LOCUS       sequence-351694-        6028 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Mammalian expression of Vav2(DH-PH-CRD) fused to mCherry-Zdk1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6028)
  AUTHORS   Yoshikawa M, Yoshii T, Ikuta M, Tsukiji S
  TITLE     Synthetic Protein Condensates That Inducibly Recruit and Release 
            Protein Activity in Living Cells.
  JOURNAL   J Am Chem Soc. 2021 May 5;143(17):6434-6446. doi: 
            10.1021/jacs.0c12375. Epub 2021 Apr 23.
  PUBMED    33890764
REFERENCE   2  (bases 1 to 6028)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1021/jacs.0c12375"; journalName: "J Am Chem Soc"; date: 
            "2021-05-5- 5"; volume: "143"; issue: "17"; pages: "6434-6446"
COMMENT     Sequence Label: pCMV-mCherry-Zdk1-Vav2cat
FEATURES             Location/Qualifiers
     source          1..6028
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        66..369
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        370..573
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     524..544
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      612..621
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             618..1325
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(763..781)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1031..1050
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             1347..1523
                     /codon_start=1
                     /product="""Zdark"" protein, derived from the Z subunit of 
                     protein A, that binds to the dark state of LOV2 and is 
                     released upon irradiation (Wang et al., 2016)"
                     /label=Zdk1
                     /translation="MVDNKFNKEKTRAGAEIHSLPNLNVEQKFAFIVSLFDDPSQSANL
                     LAEAKKLNDAQAPK"
     polyA_signal    2821..2942
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2858..2877)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2912..2931
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2949..3404)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3086..3107)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3298..3317
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3431..3535
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3537..3894
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3537..3728
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3560..3580)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3745..3880
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3929..4723
                     /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             3929..4723
                     /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(3983..4002)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4593..4612
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4911..4930)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4955..5002
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5331..5919
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5820..5839
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tgcattagtt attaatagta atcaattacg gggtcattag ttcatagccc atatatggag
       61 ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa cgacccccgc
      121 ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac tttccattga
      181 cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca agtgtatcat
      241 atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc
      301 cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt agtcatcgct
      361 attaccatgg tgatgcggtt ttggcagtac atcaatgggc gtggatagcg gtttgactca
      421 cggggatttc caagtctcca ccccattgac gtcaatggga gtttgttttg gcaccaaaat
      481 caacgggact ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat gggcggtagg
      541 cgtgtacggt gggaggtcta tataagcaga gctggtttag tgaaccgtca gatccgctag
      601 cgctaccggt cgccaccatg gtgagcaagg gcgaggagga taacatggcc atcatcaagg
      661 agttcatgcg cttcaaggtg cacatggagg gctccgtgaa cggccacgag ttcgagatcg
      721 agggcgaggg cgagggccgc ccctacgagg gcacccagac cgccaagctg aaggtgacca
      781 agggtggccc cctgcccttc gcctgggaca tcctgtcccc tcagttcatg tacggctcca
      841 aggcctacgt gaagcacccc gccgacatcc ccgactactt gaagctgtcc ttccccgagg
      901 gcttcaagtg ggagcgcgtg atgaacttcg aggacggcgg cgtggtgacc gtgacccagg
      961 actcctccct gcaggacggc gagttcatct acaaggtgaa gctgcgcggc accaacttcc
     1021 cctccgacgg ccccgtaatg cagaagaaga ccatgggctg ggaggcctcc tccgagcgga
     1081 tgtaccccga ggacggcgcc ctgaagggcg agatcaagca gaggctgaag ctgaaggacg
     1141 gcggccacta cgacgctgag gtcaagacca cctacaaggc caagaagccc gtgcagctgc
     1201 ccggcgccta caacgtcaac atcaagttgg acatcacctc ccacaacgag gactacacca
     1261 tcgtggaaca gtacgaacgc gccgagggcc gccactccac cggcggcatg gacgagctgt
     1321 acaagtccgg actcagatct cgagctatgg tggataacaa attcaataaa gaaaagacgc
     1381 gtgccggtgc ggaaatccat tctctgccaa acctgaatgt tgagcagaag tttgccttca
     1441 tcgtgagcct gtttgatgat ccatctcaga gcgccaatct gctggccgaa gccaaaaaac
     1501 tgaacgatgc ccaggcccca aaaactagta gatctcgagg gcccatgaaa atgggcatga
     1561 ctgaggacga caagagaagc tgctgcttgt tagagattca ggagaccgag gccaagtact
     1621 accgcaccct ggaggacatt gagaagaact acatgggtcc cttgcggctg gtgctgagcc
     1681 cggcggatat ggctgctgtc ttcatcaacc tggaggacct catcaaggtg catcacagct
