LOCUS       sequence-346295-        5694 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Constitutively active, mitochondrial localized RhoA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5694)
  AUTHORS   Mahlandt EK, Kreider-Letterman G, Chertkova AO, Garcia-Mata R, 
            Goedhart J
  TITLE     Cell-based optimization and characterization of genetically encoded 
            location-based biosensors for Cdc42 or Rac activity.
  JOURNAL   J Cell Sci. 2023 May 15;136(10):jcs260802. doi: 10.1242/jcs.260802. 
            Epub 2023 May 25.
  PUBMED    37226883
REFERENCE   2  (bases 1 to 5694)
  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: "J Cell 
            Sci."; date: "2023-05-15"; volume: "136(10):jcs260802. doi"; pages: 
            " 10.1242/jcs.260802. Epub 2023 May 25"
COMMENT     Sequence Label: TOMM20-mTurquoise-RhoA-G14V-deltaCaaX
FEATURES             Location/Qualifiers
     source          1..5694
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        133..336
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     287..307
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      365..374
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      830..839
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(836..838,839..841,842..1552)
                     /codon_start=1
                     /product="enhanced monomeric variant of CFP (Goedhart et 
                     al., 2010)"
                     /label=mTurquoise
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLSWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISDNVYITADKQKNGIK
                     ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(881..902)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1142..1161)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1489..1510
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2250..2371
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2287..2306)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2341..2360
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2378..2833)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2515..2536)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2727..2746
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2860..2964
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2966..3323
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2966..3157
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2989..3009)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3174..3309
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3358..4152
                     /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             3358..4152
                     /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(3412..3431)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4022..4041
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4340..4359)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4384..4431
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4760..5348
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5249..5268
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        join(5523..5694,1..132)
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 catatgccaa gtacgccccc tattgacgtc aatgacggta aatggcccgc ctggcattat
       61 gcccagtaca tgaccttatg ggactttcct acttggcagt acatctacgt attagtcatc
      121 gctattacca tggtgatgcg gttttggcag tacatcaatg ggcgtggata gcggtttgac
      181 tcacggggat ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa
      241 aatcaacggg actttccaaa atgtcgtaac aactccgccc cattgacgca aatgggcggt
      301 aggcgtgtac ggtgggaggt ctatataagc agagctggtt tagtgaaccg tcagatccgc
      361 tagcgccacc atggtgggtc ggaacagcgc catcgccgcc ggtgtatgcg gggccctttt
      421 cattgggtac tgcatctact tcgaccgcaa aagacgaagt gaccccaact tcaagaacag
      481 gcttcgagaa cgaagaaaga aacagaagct tgccaaggag agagctgggc tttccaagtt
      541 acctgacctt aaagatgctg aagctgttca gaagttcttc cttgaagaaa tacagcttgg
      601 tgaagagtta ctagctcaag gtgaatatga gaagggcgta gaccatctga caaatgcaat
      661 tgctgtgtgt ggacagccac agcagttact gcaggtctta cagcaaactc ttccaccacc
      721 agtgttccag atgcttctga ctaagctccc aacaattagt cagagaattg taagtgctca
      781 gagcttggct gaagatgatg tggaaggcgg tagcggggat ccaccggtcg ccaccatggt
      841 gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc tggacggcga
      901 cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa
      961 gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc ccaccctcgt
     1021 gaccaccctg tcctggggcg tgcagtgctt cgcccgctac cccgaccaca tgaagcagca
     1081 cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa
     1141 ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa
     1201 ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg ggcacaagct
     1261 ggagtacaac tacatcagcg acaacgtcta tatcaccgcc gacaagcaga agaacggcat
     1321 caaggccaac ttcaagatcc gccacaacat cgaggacggc ggcgtgcagc tcgccgacca
     1381 ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca accactacct
