LOCUS       sequence-175149-        6305 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  FRB-tagged N-terminal Venus (YFP) fragment for BiFC; localizes to 
            actin cytoskeleton..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6305)
  AUTHORS   Hertel F, Mo GC, Duwe S, Dedecker P, Zhang J
  TITLE     RefSOFI for Mapping Nanoscale Organization of Protein-Protein 
            Interactions in Living Cells.
  JOURNAL   Cell Rep. 2016 Jan 12;14(2):390-400. doi: 
            10.1016/j.celrep.2015.12.036. Epub 2015 Dec 31.
  PUBMED    26748717
REFERENCE   2  (bases 1 to 6305)
  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: 
            "10.1016/j.celrep.2015.12.036"; journalName: "Cell Rep"; date: 
            "2016-01-12- 12"; volume: "14"; issue: "2"; pages: "390-400"
COMMENT     Sequence Label: pcDNA3-LifeAct-FRB-Venus-173-N
FEATURES             Location/Qualifiers
     source          1..6305
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1..51
                     /codon_start=1
                     /product="peptide that binds filamentous actin (Riedl et 
                     al., 2008)"
                     /label=Lifeact
                     /note="first 17 amino acids of budding yeast Abp140"
                     /translation="MGVADLIKKFESISKEE"
     CDS             61..339
                     /codon_start=1
                     /product="T2098L mutant of FKBP-rapamycin binding domain of
                     human FRAP"
                     /label=FRB*
                     /note="binds synthetic ligands such as AP21967"
                     /translation="ILWHEMWHEGLEEASRLYFGERNVKGMFEVLEPLHAMMERGPQTL
                     KETSFNQAYGRDLMEAQEWCRKYMKSGNVKDLLQAWDLYYHVFRRISK"
     CDS             join(376..378,379..381,382..891)
                     /codon_start=1
                     /product="N-terminal fragment of mVenus for use in 
                     bimolecular fluorescence complementation (BiFC) (Kodama and
                     Hu, 2010)"
                     /label=VN173
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KLICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK
                     ANFKIRHNI"
     primer_bind     complement(421..442)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(682..701)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     promoter        complement(955..973)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(956..973)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(993..1010)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    999..1223
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1269..1697
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1356..1375)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1566..1587
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(1706..1726)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        1711..2040
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      1891..2026
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     1953..1972
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2107..2901
                     /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(2161..2180)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2771..2790
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3075..3196
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3112..3131)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3166..3185
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3245..3261)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3245..3261)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3258..3280)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3269..3285
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(3293..3299,3300..3317,3318..3323))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3338..3359
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3476..3493)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3647..4235)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3727..3746)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4406..5197,5198..5266))
                     /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     5029..5048
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5267..5371)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(5446..5465)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        5637..6016
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6017..6220
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     6171..6191
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     6265..6284
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        6265..6283
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      join(6300..6305,1..4)
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
ORIGIN
        1 atgggcgtgg ccgacctgat caagaagttc gagagcatca gcaaggagga gggggatccc
       61 atcctctggc atgagatgtg gcatgaaggc ctggaagagg catctcgttt gtactttggg
      121 gaaaggaacg tgaaaggcat gtttgaggtg ctggagccct tgcatgctat gatggaacgg
      181 ggcccccaga ctctgaagga aacatccttt aatcaggcct atggtcgaga tttaatggag
      241 gcccaagagt ggtgcaggaa gtacatgaaa tcagggaatg tcaaggacct cctccaagcc
      301 tgggacctct attatcatgt gttccgacga atctcaaaga ctagtggtac cggtggaggt
      361 agtggaggtg gaagcatggt gagcaagggc gaggagctgt tcaccggggt ggtgcccatc
      421 ctggtcgagc tggacggcga cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag
      481 ggcgatgcca cctacggcaa gctgaccctg aagctgatct gcaccaccgg caagctgccc
      541 gtgccctggc ccaccctcgt gaccaccctg ggctacggcc tgcagtgctt cgcccgctac
      601 cccgaccaca tgaagcagca cgacttcttc aagtccgcca tgcccgaagg ctacgtccag
      661 gagcgcacca tcttcttcaa ggacgacggc aactacaaga cccgcgccga ggtgaagttc
      721 gagggcgaca ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc
      781 aacatcctgg ggcacaagct ggagtacaac tacaacagcc acaacgtcta tatcaccgcc
      841 gacaagcaga agaacggcat caaggccaac ttcaagatcc gccacaacat ctaagaattc
      901 tgcagatatc catcacactg gcggccgctc gagcatgcat ctagagggcc ctattctata
      961 gtgtcaccta aatgctagag ctcgctgatc agcctcgact gtgccttcta gttgccagcc
     1021 atctgttgtt tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt
