LOCUS       sequence-261659-        6570 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  RhoGEF.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6570)
  AUTHORS   Reinhard NR, Van Der Niet S, Chertkova A, Postma M, Hordijk PL, 
            Gadella TWJ Jr, Goedhart J
  TITLE     Identification of guanine nucleotide exchange factors that increase 
            Cdc42 activity in primary human endothelial cells.
  JOURNAL   Small GTPases. 2019 Aug 30:1-15. doi: 10.1080/21541248.2019.1658509.
  PUBMED    31469028
REFERENCE   2  (bases 1 to 6570)
  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: "Small 
            GTPases. 2019 Aug 30:1-15. doi: 10.1080/21541248.2019.1658509."
COMMENT     Sequence Label: mCherry-AlphaPix
FEATURES             Location/Qualifiers
     source          1..6570
                     /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      607..616
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             613..1320
                     /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(758..776)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1026..1045
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    3358..3479
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3395..3414)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3449..3468
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3486..3941)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3623..3644)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3835..3854
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3968..4072
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4074..4431
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4074..4265
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4097..4117)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4282..4417
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4466..5260
                     /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             4466..5260
                     /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(4520..4539)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5130..5149
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5448..5467)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5492..5539
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5868..6456
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6357..6376
                     /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 ccatggtgag caagggcgag gaggataaca tggccatcat caaggagttc
      661 atgcgcttca aggtgcacat ggagggctcc gtgaacggcc acgagttcga gatcgagggc
      721 gagggcgagg gccgccccta cgagggcacc cagaccgcca agctgaaggt gaccaagggt
      781 ggccccctgc ccttcgcctg ggacatcctg tcccctcagt tcatgtacgg ctccaaggcc
      841 tacgtgaagc accccgccga catccccgac tacttgaagc tgtccttccc cgagggcttc
      901 aagtgggagc gcgtgatgaa cttcgaggac ggcggcgtgg tgaccgtgac ccaggactcc
      961 tccctgcagg acggcgagtt catctacaag gtgaagctgc gcggcaccaa cttcccctcc
     1021 gacggccccg taatgcagaa gaagaccatg ggctgggagg cctcctccga gcggatgtac
     1081 cccgaggacg gcgccctgaa gggcgagatc aagcagaggc tgaagctgaa ggacggcggc
     1141 cactacgacg ctgaggtcaa gaccacctac aaggccaaga agcccgtgca gctgcccggc
     1201 gcctacaacg tcaacatcaa gttggacatc acctcccaca acgaggacta caccatcgtg
     1261 gaacagtacg aacgcgccga gggccgccac tccaccggcg gcatggacga gctgtacaag
     1321 tccggactca gatctcgagc tcaagcttcg aattccatga cggaaaatgg aagtcatcag
     1381 ttgatagtaa aagcaagatt caactttaag cagactaatg aggatgaact gtcagtttgt
     1441 aagggggaca tcatttacgt cacacgagtt gaagaaggag gctggtggga aggcacatta
     1501 aatgggagaa caggctggtt ccccagtaat tatgtccgtg aaattaaatc cagtgagaga
     1561 cctctctccc caaaagccgt caaaggattt gaaactgctc cacttaccaa gaattattat
     1621 actgtggtgt tacagaacat cctggacact gaaaaagaat atgctaaaga acttcagtct
     1681 cttcttgtta cttacttaag acccctgcag tccaataaca atctgagtac tgtggaggtt
     1741 acatctttac tgggaaactt cgaggaagta tgcacatttc aacagacact ctgccaagcc
     1801 ttggaagaat gttcaaagtt tccagaaaac cagcacaaag taggaggttg tctactgagt
     1861 ctcatgcctc attttaaatc tatgtatctg gcttactgtg caaaccatcc ttcagctgta
     1921 aatgtgctca ctcagcacag tgatgagttg gaacaattca tggaaaatca aggtgcatcg
     1981 agcccaggta tcctcatttt aacaacaaac ctcagcaaac cattcatgcg actggagaaa
     2041 tatgttactc tcttgcaaga gttagaacgg catatggagg atactcatcc agatcatcag
     2101 gatattctga aagcaatcgt agcattcaaa actctcatgg ggcaatgtca agatctgagg
     2161 aagagaaaac agctggagtt acagatactg tccgaaccta ttcaggcatg ggaaggagaa
     2221 gatattaaaa acttgggaaa tgtgattttt atgtcacaag taatggtgca gtatggagca
     2281 tgtgaggaaa aagaggagcg gtaccttatg ttattttcaa atgtcctgat aatgttatct
     2341 gcaagtcctc ggatgagtgg ctttatctat cagggaaaaa taccaatagc aggaacggtg
     2401 gtgactagat tagatgaaat tgaagggaat gactgcacat ttgaaatcac tggtaacaca
     2461 gtggagagaa ttgtggtcca ttgtaacaac aaccaggact tccaggaatg gttggagcag
     2521 ctgaacagac tgatcagagg acctgcctct tgcagttcat tatccaaaac ctcatcgtca
