LOCUS       sequence-118094-        6432 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses a fusion of mCherry with full-length p63RhoGEF in 
            mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6432)
  AUTHORS   van Unen J, Reinhard NR, Yin T, Wu YI, Postma M, Gadella TW, 
            Goedhart J
  TITLE     Plasma membrane restricted RhoGEF activity is sufficient for 
            RhoA-mediated actin polymerization.
  JOURNAL   Sci Rep. 2015 Oct 5;5:14693. doi: 10.1038/srep14693.
  PUBMED    26435194
REFERENCE   2  (bases 1 to 6432)
  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: "Sci Rep."; 
            date: "2015-10-5"; volume: "5:14693. doi"; pages: " 
            10.1038/srep14693"
COMMENT     Sequence Label: pmCherry-C1-p63RhoGEF
FEATURES             Location/Qualifiers
     source          1..6432
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      619..628
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             625..1332
                     /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(770..788)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1038..1057
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             1336..1419
                     /codon_start=1
                     /product="palmitoylation signal consisting of the first 29 
                     amino acids of p63RhoGEF (Goedhart et al., 2013)"
                     /label=palmitoylation signal
                     /translation="RGGHKGGRCACPRVIRKVLAKCGCCFAR"
     regulatory      1894..1903
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    3220..3341
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3257..3276)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3311..3330
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3348..3803)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3485..3506)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3697..3716
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3830..3934
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3936..4293
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3936..4127
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3959..3979)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4144..4279
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4328..5122
                     /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             4328..5122
                     /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(4382..4401)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4992..5011
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5310..5329)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5354..5401
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5730..6318
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6219..6238
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggtcgcca ccccggtcgc caccatggtg agcaagggcg aggaggataa catggccatc
      661 atcaaggagt tcatgcgctt caaggtgcac atggagggct ccgtgaacgg ccacgagttc
      721 gagatcgagg gcgagggcga gggccgcccc tacgagggca cccagaccgc caagctgaag
      781 gtgaccaagg gtggccccct gcccttcgcc tgggacatcc tgtcccctca gttcatgtac
      841 ggctccaagg cctacgtgaa gcaccccgcc gacatccccg actacttgaa gctgtccttc
      901 cccgagggct tcaagtggga gcgcgtgatg aacttcgagg acggcggcgt ggtgaccgtg
      961 acccaggact cctccctgca ggacggcgag ttcatctaca aggtgaagct gcgcggcacc
     1021 aacttcccct ccgacggccc cgtaatgcag aagaagacca tgggctggga ggcctcctcc
     1081 gagcggatgt accccgagga cggcgccctg aagggcgaga tcaagcagag gctgaagctg
     1141 aaggacggcg gccactacga cgctgaggtc aagaccacct acaaggccaa gaagcccgtg
     1201 cagctgcccg gcgcctacaa cgtcaacatc aagttggaca tcacctccca caacgaggac
     1261 tacaccatcg tggaacagta cgaacgcgcc gagggccgcc actccaccgg cggcatggac
     1321 gagctgtaca agtcccgggg ggggcacaaa gggggtcgct gtgcctgtcc ccgtgtgatc
     1381 cgaaaagtgc tggcaaaatg cggctgctgc ttcgcccggg ggggacgtga atcctattcc
     1441 attgcgggca gtgaggggag tatatcggct tctgctgcct ccggtctggc tgccccctct
     1501 ggccccagct ctggcctcag ctctggcccc tgttccccag gccccccagg gcccgtcagt
     1561 ggcctgagga gatggttgga tcattccaaa cattgtctca gtgtggaaac tgaggcagac
     1621 agtggtcagg caggaccata tgagaactgg atgttggagc cagctctagc cacaggagag
     1681 gagctgccgg aactgacctt gctgaccaca ctgttggagg gccctggaga taagacgcag
     1741 ccacctgaag aggagacttt gtcccaagcc cctgagagtg aggaggaaca gaagaagaag
     1801 gctctggaaa ggagtatgta tgtcctgagt gaactggtag aaacagagaa aatgtacgtg
     1861 gacgacttgg ggcagattgt ggagggttat atggccacca tggctgctca gggggtcccc
     1921 gagagtcttc gaggccgtga caggattgtg tttgggaata tccagcaaat ctatgagtgg
     1981 caccgagact atttcttgca agagctacaa cggtgtctga aagatcctga ttggctggct
     2041 cagctattca tcaaacacga gcgccggctg catatgtatg tggtgtactg tcagaataag
     2101 cccaagtcag agcatgtggt gtcagagttt ggggacagct actttgagga gctccggcag
     2161 cagctggggc accgcctgca gctgaacgac ctcctcatca aacctgtgca gcggatcatg
     2221 aaataccagc tgctgctcaa ggattttctc aagtattaca atagagctgg gatggatact
     2281 gcagacctag agcaagctgt ggaggtcatg tgctttgtgc ccaagcgctg caacgatatg
