtggccggc tgggtgtggc
     4441 ggaccgctat caggacatag cgttggctac ccgtgatatt gctgaagagc ttggcggcga
     4501 atgggctgac cgcttcctcg tgctttacgg tatcgccgct cccgattcgc agcgcatcgc
     4561 cttctatcgc cttcttgacg agttcttctg agcgggactc tggggttcga aatgaccgac
     4621 caagcgacgc ccaacctgcc atcacgatgg ccgcaataaa atatctttat tttcattaca
     4681 tctgtgtgtt ggttttttgt gtgaatcgat agcgataagg atccgcgtat ggtgcactct
     4741 cagtacaatc tgctctgatg ccgcatagtt aagccagccc cgacacccgc caacacccgc
     4801 tgacgcgccc tgacgggctt gtctgctccc ggcatccgct tacagacaag ctgtgaccgt
     4861 ctccgggagc tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg cgagacgaaa
     4921 gggcctcgtg atacgcctat ttttataggt taatgtcatg ataataatgg tttcttagac
     4981 gtcaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat
     5041 acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg
     5101 aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc
     5161 attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga
     5221 tcagttgggt gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga
     5281 gagttttcgc cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg
     5341 cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc
     5401 tcagaatgac ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac
     5461 agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact
     5521 tctgacaacg atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca
     5581 tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg
     5641 tgacaccacg atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact
     5701 acttactcta gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg
     5761 accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg
     5821 tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat
     5881 cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc
     5941 tgagataggt gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat
     6001 actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt
     6061 tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc
     6121 cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt
     6181 gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac
     6241 tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt
     6301 gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct
     6361 gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga
     6421 ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac
     6481 acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg
     6541 agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt
     6601 cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc
     6661 tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg
     6721 gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc
     6781 ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc
     6841 ctttgagtga gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag
     6901 cgaggaagcg gaagagcgcc caatacgcaa accgcctctc cccgcgcgtt ggccgattca
     6961 ttaatgcaga gcttgcaatt cgcgcttttt caatattatt gaagcattta tcagggttat
     7021 tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg
     7081 cgcacatttc cccgaaaagt gccacctgac gtctaagaaa ccattattat catgacatta
     7141 acctataaaa ataggcgtat tacgaggccc tttcacg
//