     2581 tcatgtagtg ctcattcttc ttttagctct accggacagc cccgaggacc cttggaacct
     2641 cctcaaatta taaaaccgtg gagtttaagt tgtctacgac ctgcacctcc acttagacca
     2701 tcagcagcac taggttataa agagaggatg tcttatatct taaaggagtc tagtaaaagc
     2761 cctaaaacga tgaagaaatt tcttcataaa aggaagactg agagaaaacc atcggaggag
     2821 gaatatgtga ttaggaaaag tacagctgct ctggaagagg atgctcaaat ccttaaagtg
     2881 atcgaagcct actgcaccag cgcaaatttt caacaaggcc atggctcaag tactcgaaaa
     2941 gattccattc cacaagtcct actccctgag gaagagaaac tcatcattga agaaaccaga
     3001 agcaacggcc agaccatcat ggaagaaaag agccttgttg atactgttta cgccttgaag
     3061 gacgaggtca gagaactgaa gcaggaaaat aaaagaatga agcaatgcct ggaagaagaa
     3121 ctgaaatcaa gaagggacct agaaaagctg gtgcggaggc ttttgaagca aacagatgag
     3181 tgtattcgag gcgagtccag tagcaagacc tcaattcttc cataataagg atccaccgga
     3241 tctagataac tgatcataat cagccatacc acatttgtag aggttttact tgctttaaaa
     3301 aacctcccac acctccccct gaacctgaaa cataaaatga atgcaattgt tgttgttaac
     3361 ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat
     3421 aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttaa
     3481 ggcgtaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt gttaaatcag
     3541 ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa aagaatagac
     3601 cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa agaacgtgga
     3661 ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac gtgaaccatc
     3721 accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg
     3781 gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa aggaagggaa
     3841 gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac
     3901 caccacaccc gccgcgctta atgcgccgct acagggcgcg tcaggtggca cttttcgggg
     3961 aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct
     4021 catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga gtcctgaggc
     4081 ggaaagaacc agctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca
     4141 gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc
     4201 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata
     4261 gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg
     4321 ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc ggcctctgag
     4381 ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa agatcgatca
     4441 agagacagga tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg caggttctcc
     4501 ggccgcttgg gtggagaggc tattcggcta tgactgggca caacagacaa tcggctgctc
     4561 tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg gttctttttg tcaagaccga
     4621 cctgtccggt gccctgaatg aactgcaaga cgaggcagcg cggctatcgt ggctggccac
     4681 gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa gggactggct
     4741 gctattgggc gaagtgccgg ggcaggatct cctgtcatct caccttgctc ctgccgagaa
     4801 agtatccatc atggctgatg caatgcggcg gctgcatacg cttgatccgg ctacctgccc
     4861 attcgaccac caagcgaaac atcgcatcga gcgagcacgt actcggatgg aagccggtct
     4921 tgtcgatcag gatgatctgg acgaagagca tcaggggctc gcgccagccg aactgttcgc
     4981 caggctcaag gcgagcatgc ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg
     5041 cttgccgaat atcatggtgg aaaatggccg cttttctgga ttcatcgact gtggccggct
     5101 gggtgtggcg gaccgctatc aggacatagc gttggctacc cgtgatattg ctgaagagct
     5161 tggcggcgaa tgggctgacc gcttcctcgt gctttacggt atcgccgctc ccgattcgca
     5221 gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga gcgggactct ggggttcgaa
     5281 atgaccgacc aagcgacgcc caacctgcca tcacgagatt tcgattccac cgccgccttc
     5341 tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg gctggatgat cctccagcgc
     5401 ggggatctca tgctggagtt cttcgcccac cctaggggga ggctaactga aacacggaag
     5461 gagacaatac cggaaggaac ccgcgctatg acggcaataa aaagacagaa taaaacgcac
     5521 ggtgttgggt cgtttgttca taaacgcggg gttcggtccc agggctggca ctctgtcgat
     5581 accccaccga gaccccattg gggccaatac gcccgcgttt cttccttttc cccaccccac
     5641 cccccaagtt cgggtgaagg cccagggctc gcagccaacg tcggggcggc aggccctgcc
     5701 atagcctcag gttactcata tatactttag attgatttaa aacttcattt ttaatttaaa
     5761 aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt
     5821 tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt
     5881 tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt
     5941 ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag
     6001 ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa gaactctgta
     6061 gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat
     6121 aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg
     6181 ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg
     6241 agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac
     6301 aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga
     6361 aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt
     6421 ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta
     6481 cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat
     6541 tctgtggata accgtattac cgccatgcat
//
