LOCUS       sequence-120972-        7608 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  FRET-based biosensor reporting on endogenous Cdc42 activation (for 
            expression in E. coli, insect and vertebrate cells).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7608)
  AUTHORS   Martin K, Reimann A, Fritz RD, Ryu H, Jeon NL, Pertz O
  TITLE     Spatio-temporal co-ordination of RhoA, Rac1 and Cdc42 activation 
            during prototypical edge protrusion and retraction dynamics.
  JOURNAL   Sci Rep. 2016 Feb 25;6:21901. doi: 10.1038/srep21901.
  PUBMED    26912264
REFERENCE   2  (bases 1 to 7608)
  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: "2016-02-25"; volume: "6:21901. doi"; pages: " 
            10.1038/srep21901"
COMMENT     Sequence Label: pTriEx4-Cdc42-2G
FEATURES             Location/Qualifiers
     source          1..7608
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_recomb     181..1008
                     /label=baculovirus recombination region (lef2/ORF603)
                     /note="contains ORF603 and part of lef2"
     CDS             181..305
                     /codon_start=3
                     /product="baculovirus late expression factor 2"
                     /label=lef2
                     /translation="HLLIKSQDVYKPPNCQKMKTVDKLCPFAGNCKGLNPICNY"
     CDS             complement(343..948)
                     /codon_start=1
                     /product="baculovirus ORF603 protein"
                     /label=ORF603
                     /translation="MAVIFNNKQLLADNSIEKGGELFLFNGSYNILESYVNPVLLKNGV
                     IELEEAAYYAGNILYKTDDPKFIDYINLIIKATHSEELPENSTVVNYRKTMRSGTIHPI
                     KKDIYIYDNKKFTLYDRYIYGYDNNYVNFYEEKNEKEKEYEEEDDKASSLCENKIILSQ
                     INCESFENDFKYYLSDYNYAFSIIDNTTNVLVAFGLYR"
     enhancer        1076..1379
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1381..1583
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1534..1554
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1786..1808
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1870..1889
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     primer_bind     1931..1950
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1931..1949
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1950..1974
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        1990..2099
                     /label=p10 promoter
                     /note="baculovirus promoter for expression in insect cells"
     CDS             2120..2137
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             2147..2176
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     primer_bind     3541..3562
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             join(3620..3622,3623..3625,3626..4138)
                     /codon_start=1
                     /product="N-terminal fragment of mVenus for use in 
                     bimolecular fluorescence complementation (BiFC) (Kodama and
                     Hu, 2010)"
                     /label=VN173
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KLICTTGKLPVPWPTLVTTLGYGLMCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK
                     ANFKIRHNIE"
     primer_bind     complement(3665..3686)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3926..3945)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     CDS             4790..4813
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     primer_bind     complement(4903..4922)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    4968..5023
