LOCUS Exported 7038 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION ""Cdc42 was modified with a FRET 'binding antenna' that selectively reported Cdc42 binding to endogenous GDI..."" -Hodgson et al. ACCESSION . VERSION . KEYWORDS pTriEX-Antenna-GDI Cdc42 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7038) 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 7038) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7038 /organism="synthetic DNA construct" /mol_type="other DNA" CDS complement(345..1058) /codon_start=1 /product="enhanced monomeric variant of CFP (Rizzo et al., 2004)" /label=mCerulean /note=" mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNAISDNVYITADKQKNGIK ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL EFVTAAGITLGMDELY" CDS complement(1068..1085) /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" promoter complement(1106..1215) /label=p10 promoter /note="baculovirus promoter for expression in insect cells" protein_bind 1231..1255 /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)." primer_bind complement(1255..1274) /label=T7 /note="T7 promoter, forward primer" promoter complement(1256..1274) /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" primer_bind complement(1316..1335) /label=pCAG-F /note="Rabbit beta-globin intron, for pCAG plasmids, forward primer" primer_bind complement(1397..1419) /label=pCAGGS-5 /note="Chimeric intron in CAG promoter, forward primer" promoter complement(1625..1824) /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind complement(1651..1671) /label=CMV-F /note="Human CMV immediate early promoter, forward primer" enhancer 1826..2129 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" misc_recomb 2197..3024 /label=baculovirus recombination region (lef2/ORF603) /note="contains ORF603 and part of lef2" CDS 2257..2862 /codon_start=1 /product="baculovirus ORF603 protein" /label=ORF603 /translation="MAVIFNNKQLLADNSIEKGGELFLFNGSYNILESYVNPVLLKNGV IELEEAAYYAGNILYKTDDPKFIDYINLIIKATHSEELPENSTVVNYRKTMRSGTIHPI KKDIYIYDNKKFTLYDRYIYGYDNNYVNFYEEKNEKEKEYEEEDDKASSLCENKIILSQ INCESFENDFKYYLSDYNYAFSIIDNTTNVLVAFGLYR" CDS complement(2900..3024) /codon_start=3 /product="baculovirus late expression factor 2" /label=lef2 /translation="HLLIKSQDVYKPPNCQKMKTVDKLCPFAGNCKGLNPICNY" CDS 3267..4127 /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 complement(3485..3504) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 4140..4727 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 4629..4648 /label=pBR322ori-F /note="pBR322 origin, forward primer" protein_bind complement(4895..4928) /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." misc_recomb 4937..5642 /label=baculovirus recombination region (ORF1629) /note="contains part of ORF1629" CDS 4937..5525 /codon_start=2 /product="baculovirus capsid-associated protein" /label=ORF1629 /note="required for viral replication" /translation="IPKSSTTNLIADVLADTINRRRVAMAKSSSEATSNDEGWDDDDNR PNKANTPDVKYVQALFNVFTSSQLYTNDSDERNTKAHNILNDVEPLLQNKTQTNIDKAR LLLQDLASFVALSENPLDSPAIGSEKQPLFETNRNLFYKSIEDLIFKFRYKDAENHLIF ALTYHPKDYKFNELLKYVQQLSVNQQRTESSA" 3'UTR 5182..5554 /label=polyhedrin 3' UTR /note="3' UTR from the Autographa californica nucleopolyhedrovirus polyhedrin gene (Suzuki et al., 2006)" terminator 5655..5702 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" primer_bind 5772..5791 /label=rbglobpA-R /note="Rabbit beta-globin polyA, reverse primer. Also called rb-glob-pA-term-R" polyA_signal 