LOCUS       sequence-261607-        6749 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 6749)
  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 6749)
  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-BetaPix
FEATURES             Location/Qualifiers
     source          1..6749
                     /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"
     CDS             1384..1413
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     polyA_signal    3548..3669
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3585..3604)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3639..3658
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3676..4131)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3813..3834)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4025..4044
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4158..4262
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4264..4621
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4264..4455
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4287..4307)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4472..4607
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4656..5450
                     /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             4656..5450
                     /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(4710..4729)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5320..5339
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5638..5657)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5682..5729
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6058..6646
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6547..6566
                     /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 aattctgcag tcgacggtac cgcgggcccc
     1381 atggagcaga agctgatctc cgaggaggac ctgggatccg acatgaccga taatagcaac
     1441 aatcaactgg tagtaagagc aaagtttaac ttccagcaga ccaatgagga cgagctttcc
     1501 ttctcaaaag gagacgtcat ccatgtcacc cgtgtggaag agggaggctg gtgggagggc
     1561 acactcaacg gccggaccgg ctggttcccc agcaactacg tgcgcgaggt caaggccagc
     1621 gagaagcctg tgtctcccaa atcaggaaca ctgaagagcc ctcccaaagg atttgatacg
     1681 actgccataa acaaaagcta ttacaatgtg gtgctacaga atattttaga aacagaaaat
     1741 gaatattcta aagaacttca gactgtgctt tcaacgtacc tacggccatt gcagaccagt
     1801 gagaagttaa gttcagcaaa catttcatat ttaatgggaa atctagaaga aatatgttct
     1861 ttccagcaaa tgctcgtaca gtctttagaa gaatgcacca agttgcccga agctcagcag
     1921 agagtcggag gctgcttttt aaacctgatg ccacagatga aaaccctgta cctcacgtat
     1981 tgtgccaatc acccttctgc agtgaatgtc ctcacggaac acagtgagga gttgggggag
     2041 ttcatggaga ccaaaggtgc cagcagccct gggattctcg tgctgaccac gggcctgagc
     2101 aaacccttca tgcgcctgga taaataccct acgctgctca aagagctcga gagacacatg
     2161 gaggattatc atacagatag acaagatatt caaaaatcca tggctgcctt caaaaacctt
     2221 tcagcccaat gtcaagaagt ccggaagagg aaagagcttg agctgcagat cctgacggaa
     2281 gccatccgga actgggaggg cgatgacatt aaaactctgg gcaacgtcac ttacatgtcc
     2341 caggtcctga ttcagtgtgc cggaagtgag gaaaagaatg aaagatatct tctactcttc
     2401 ccaaatgttt tgctaatgtt gtctgccagt cctaggatga gtggctttat ctatcaggga
     2461 aagcttccaa cgacaggaat gacaatcaca aagcttgagg acagtgaaaa tcatagaaat
     2521 gcatttgaaa tatcagggag catgattgag cggatattag tgtcgtgcaa caaccagcag
     2581 gatctgcagg aatgggtgga gcacctacag aagcaaacga aggtcacgtc tgtgggaaac
     2641 cccaccataa agcctcattc agtgccatct cataccctcc cctcccaccc ggtcactccg
     2701 tccagcaagc acgcagacag caagcccgcg ccgctgacgc ccgcctacca cacgctgccc
     2761 cacccctccc accacggcac cccgcacacc accatcaact ggggacccct ggagcctccg
     2821 aaaacaccca agccctggag cctgagctgc ctgcggcccg cgcctcccct ccggccctca
     2881 gctgctctct gctacaagga ggatcttagt aagagcccta agaccatgaa aaagctgctg
     2941 cccaagcgca aacctgaacg gaagccttca gatgaggagt tcgcgtcccg gaaaagcaca
     3001 gctgctttgg aagaagatgc tcagattctg aaagtcattg aagcttactg caccagcgcc
     3061 aaaacaaggc aaacactcaa ttcaagttca cgcaaagaat ctgctccaca agttttgctt
     3121 ccagaagaag agaaaattat agtggaagaa actaaaagta atggtcagac agtgatagaa
     3181 gaaaagagtc ttgtggatac cgtatatgca ttaaaggatg aagttcaaga attaagacag
     3241 gacaacaaaa agatgaagaa atctctagag gaagaacaga gagcccgcaa agacctggag