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(5022..5041)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     terminator      5111..5158
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     misc_recomb     5171..5876
                     /label=baculovirus recombination region (ORF1629)
                     /note="contains part of ORF1629"
     3'UTR           5259..5631
                     /label=polyhedrin 3' UTR
                     /note="3' UTR from the Autographa californica 
                     nucleopolyhedrovirus polyhedrin gene (Suzuki et al., 2006)"
     CDS             complement(5288..5876)
                     /codon_start=2
                     /product="baculovirus capsid-associated protein"
                     /label=ORF1629
                     /note="required for viral replication"
                     /translation="IPKSSTTNLIADVLADTINRRRVAMAKSSSEATSNDEGWDDDDNR
                     PNKANTPDVKYVQALFNVFTSSQLYTNDSDERNTKAHNILNDVEPLLQNKTQTNIDKAR
                     LLLQDLASFVALSENPLDSPAIGSEKQPLFETNRNLFYKSIEDLIFKFRYKDAENHLIF
                     ALTYHPKDYKFNELLKYVQQLSVNQQRTESSA"
     protein_bind    complement(5885..5918)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     rep_origin      complement(6086..6673)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6165..6184)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6686..7477,7478..7546))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     7309..7328
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 tcctgcatct tttaatcaaa tcccaagatg tgtataaacg cgccggtatg tacaggaaga
       61 ggtttatact aaactgttac attgcaaacg tggtttcgtg tgccaagtgt gaaaaccgat
      121 gtttaatcaa ggctctgacg catttctaca accacgactc caagtgtgtg ggtgaagtca
      181 tgcatctttt aatcaaatcc caagatgtgt ataaaccacc aaactgccaa aaaatgaaaa
      241 ctgtcgacaa gctctgtccg tttgctggca actgcaaggg tctcaatcct atttgtaatt
      301 attgaataat aaaacaatta taaatgtcaa atttgttttt tattaacgat acaaaccaaa
      361 cgcaacaaga acatttgtag tattatctat aattgaaaac gcgtagttat aatcgctgag
      421 gtaatattta aaatcatttt caaatgattc acagttaatt tgcgacaata taattttatt
      481 ttcacataaa ctagacgcct tgtcgtcttc ttcttcgtat tccttctctt tttcattttt
      541 ctcttcataa aaattaacat agttattatc gtatccatat atgtatctat cgtatagagt
      601 aaattttttg ttgtcataaa tatatatgtc ttttttaatg gggtgtatag taccgctgcg
      661 catagttttt ctgtaattta caacagtgct attttctggt agttcttcgg agtgtgttgc
      721 tttaattatt aaatttatat aatcaatgaa tttgggatcg tcggttttgt acaatatgtt
      781 gccggcatag tacgcagctt cttctagttc aattacacca ttttttagca gcaccggatt
      841 aacataactt tccaaaatgt tgtacgaacc gttaaacaaa aacagttcac ctcccttttc
      901 tatactattg tctgcgagca gttgtttgtt gttaaaaata acagccattg taatgagacg
      961 cacaaactaa tatcacaaac tggaaatgtc tatcaatata tagttgctga tggccggcct
     1021 attaatagta atcaattacg gggtcattag ttcatagccc atatatggag ttccgcgtta
     1081 cataacttac ggtaaatggc ccgcctggct gaccgcccaa cgacccccgc ccattgacgt
     1141 caataatgac gtatgttccc atagtaacgc caatagggac tttccattga cgtcaatggg
     1201 tggagtattt acggtaaact gcccacttgg cagtacatca agtgtatcat atgccaagtc
     1261 cgccccctat tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc cagtacatga
     1321 ccttacggga ctttcctact tggcagtaca tctacgtatt agtcatcgct attaccatgc
     1381 tgatgcggtt ttggcagtac accaatgggc gtggatagcg gtttgactca cggggatttc
     1441 caagtctcca ccccattgac gtcaatggga gtttgttttg gcaccaaaat caacgggact
     1501 ttccaaaatg tcgtaataac cccgccccgt tgacgcaaat gggcggtagg cgtgtacggt
     1561 gggaggtcta tataagcaga cgtcgtttag tgaaccgtca gatcactaga tgctttattg
     1621 cggtagttta tcacagttaa attgctaacg ccagtctcga acttaacgtg cagaagttgg