5790..5845 /label=-beta-globin poly(A) signal /note="rabbit beta-globin polyadenylation signal (Gil and Proudfoot, 1987)" primer_bind 5891..5910 /label=Bglob-pA-R /note="Rabbit beta-globin polyA region, reverse primer" CDS complement(6000..6023) /codon_start=1 /product="8xHis affinity tag" /label=8xHis /translation="HHHHHHHH" CDS complement(6033..6608) /codon_start=1 /product="human Rho family small GTPase Cdc42" /label=Cdc42 /translation="MQTIKCVVVGDGAVGKTCLLISYTTNKFPSEYVPTVFDNYAVTVM IGGEPYTLGLFDTAGQEDYDRLRPLSYPQTDVFLVCFSVVSPSSFENVKEKWVPEITHH CPKTPFLLVGTQIDLRDDPSTIEKLAKNKQKPITPETAEKLARDLKAVKYVECSALTQK GLKNVFDEAILAALEPPEPKKSRRCVLL" CDS complement(6783..7034) /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" ORIGIN 1 gtgatataga cgttgtggct gttgtagttg tactccagct tgtgccccag gatgttgccg 61 tcctccttga agtcgatgcc cttcagctcg atgcggttca ccagggtgtc gccctcgaac 121 ttcacctcgg cgcgggtctt gtagttgccg tcgtccttga agaagatggt gcgctcctgg 181 acgtagcctt cgggcatggc ggacttgaag aagtcgtgct gcttcatgtg gtcggggtag 241 cgggcgaagc acatcaggcc gtagcccagg gtggtcacga gggtgggcca gggcacgggc 301 agcttgccgg tcattgcggc cgcaccagaa ccagataagc ttttgtacag ctcgtccatg 361 ccgagagtga tcccggcggc ggtcacgaac tccagcagga ccatgtgatc gcgcttctcg 421 ttggggtctt tgctcagctt ggactgggtg ctcaggtagt ggttgtcggg cagcagcacg 481 gggccgtcgc cgatgggggt gttctgctgg tagtggtcgg cgagctgcac gctgccgtcc 541 tcgatgttgt ggcggatctt gaagttggcc ttgatgccgt tcttctgctt gtcggcggtg 601 atatagacgt tgtcgctgat ggcgttgtac tccagcttgt gccccaggat gttgccgtcc 661 tccttgaagt cgatgccctt cagctcgatg cggttcacca gggtgtcgcc ctcgaacttc 721 acctcggcgc gggtcttgta gttgccgtcg tccttgaaga agatggtgcg ctcctggacg 781 tagccttcgg gcatggcgga cttgaagaag tcgtgctgct tcatgtggtc ggggtagcgg 841 gcgaagcact gcacgcccca ggtcagggtg gtcacgaggg tgggccaggg cacgggcagc 901 ttgccggtgg tgcagatgaa cttcagggtc agcttgccgt aggtggcatc gccctcgccc 961 tcgccggaca cgctgaactt gtggccgttt acgtcgccgt ccagctcgac caggatgggc 1021 accaccccgg tgaacagctc ctcgcccttg ctcaccattc cggatccgtg atggtggtga 1081 tggtgtgcca tggtatatct cctttgattg taaataaaat gtaatttaca gtatagtatt 1141 ttaattaata tacaaatgat ttgataataa ttcttattta actataatat attgtgttgg 1201 gttgaattaa aggtcccgga ttaactacgg ggaattgtta tccgctcaca attcccctat 1261 agtgagtcgt attaatttcg gtccgatcca attctttgcc aaaatgatga gacagcacaa 1321 taaccagcac gttgcccagg agctgtagga aaaagaagaa ggcatgaaca tggttagcag 1381 aggctctaga gccgccggtc acacgccaga agccgaaccc cgccctgccc cgtccccccc 1441 gaaggcagcc gtccccccgc ggacagcccc gagccgctcc ccccgcacaa agggcccgat 1501 acttacctgc ccagtgcctc acgaccaact tctgcacgtt aagttcgaga ctggcgttag 1561 caatttaact gtgataaact accgcaataa agcatctagt gatctgacgg ttcactaaac 1621 gacgtctgct tatatagacc tcccaccgta cacgcctacc gcccatttgc gtcaacgggg 1681 cggggttatt acgacatttt ggaaagtccc gttgattttg gtgccaaaac aaactcccat 1741 tgacgtcaat ggggtggaga cttggaaatc cccgtgagtc aaaccgctat ccacgcccat 1801 tggtgtactg ccaaaaccgc atcagcatgg taatagcgat gactaatacg tagatgtact 1861 gccaagtagg aaagtcccgt aaggtcatgt actgggcata atgccaggcg ggccatttac 1921 cgtcattgac gtcaataggg ggcggacttg gcatatgata cacttgatgt actgccaagt 1981 gggcagttta ccgtaaatac tccacccatt gacgtcaatg gaaagtccct attggcgtta 2041 ctatgggaac atacgtcatt attgacgtca atgggcgggg gtcgttgggc ggtcagccag 2101 gcgggccatt taccgtaagt tatgtaacgc ggaactccat atatgggcta tgaactaatg 2161 accccgtaat tgattactat taataggccg gccatcagca actatatatt gatagacatt 2221 tccagtttgt gatattagtt tgtgcgtctc attacaatgg ctgttatttt taacaacaaa 2281 caactgctcg cagacaatag tatagaaaag ggaggtgaac tgtttttgtt taacggttcg 