     2341 atgacgctgg ggagattgcg gggatttgag ggcaaactga ctgctcaggg gaagctcttg
     2401 ggccaggaca ctttctgggt caccgagcct gaggctggag ggctgctgtc ttcccgaggt
     2461 cgagagaggc gcgtcttcct ctttgagcaa atcatcatct tcagtgaagc cctgggagga
     2521 ggagtgagag gtggaacaca gcctggatat gtatacaaga acagcattaa ggtgagctgc
     2581 ctgggactgg aggggaacct ccaaggtgac ccttgccgct ttgcactgac ctccagaggg
     2641 ccagagggtg ggatccagcg ctatgtcctg caggctgcag accctgctat cagtcaggcc
     2701 tggatcaagc atgtggctca gatcttggag agccaacggg acttcctcaa cgcattgcag
     2761 tcacccattg agtaccagag acgggagagc cagaccaaca gcctggggcg gccaagaggg
     2821 cctggagtgg ggagccctgg aagaattcgg cttggagatc aggcccaggg cagcacacac
     2881 acacccatca atggctctct cccctctctg ctgctgtcac ccaaagggga ggtggccaga
     2941 gccctcttgc cactggataa acaggccctt ggtgacatcc cccaggctcc ccatgactct
     3001 cctccagtct ctccaactcc aaaaacccct ccctgccaag ccagacttgc caagctggat
     3061 gaagatgagc tgtaaggtac cgcgggcccg ggatccaccg gatctagata actgatcata
     3121 atcagccata ccacatttgt agaggtttta cttgctttaa aaaacctccc acacctcccc
     3181 ctgaacctga aacataaaat gaatgcaatt gttgttgtta acttgtttat tgcagcttat
     3241 aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg
     3301 cattctagtt gtggtttgtc caaactcatc aatgtatctt aacgcgtaaa ttgtaagcgt
     3361 taatattttg ttaaaattcg cgttaaattt ttgttaaatc agctcatttt ttaaccaata
     3421 ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag accgagatag ggttgagtgt
     3481 tgttccagtt tggaacaaga gtccactatt aaagaacgtg gactccaacg tcaaagggcg
     3541 aaaaaccgtc tatcagggcg atggcccact acgtgaacca tcaccctaat caagtttttt
     3601 ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa gggagccccc gatttagagc
     3661 ttgacgggga aagccggcga acgtggcgag aaaggaaggg aagaaagcga aaggagcggg
     3721 cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta accaccacac ccgccgcgct
     3781 taatgcgccg ctacagggcg cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc
     3841 tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg
     3901 ataaatgctt caataatatt gaaaaaggaa gagtcctgag gcggaaagaa ccagctgtgg
     3961 aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa
     4021 agcatgcatc tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc
     4081 agaagtatgc aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg
     4141 cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt
     4201 ttttttattt atgcagaggc cgaggccgcc tcggcctctg agctattcca gaagtagtga
     4261 ggaggctttt ttggaggcct aggcttttgc aaagatcgat caagagacag gatgaggatc
     4321 gtttcgcatg attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag
     4381 gctattcggc tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg
     4441 gctgtcagcg caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa
     4501 tgaactgcaa gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc
     4561 agctgtgctc gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc
     4621 ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca tcatggctga
     4681 tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa
     4741 acatcgcatc gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct
     4801 ggacgaagag catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgagcat
     4861 gcccgacggc gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt
     4921 ggaaaatggc cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta
     4981 tcaggacata gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga
     5041 ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg
     5101 ccttcttgac gagttcttct gagcgggact ctggggttcg aaatgaccga ccaagcgacg
     5161 cccaacctgc catcacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc
     5221 ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag
     5281 ttcttcgccc accctagggg gaggctaact gaaacacgga aggagacaat accggaagga
     5341 acccgcgcta tgacggcaat aaaaagacag aataaaacgc acggtgttgg gtcgtttgtt
     5401 cataaacgcg gggttcggtc ccagggctgg cactctgtcg ataccccacc gagaccccat
     5461 tggggccaat acgcccgcgt ttcttccttt tccccacccc accccccaag ttcgggtgaa
     5521 ggcccagggc tcgcagccaa cgtcggggcg gcaggccctg ccatagcctc aggttactca
     5581 tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc
     5641 ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca
     5701 gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc
     5761 tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta
     5821 ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgtcctt
     5881 ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc
     5941 gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg
     6001 ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg
     6061 tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag
     6121 ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc
     6181 agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat
     6241 agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg
     6301 gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc
     6361 tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt
     6421 accgccatgc at
//