     1681 tcgtgaggca ctgggcaggt aagtatcggg ccctttgtgc ggggggagcg gctcggggct
     1741 gtccgcgggg ggacggctgc cttcgggggg gacggggcag ggcggggttc ggcttctggc
     1801 gtgtgaccgg cggctctaga gcctctgcta accatgttca tgccttcttc tttttcctac
     1861 agctcctggg caacgtgctg gttattgtgc tgtctcatca ttttggcaaa gaattggatc
     1921 ggaccgaaat taatacgact cactataggg gaattgtgag cggataacaa ttccccggag
     1981 ttaatccggg acctttaatt caacccaaca caatatatta tagttaaata agaattatta
     2041 tcaaatcatt tgtatattaa ttaaaatact atactgtaaa ttacatttta tttacaatca
     2101 aaggagatat accatggcac accatcacca ccatcacggg agcggggagc agaaactaat
     2161 atcagaggag gatcttggct caggatcctc caccgacggc atggacgagc tgtacaaggg
     2221 tgctagcgga ggaatggtga gcaagggcga ggagaccaca atgggcgtaa tcaagcccga
     2281 catgaagatc aagctgaaga tggagggcaa cgtgaatggc cacgccttcg tgatcgaggg
     2341 cgagggcgag ggcaagccct acgacggcac caacaccatc aacctggagg tgaaggaggg
     2401 agcccccctg cccttctcct acgacattct gaccaccgcg ttcgcctacg gcaacagggc
     2461 cttcaccaag taccccgacg acatccccaa ctacttcaag cagtccttcc ccgagggcta
     2521 ctcttgggag cgcaccatga ccttcgagga caagggcatc gtgaaggtga agtccgacat
     2581 ctccatggag gaggactcct tcatctacga gatacacctc aagggcgaga acttcccccc
     2641 caacggcccc gtgatgcaga agaagaccac cggctgggac gcctccaccg agaggatgta
     2701 cgtgcgcgac ggcgtgctga agggcgacgt caagcacaag ctgctgctgg agggcggcgg
     2761 ccaccaccgc gttgacttca agaccatcta cagggccaag aaggcggtga agctgcccga
     2821 ctatcacttt gtggaccacc gcatcgagat cctgaaccac gacaaggact acaacaaggt
     2881 gaccgtttac gagagcgccg tggcccgcaa ctccggagac atccagaacc ctgacatcac
     2941 gagttcacga taccgtgggc tcccagcacc tggacctagc ccagctgata agaaacgctc
     3001 agggaagaag aagatcagca aagctgatat tggtgcaccc agtggattca agcatgtcag
     3061 ccacgtgggg tgggaccccc agaatggatt tgacgtgaac aacctcgacc ccgacctgcg
     3121 cagtctgttc tccagggcag gaatcagcga ggcccagctc accgacgccg agacctctaa
     3181 acttatctac gacttcattg aggaccaggg tgggctggag gctgtgcggc aggagatgag
     3241 gcgccaggag ccacttccgc ccgggacttc tggttctggt aaacctggtt ctggtgaagg
     3301 ttctactaaa gggacttctg gttctggtaa acctggttct ggtgaaggtt ctactaaagg
     3361 gacttctggt tctggtaaac ctggttctgg tgaaggttct actaaaggga cttctggttc
     3421 tggtaaacct ggttctggtg aaggttctac taaaggggcg gccgccatgc tgctgcccga
     3481 caaccactac ctgagctacc agtccgccct gagcaaagac cccaacgaga agcgcgatca
     3541 catggtcctg ctggagttcg tgaccgccgc cgggatcact ctcggcatgg acgagctgta
     3601 caagggtggc agcggtggca tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc
     3661 catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg
     3721 cgagggcgat gccacctacg gcaagctgac cctgaagctg atctgcacca ccggcaagct
     3781 gcccgtgccc tggcccaccc tcgtgaccac cctgggctac ggcctgatgt gcttcgcccg
     3841 ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacgt
     3901 ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg ccgaggtgaa
     3961 gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact tcaaggagga
     4021 cggcaacatc ctggggcaca agctggagta caactacaac agccacaacg tctatatcac
     4081 cgccgacaag cagaagaacg gcatcaaggc caacttcaag atccgccaca acatcgagga
     4141 cggcggcgtg cagctcgccg accactacca gcagaacacc cccatcggcg acggccccgt
     4201 gggtaccatg cagacaatta agtgtgttgt tgttggtgat ggtgctgttg gtaaaacatg
     4261 tctcctgata tcctacacaa caaacaaatt cccatcggaa tatgtaccaa ctgtttttga
     4321 caactatgca gtcacagtta tgattggtgg agagccatac actcttggac tttttgatac
     4381 tgcagggcaa gaggattatg acagactacg accgctaagt tatccacaga cagatgtttt
     4441 tctagtatgt ttctcagtgg tctctccatc ctcatttgaa aatgtgaaag aaaagtgggt
     4501 gcctgagata actcaccact gtccaaagac tcctttcttg cttgttggga cccaaattga