     1081 cctttcctaa taaaatgagg aaattgcatc gcattgtctg agtaggtgtc attctattct
     1141 ggggggtggg gtggggcagg acagcaaggg ggaggattgg gaagacaata gcaggcatgc
     1201 tggggatgcg gtgggctcta tggcttctga ggcggaaaga accagctggg gctctagggg
     1261 gtatccccac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag
     1321 cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct tcccttcctt
     1381 tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt
     1441 ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg atggttcacg
     1501 tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt
     1561 taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg tctattcttt
     1621 tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca
     1681 aaaatttaac gcgaattaat tctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca
     1741 ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc aaccaggtgt
     1801 ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca
     1861 gcaaccatag tcccgcccct aactccgccc atcccgcccc taactccgcc cagttccgcc
     1921 cattctccgc cccatggctg actaattttt tttatttatg cagaggccga ggccgcctct
     1981 gcctctgagc tattccagaa gtagtgagga ggcttttttg gaggcctagg cttttgcaaa
     2041 aagctcccgg gagcttgtat atccattttc ggatctgatc aagagacagg atgaggatcg
     2101 tttcgcatga ttgaacaaga tggattgcac gcaggttctc cggccgcttg ggtggagagg
     2161 ctattcggct atgactgggc acaacagaca atcggctgct ctgatgccgc cgtgttccgg
     2221 ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg acctgtccgg tgccctgaat
     2281 gaactgcagg acgaggcagc gcggctatcg tggctggcca cgacgggcgt tccttgcgca
     2341 gctgtgctcg acgttgtcac tgaagcggga agggactggc tgctattggg cgaagtgccg
     2401 gggcaggatc tcctgtcatc tcaccttgct cctgccgaga aagtatccat catggctgat
     2461 gcaatgcggc ggctgcatac gcttgatccg gctacctgcc cattcgacca ccaagcgaaa
     2521 catcgcatcg agcgagcacg tactcggatg gaagccggtc ttgtcgatca ggatgatctg
     2581 gacgaagagc atcaggggct cgcgccagcc gaactgttcg ccaggctcaa ggcgcgcatg
     2641 cccgacggcg aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg
     2701 gaaaatggcc gcttttctgg attcatcgac tgtggccggc tgggtgtggc ggaccgctat
     2761 caggacatag cgttggctac ccgtgatatt gctgaagagc ttggcggcga atgggctgac
     2821 cgcttcctcg tgctttacgg tatcgccgct cccgattcgc agcgcatcgc cttctatcgc
     2881 cttcttgacg agttcttctg agcgggactc tggggttcga aatgaccgac caagcgacgc
     2941 ccaacctgcc atcacgagat ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg
     3001 gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc atgctggagt
     3061 tcttcgccca ccccaacttg tttattgcag cttataatgg ttacaaataa agcaatagca
     3121 tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac
     3181 tcatcaatgt atcttatcat gtctgtatac cgtcgacctc tagctagagc ttggcgtaat
     3241 catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac
     3301 gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa
     3361 ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat
     3421 gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc
     3481 tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg
     3541 cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag
     3601 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc
     3661 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
     3721 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
     3781 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
     3841 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
     3901 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
     3961 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
     4021 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
     4081 ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
     4141 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
     4201 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg
     4261 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa
     4321 aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta
     4381 tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag
     4441 cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga
     4501 tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac
     4561 cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc
     4621 ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta
     4681 gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac
     4741 gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat
     4801 gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa
     4861 gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg
     4921 tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag
     4981 aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc
     5041 cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct
     5101 caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat
     5161 cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg
     5221 ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc
     5281 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
     5341 tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg
     5401 tcgacggatc gggagatctc ccgatcccct atggtgcact ctcagtacaa tctgctctga
     5461 tgccgcatag ttaagccagt atctgctccc tgcttgtgtg ttggaggtcg ctgagtagtg
     5521 cgcgagcaaa atttaagcta caacaaggca aggcttgacc gacaattgca tgaagaatct
     5581 gcttagggtt aggcgttttg cgctgcttcg cgatgtacgg gccagatata cgcgttgaca
     5641 ttgattattg actagttatt aatagtaatc aattacgggg tcattagttc atagcccata
     5701 tatggagttc cgcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga
     5761 cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt
     5821 ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt
     5881 gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca
     5941 ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt
     6001 catcgctatt accatggtga tgcggttttg gcagtacatc aatgggcgtg gatagcggtt
     6061 tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt tgttttggca
     6121 ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga cgcaaatggg
     6181 cggtaggcgt gtacggtggg aggtctatat aagcagagct ctctggctaa ctagagaacc
     6241 cactgcttac tggcttatcg aaattaatac gactcactat agggagaccc aagcttgccg
     6301 ccacc
//
