LOCUS       sequence-372008-        7044 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of Antenna-RhoA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7044)
  AUTHORS   Hodgson L, Spiering D, Sabouri-Ghomi M, Dagliyan O, DerMardirossian 
            C, Danuser G, Hahn KM
  TITLE     FRET binding antenna reports spatiotemporal dynamics of GDI-Cdc42 
            GTPase interactions.
  JOURNAL   Nat Chem Biol. 2016 Aug 8. doi: 10.1038/nchembio.2145.
  PUBMED    27501396
REFERENCE   2  (bases 1 to 7044)
  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: "Nat Chem 
            Biol. 2016 Aug 8. doi: 10.1038/nchembio.2145."
COMMENT     Sequence Label: pTriEX-Antenna-GDI RhoA
FEATURES             Location/Qualifiers
     source          1..7044
                     /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             join(2147..2149,2150..2152,2153..2863)
                     /codon_start=1
                     /product="enhanced monomeric variant of CFP (Rizzo et al., 
                     2004)"
                     /label=mCerulean
                     /note=" mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNAISDNVYITADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             3209..3460
                     /codon_start=1
                     /product="C-terminal fragment of mVenus for use in 
                     bimolecular fluorescence complementation (BiFC) (Kodama and
                     Hu, 2010)"
                     /label=VC155
                     /translation="DKQKNGIKANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNH
                     YLSYQSALSKDPNEKRDHMVLLEFVTAAGITLGMDELYK"
     CDS             3635..4213
                     /codon_start=1
                     /product="human Rho family small GTPase RhoA"
                     /label=RhoA
                     /translation="AAIRKKLVIVGDGACGKTCLLIVFSKDQFPEVYVPTVFENYVADI
                     EVDGKQVELALWDTAGQEDYDRLRPLSYPDTDVILMCFSIDSPDSLENIPEKWTPEVKH
                     FCPNVPIILVGNKKDLRNDEHTRRELAKMKQEPVKPEEGRDMANRIGAFGYMECSAKTK
                     DGVREVFEMATRAALQARRGKKKSGCLVL"
     CDS             4226..4249
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     primer_bind     complement(4339..4358)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    4404..4459
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(4458..4477)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     terminator      4547..4594
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     misc_recomb     4607..5312
                     /label=baculovirus recombination region (ORF1629)
                     /note="contains part of ORF1629"
     3'UTR           4695..5067
                     /label=polyhedrin 3' UTR
                     /note="3' UTR from the Autographa californica 
                     nucleopolyhedrovirus polyhedrin gene (Suzuki et al., 2006)"
     CDS             complement(4724..5312)
                     /codon_start=2
                     /product="baculovirus capsid-associated protein"
                     /label=ORF1629
                     /note="required for viral replication"
                     /translation="IPKSSTTNLIADVLADTINRRRVAMAKSSSEATSNDEGWDDDDNR
                     PNKANTPDVKYVQALFNVFTSSQLYTNDSDERNTKAHNILNDVEPLLQNKTQTNIDKAR
                     LLLQDLASFVALSENPLDSPAIGSEKQPLFETNRNLFYKSIEDLIFKFRYKDAENHLIF
                     ALTYHPKDYKFNELLKYVQQLSVNQQRTESSA"
     protein_bind    complement(5321..5354)
                     /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(5522..6109)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5601..5620)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6122..6913,6914..6982))
                     /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     6745..6764
                     /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 ttccccgtag
     1981 ttaatccggg acctttaatt caacccaaca caatatatta tagttaaata agaattatta
     2041 tcaaatcatt tgtatattaa ttaaaatact atactgtaaa ttacatttta tttacaatca
     2101 aaggagatat accatggcac accatcacca ccatcacgga tccggaatgg tgagcaaggg
     2161 cgaggagctg ttcaccgggg tggtgcccat cctggtcgag ctggacggcg acgtaaacgg
     2221 ccacaagttc agcgtgtccg gcgagggcga gggcgatgcc acctacggca agctgaccct
     2281 gaagttcatc tgcaccaccg gcaagctgcc cgtgccctgg cccaccctcg tgaccaccct
     2341 gacctggggc gtgcagtgct tcgcccgcta ccccgaccac atgaagcagc acgacttctt
     2401 caagtccgcc atgcccgaag gctacgtcca ggagcgcacc atcttcttca aggacgacgg
     2461 caactacaag acccgcgccg aggtgaagtt cgagggcgac accctggtga accgcatcga
     2521 gctgaagggc atcgacttca aggaggacgg caacatcctg gggcacaagc tggagtacaa
     2581 cgccatcagc gacaacgtct atatcaccgc cgacaagcag aagaacggca tcaaggccaa
     2641 cttcaagatc cgccacaaca tcgaggacgg cagcgtgcag ctcgccgacc actaccagca
     2701 gaacaccccc atcggcgacg gccccgtgct gctgcccgac aaccactacc tgagcaccca
     2761 gtccaagctg agcaaagacc ccaacgagaa gcgcgatcac atggtcctgc tggagttcgt
     2821 gaccgccgcc gggatcactc tcggcatgga cgagctgtac aaaagcttat ctggttctgg