     4561 tctcagagat gacccctcta ctattgagaa acttgccaag aacaaacaga agcctattac
     4621 tccagagact gctgaaaagc tggcgcggga tctgaaggct gtcaagtatg tggagtgctc
     4681 tgccctcaca cagaaaggcc taaagaatgt gtttgatgaa gcaatattgg ctgccctgga
     4741 gcctccagaa ccgaagaaga gccgcaggtg tgtgctgcta tgactcgagc accaccatca
     4801 ccatcaccat cactaagtga ttaacctcag gtgcaggctg cctatcagaa ggtggtggct
     4861 ggtgtggcca atgccctggc tcacaaatac cactgagatc gatctttttc cctctgccaa
     4921 aaattatggg gacatcatga agccccttga gcatctgact tctggctaat aaaggaaatt
     4981 tattttcatt gcaatagtgt gttggaattt tttgtgtctc tcactcggaa ggacatatgg
     5041 gagggcaaat catttaaaac atcagaatga gtatttggtt tagagtttgg caacatatgc
     5101 ccatatgtaa ctagcataac cccttggggc ctctaaacgg gtcttgaggg gttttttgct
     5161 gaaagcatgc ggaggaaatt ctccttgaag tttccctggt gttcaaagta aaggagtttg
     5221 caccagacgc acctctgttc actggtccgg cgtattaaaa cacgatacat tgttattagt
     5281 acatttatta agcgctagat tctgtgcgtt gttgatttac agacaattgt tgtacgtatt
     5341 ttaataattc attaaattta taatctttag ggtggtatgt tagagcgaaa atcaaatgat
     5401 tttcagcgtc tttatatctg aatttaaata ttaaatcctc aatagatttg taaaataggt
     5461 ttcgattagt ttcaaacaag ggttgttttt ccgaaccgat ggctggacta tctaatggat
     5521 tttcgctcaa cgccacaaaa cttgccaaat cttgtagcag caatctagct ttgtcgatat
     5581 tcgtttgtgt tttgttttgt aataaaggtt cgacgtcgtt caaaatatta tgcgcttttg
     5641 tatttctttc atcactgtcg ttagtgtaca attgactcga cgtaaacacg ttaaatagag
     5701 cttggacata tttaacatcg ggcgtgttag ctttattagg ccgattatcg tcgtcgtccc
     5761 aaccctcgtc gttagaagtt gcttccgaag acgattttgc catagccaca cgacgcctat
     5821 taattgtgtc ggctaacacg tccgcgatca aatttgtagt tgagcttttt ggaattgcga
     5881 tcgcataact tcgtatagca tacattatac gaagttataa gctcggaacg ctgcgctcgg
     5941 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag
     6001 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
     6061 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg ccccctgacg agcatcacaa
     6121 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt
     6181 tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct
     6241 gtccgccttt ctcccttcgg gaagcgtggc gctttctcaa tgctcacgct gtaggtatct
     6301 cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc
     6361 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt
     6421 atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc
     6481 tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat
     6541 ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa
     6601 acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa
     6661 aaaaggatct caagaagatc ctttgttacc aatgcttaat cagtgaggca cctatctcag
     6721 cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga
     6781 tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac
     6841 cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc
     6901 ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta
     6961 gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac
     7021 gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat
     7081 gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa
     7141 gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg
     7201 tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag
     7261 aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc
     7321 cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct
     7381 caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat
     7441 cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg
     7501 ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc
     7561 aatattattg aagcatttat cagggttatt gtctcatgtc cgcgcgtt
//