     3301 aagctggtga ggaaagtcct gaagaacatg aatgatcctg cctgggatga gaccaatcta
     3361 taagggacgt cctcagttct ttctgttgaa gaccagttct gaggtgaagc tggaattggg
     3421 atccaccgga tctagataac tgatcataat cagccatacc acatttgtag aggttttact
     3481 tgctttaaaa aacctcccac acctccccct gaacctgaaa cataaaatga atgcaattgt
     3541 tgttgttaac ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa
     3601 tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa
     3661 tgtatcttaa cgcgtaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt
     3721 gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa
     3781 aagaatagac cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa
     3841 agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac
     3901 gtgaaccatc accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga
     3961 accctaaagg gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa
     4021 aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc
     4081 tgcgcgtaac caccacaccc gccgcgctta atgcgccgct acagggcgcg tcaggtggca
     4141 cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata
     4201 tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga
     4261 gtcctgaggc ggaaagaacc agctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc
     4321 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg
     4381 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc
     4441 agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc
     4501 ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc
     4561 ggcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa
     4621 agatcgatca agagacagga tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg
     4681 caggttctcc ggccgcttgg gtggagaggc tattcggcta tgactgggca caacagacaa
     4741 tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg gttctttttg
     4801 tcaagaccga cctgtccggt gccctgaatg aactgcaaga cgaggcagcg cggctatcgt
     4861 ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa
     4921 gggactggct gctattgggc gaagtgccgg ggcaggatct cctgtcatct caccttgctc
     4981 ctgccgagaa agtatccatc atggctgatg caatgcggcg gctgcatacg cttgatccgg
     5041 ctacctgccc attcgaccac caagcgaaac atcgcatcga gcgagcacgt actcggatgg
     5101 aagccggtct tgtcgatcag gatgatctgg acgaagagca tcaggggctc gcgccagccg
     5161 aactgttcgc caggctcaag gcgagcatgc ccgacggcga ggatctcgtc gtgacccatg
     5221 gcgatgcctg cttgccgaat atcatggtgg aaaatggccg cttttctgga ttcatcgact
     5281 gtggccggct gggtgtggcg gaccgctatc aggacatagc gttggctacc cgtgatattg
     5341 ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt gctttacggt atcgccgctc
     5401 ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga gcgggactct
     5461 ggggttcgaa atgaccgacc aagcgacgcc caacctgcca tcacgagatt tcgattccac
     5521 cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg gctggatgat
     5581 cctccagcgc ggggatctca tgctggagtt cttcgcccac cctaggggga ggctaactga
     5641 aacacggaag gagacaatac cggaaggaac ccgcgctatg acggcaataa aaagacagaa
     5701 taaaacgcac ggtgttgggt cgtttgttca taaacgcggg gttcggtccc agggctggca
     5761 ctctgtcgat accccaccga gaccccattg gggccaatac gcccgcgttt cttccttttc
     5821 cccaccccac cccccaagtt cgggtgaagg cccagggctc gcagccaacg tcggggcggc
     5881 aggccctgcc atagcctcag gttactcata tatactttag attgatttaa aacttcattt
     5941 ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta
     6001 acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg
     6061 agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc
     6121 ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag
     6181 cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa
     6241 gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc
     6301 cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc
     6361 gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta
     6421 caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag
     6481 aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct
     6541 tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga
     6601 gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc
     6661 ggccttttta cggttcctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt
     6721 ctgtggataa ccgtattacc gccatgcat
//