     1741 ttctgcgagc catcgatgtg tccatgatgg ctggtggcag taccctggct aaggtctttc
     1801 tggagtttaa ggaaaggctc ctgatctatg gagagtactg tagccacatg gaacacgctc
     1861 agagtacact gaaccagctc ctcgccagcc gagaggactt caggcagaaa gtggaggagt
     1921 gcacactcag ggttcaggat ggcaagttca agctgcaaga cctgctggtg gtgcccatgc
     1981 aacgggtgct gaagtaccac ctgctgctca aggagctcct gagccattct gcagaccgac
     2041 cagaaagaca acagctcaaa gaagccctgg aagccatgca ggacttggcc atgtacatca
     2101 atgaagtgaa gcgggacaag gagaccttga agaagattag cgagttccag tgctccatag
     2161 aaaacctgca agtgaagctg gaggaatttg ggaggccaaa gattgacggg gagcttaaag
     2221 tccggtccat agtcaaccac accaagcaag acaggtacct gttcctattt gacaaggtgg
     2281 tcatcgtgtg caagaggaag ggctacagct atgagctgaa ggaggtcatt gagctgctct
     2341 tccacaagat gaccgatgac ccgatgcaca acaaggacat caagaagtgg tcctatggct
     2401 tctacctgat tcacctccaa ggaaagcaag gctttcagtt cttctgcaag acggaagaca
     2461 tgaagcggaa gtggatggag cagttcgaga tggccatgtc aaacatcaag ccagataagg
     2521 ccaatgccaa ccatcatagc ttccagatgt acacattcga caagactacc aactgcaaag
     2581 cctgcaagat gtttctcagg ggtaccttct accagggata cctgtgtacc agatgtggcg
     2641 tcggggcaca caaggaatgc ctggaggtga tccccccctg caagtaaccc gggatccacc
     2701 ggatctagat aactgatcat aatcagccat accacatttg tagaggtttt acttgcttta
     2761 aaaaacctcc cacacctccc cctgaacctg aaacataaaa tgaatgcaat tgttgttgtt
     2821 aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca
     2881 aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct
     2941 taaggcgtaa attgtaagcg ttaatatttt gttaaaattc gcgttaaatt tttgttaaat
     3001 cagctcattt tttaaccaat aggccgaaat cggcaaaatc ccttataaat caaaagaata
     3061 gaccgagata gggttgagtg ttgttccagt ttggaacaag agtccactat taaagaacgt
     3121 ggactccaac gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac tacgtgaacc
     3181 atcaccctaa tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc ggaaccctaa
     3241 agggagcccc cgatttagag cttgacgggg aaagccggcg aacgtggcga gaaaggaagg
     3301 gaagaaagcg aaaggagcgg gcgctagggc gctggcaagt gtagcggtca cgctgcgcgt
     3361 aaccaccaca cccgccgcgc ttaatgcgcc gctacagggc gcgtcaggtg gcacttttcg
     3421 gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc
     3481 gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga agagtcctga
     3541 ggcggaaaga accagctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc
     3601 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag
     3661 tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc
     3721 atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct
     3781 ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc ctcggcctct
     3841 gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg caaagatcga
     3901 tcaagagaca ggatgaggat cgtttcgcat gattgaacaa gatggattgc acgcaggttc
     3961 tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga caatcggctg
     4021 ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac
     4081 cgacctgtcc ggtgccctga atgaactgca agacgaggca gcgcggctat cgtggctggc
     4141 cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg gaagggactg
     4201 gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg ctcctgccga
     4261 gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc cggctacctg
     4321 cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga tggaagccgg
     4381 tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt
     4441 cgccaggctc aaggcgagca tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc
     4501 ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg actgtggccg
     4561 gctgggtgtg gcggaccgct atcaggacat agcgttggct acccgtgata ttgctgaaga
     4621 gcttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg ctcccgattc
     4681 gcagcgcatc gccttctatc gccttcttga cgagttcttc tgagcgggac tctggggttc
     4741 gaaatgaccg accaagcgac gcccaacctg ccatcacgag atttcgattc caccgccgcc
     4801 ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat gatcctccag
     4861 cgcggggatc tcatgctgga gttcttcgcc caccctaggg ggaggctaac tgaaacacgg
     4921 aaggagacaa taccggaagg aacccgcgct atgacggcaa taaaaagaca gaataaaacg
     4981 cacggtgttg ggtcgtttgt tcataaacgc ggggttcggt cccagggctg gcactctgtc
     5041 gataccccac cgagacccca ttggggccaa tacgcccgcg tttcttcctt ttccccaccc
     5101 caccccccaa gttcgggtga aggcccaggg ctcgcagcca acgtcggggc ggcaggccct
     5161 gccatagcct caggttactc atatatactt tagattgatt taaaacttca tttttaattt
     5221 aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag
     5281 ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct
     5341 ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt
     5401 tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg
     5461 cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct
     5521 gtatcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc
     5581 gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg
     5641 tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa
     5701 ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg
     5761 gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg
     5821 ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga
     5881 tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt
     5941 ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct
     6001 gattctgtgg ataaccgtat taccgcca
//