     2881 tgcggccgca atgaccggca agctgcccgt gccctggccc accctcgtga ccaccctggg
     2941 ctacggcctg atgtgcttcg cccgctaccc cgaccacatg aagcagcacg acttcttcaa
     3001 gtccgccatg cccgaaggct acgtccagga gcgcaccatc ttcttcaagg acgacggcaa
     3061 ctacaagacc cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc gcatcgagct
     3121 gaagggcatc gacttcaagg aggacggcaa catcctgggg cacaagctgg agtacaacta
     3181 caacagccac aacgtctata tcaccgccga caagcagaag aacggcatca aggccaactt
     3241 caagatccgc cacaacatcg aggacggcgg cgtgcagctc gccgaccact accagcagaa
     3301 cacccccatc ggcgacggcc ccgtgctgct gcccgacaac cactacctga gctaccagtc
     3361 cgccctgagc aaagacccca acgagaagcg cgatcacatg gtcctgctgg agttcgtgac
     3421 cgccgccggg atcactctcg gcatggacga gctgtacaag ggtggcagcg gtggcatggt
     3481 gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc tggacggcga
     3541 cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa
     3601 gctgaccctg aagttgatct gcaccaagga attcgctgcc atccggaaga aactggtgat
     3661 tgttggtgat ggagcctgtg gaaagacatg cttgctcata gtcttcagca aggaccagtt
     3721 cccagaggtg tatgtgccca cagtgtttga gaactatgtg gcagatatcg aggtggatgg
     3781 aaagcaggta gagttggctt tgtgggacac agctgggcag gaagattatg atcgcctgag
     3841 gcccctctcc tacccagata ccgatgttat actgatgtgt ttttccatcg acagccctga
     3901 tagtttagaa aacatcccag aaaagtggac cccagaagtc aagcatttct gtcccaacgt
     3961 gcccatcatc ctggttggga ataagaagga tcttcggaat gatgagcaca caaggcggga
     4021 gctagccaag atgaagcagg agccggtgaa acctgaagaa ggcagagata tggcaaacag
     4081 gattggcgct tttgggtaca tggagtgttc agcaaagacc aaagatggag tgagagaggt
     4141 ttttgaaatg gctacgagag ctgctctgca agctagacgt gggaagaaaa aatctgggtg
     4201 ccttgtcttg tgaaactaac tcgagcacca ccatcaccat caccatcact aagtgattaa
     4261 cctcaggtgc aggctgccta tcagaaggtg gtggctggtg tggccaatgc cctggctcac
     4321 aaataccact gagatcgatc tttttccctc tgccaaaaat tatggggaca tcatgaagcc
     4381 ccttgagcat ctgacttctg gctaataaag gaaatttatt ttcattgcaa tagtgtgttg
     4441 gaattttttg tgtctctcac tcggaaggac atatgggagg gcaaatcatt taaaacatca
     4501 gaatgagtat ttggtttaga gtttggcaac atatgcccat atgtaactag cataacccct
     4561 tggggcctct aaacgggtct tgaggggttt tttgctgaaa gcatgcggag gaaattctcc
     4621 ttgaagtttc cctggtgttc aaagtaaagg agtttgcacc agacgcacct ctgttcactg
     4681 gtccggcgta ttaaaacacg atacattgtt attagtacat ttattaagcg ctagattctg
     4741 tgcgttgttg atttacagac aattgttgta cgtattttaa taattcatta aatttataat
     4801 ctttagggtg gtatgttaga gcgaaaatca aatgattttc agcgtcttta tatctgaatt
     4861 taaatattaa atcctcaata gatttgtaaa ataggtttcg attagtttca aacaagggtt
     4921 gtttttccga accgatggct ggactatcta atggattttc gctcaacgcc acaaaacttg
     4981 ccaaatcttg tagcagcaat ctagctttgt cgatattcgt ttgtgttttg ttttgtaata
     5041 aaggttcgac gtcgttcaaa atattatgcg cttttgtatt tctttcatca ctgtcgttag
     5101 tgtacaattg actcgacgta aacacgttaa atagagcttg gacatattta acatcgggcg
     5161 tgttagcttt attaggccga ttatcgtcgt cgtcccaacc ctcgtcgtta gaagttgctt
     5221 ccgaagacga ttttgccata gccacacgac gcctattaat tgtgtcggct aacacgtccg
     5281 cgatcaaatt tgtagttgag ctttttggaa ttgcgatcgc ataacttcgt atagcataca
     5341 ttatacgaag ttataagctc ggaacgctgc gctcggtcgt tcggctgcgg cgagcggtat
     5401 cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga
     5461 acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt
     5521 ttttccatag gctccgcccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
     5581 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg
     5641 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
     5701 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
     5761 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
     5821 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
     5881 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc
     5941 taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac
     6001 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg
     6061 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt
     6121 gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca
     6181 tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc
     6241 ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa
     6301 accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc
     6361 agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca
     6421 acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat
     6481 tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag
     6541 cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac
     6601 tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt
     6661 ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt
     6721 gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc
     6781 tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat
     6841 ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca
     6901 gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga
     6961 cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg
     7021 gttattgtct catgtccgcg cgtt
//