     1441 gagcacccag tccaagctga gcaaagaccc caacgagaag cgcgatcaca tggtcctgct
     1501 ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtaca agtccatggc
     1561 tgccatccgg aagaaactgg tgattgttgg tgatgtagcc tgtggaaaga catgcttgct
     1621 catagtcttc agcaaggacc agttcccaga ggtgtatgtg cccacagtgt ttgagaacta
     1681 tgtggcagat atcgaggtgg atggaaagca ggtagagttg gctttgtggg acacagctgg
     1741 gcaggaagat tatgatcgcc tgaggcccct ctcctaccca gataccgatg ttatactgat
     1801 gtgtttttcc atcgacagcc ctgatagttt agaaaacatc ccagaaaagt ggaccccaga
     1861 agtcaagcat ttctgtccca acgtgcccat catcctggtt gggaataaga aggatcttcg
     1921 gaatgatgag cacacaaggc gggagctagc caagatgaag caggagccgg tgaaacctga
     1981 agaaggcaga gatatggcaa acaggattgg cgcttttggg tacatggagt gttcagcaaa
     2041 gaccaaagat ggagtgagag aggtttttga aatggctacg agagctgctc tgcaagctag
     2101 acgtgggaag aaaaaatctg ggagctaagc ggccgcgact ctagatcata atcagccata
     2161 ccacatttgt agaggtttta cttgctttaa aaaacctccc acacctcccc ctgaacctga
     2221 aacataaaat gaatgcaatt gttgttgtta acttgtttat tgcagcttat aatggttaca
     2281 aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt
     2341 gtggtttgtc caaactcatc aatgtatctt aaggcgtaaa ttgtaagcgt taatattttg
     2401 ttaaaattcg cgttaaattt ttgttaaatc agctcatttt ttaaccaata ggccgaaatc
     2461 ggcaaaatcc cttataaatc aaaagaatag accgagatag ggttgagtgt tgttccagtt
     2521 tggaacaaga gtccactatt aaagaacgtg gactccaacg tcaaagggcg aaaaaccgtc
     2581 tatcagggcg atggcccact acgtgaacca tcaccctaat caagtttttt ggggtcgagg
     2641 tgccgtaaag cactaaatcg gaaccctaaa gggagccccc gatttagagc ttgacgggga
     2701 aagccggcga acgtggcgag aaaggaaggg aagaaagcga aaggagcggg cgctagggcg
     2761 ctggcaagtg tagcggtcac gctgcgcgta accaccacac ccgccgcgct taatgcgccg
     2821 ctacagggcg cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta
     2881 tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt
     2941 caataatatt gaaaaaggaa gagtcctgag gcggaaagaa ccagctgtgg aatgtgtgtc
     3001 agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc
     3061 tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
     3121 aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc
     3181 ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt
     3241 atgcagaggc cgaggccgcc tcggcctctg agctattcca gaagtagtga ggaggctttt
     3301 ttggaggcct aggcttttgc aaagatcgat caagagacag gatgaggatc gtttcgcatg
     3361 attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc
     3421 tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg
     3481 caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcaa
     3541 gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc
     3601 gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat
     3661 ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg
     3721 cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc
     3781 gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag
     3841 catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgagcat gcccgacggc
     3901 gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc
     3961 cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata
     4021 gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc
     4081 gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac
     4141 gagttcttct gagcgggact ctggggttcg aaatgaccga ccaagcgacg cccaacctgc
     4201 catcacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt
     4261 tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc
     4321 accctagggg gaggctaact gaaacacgga aggagacaat accggaagga acccgcgcta
     4381 tgacggcaat aaaaagacag aataaaacgc acggtgttgg gtcgtttgtt cataaacgcg
     4441 gggttcggtc ccagggctgg cactctgtcg ataccccacc gagaccccat tggggccaat
     4501 acgcccgcgt ttcttccttt tccccacccc accccccaag ttcgggtgaa ggcccagggc
     4561 tcgcagccaa cgtcggggcg gcaggccctg ccatagcctc aggttactca tatatacttt
     4621 agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata
     4681 atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag
     4741 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa
     4801 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt
     4861 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc
     4921 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa
     4981 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa
     5041 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc
     5101 ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa
     5161 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa
     5221 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg
     5281 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc
     5341 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg
     5401 ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgccatgc
     5461 attagttatt aatagtaatc aattacgggg tcattagttc atagcccata tatggagttc
     5521 cgcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga cccccgccca
     5581 ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt ccattgacgt
     5641 caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt gtat
//