2341 tacaacattt tggaaagtta tgttaatccg gtgctgctaa aaaatggtgt aattgaacta 2401 gaagaagctg cgtactatgc cggcaacata ttgtacaaaa ccgacgatcc caaattcatt 2461 gattatataa atttaataat taaagcaaca cactccgaag aactaccaga aaatagcact 2521 gttgtaaatt acagaaaaac tatgcgcagc ggtactatac accccattaa aaaagacata 2581 tatatttatg acaacaaaaa atttactcta tacgatagat acatatatgg atacgataat 2641 aactatgtta atttttatga agagaaaaat gaaaaagaga aggaatacga agaagaagac 2701 gacaaggcgt ctagtttatg tgaaaataaa attatattgt cgcaaattaa ctgtgaatca 2761 tttgaaaatg attttaaata ttacctcagc gattataact acgcgttttc aattatagat 2821 aatactacaa atgttcttgt tgcgtttggt ttgtatcgtt aataaaaaac aaatttgaca 2881 tttataattg ttttattatt caataattac aaataggatt gagacccttg cagttgccag 2941 caaacggaca gagcttgtcg acagttttca ttttttggca gtttggtggt ttatacacat 3001 cttgggattt gattaaaaga tgcatgactt cacccacaca cttggagtcg tggttgtaga 3061 aatgcgtcag agccttgatt aaacatcggt tttcacactt ggcacacgaa accacgtttg 3121 caatgtaaca gtttagtata aacctcttcc tgtacatacc ggcgcgttta tacacatctt 3181 gggatttgat taaaagatgc aggaaacgcg cggacatgag acaataaccc tgataaatgc 3241 ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc 3301 ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa 3361 aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg 3421 gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag 3481 ttctgctatg tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc 3541 gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta 3601 cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg 3661 cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca 3721 acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac 3781 caaacgacga gcgtgacacc acgatgcctg tagcaatggc aacaacgttg cgcaaactat 3841 taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg 3901 ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata 3961 aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta 4021 agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa 4081 atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaacaa aggatcttct 4141 tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca 4201 gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc 4261 agcagagcgc agataccaaa tactgttctt ctagtgtagc cgtagttagg ccaccacttc 4321 aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct 4381 gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag 4441 gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc 4501 tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg 4561 agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag 4621 cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt 4681 gagcgtcgat ttttgtgatg ctcgtcaggg ggcggagcct atggaaaaac gccagcaacg 4741 cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt 4801 tatcccctga ttctgtggat aaccgtatta ccgcctttga gtgagctgat accgctcgcc 4861 gcagccgaac gaccgagcgc agcgttccga gcttataact tcgtataatg tatgctatac 4921 gaagttatgc gatcgcaatt ccaaaaagct caactacaaa tttgatcgcg gacgtgttag 4981 ccgacacaat taataggcgt cgtgtggcta tggcaaaatc gtcttcggaa gcaacttcta 5041 acgacgaggg ttgggacgac gacgataatc ggcctaataa agctaacacg cccgatgtta 5101 aatatgtcca agctctattt aacgtgttta cgtcgagtca attgtacact aacgacagtg 5161 atgaaagaaa tacaaaagcg cataatattt tgaacgacgt cgaaccttta ttacaaaaca 5221 aaacacaaac gaatatcgac aaagctagat tgctgctaca agatttggca agttttgtgg 5281 cgttgagcga aaatccatta gatagtccag ccatcggttc ggaaaaacaa cccttgtttg 5341 aaactaatcg aaacctattt tacaaatcta ttgaggattt aatatttaaa ttcagatata 5401 aagacgctga aaatcatttg attttcgctc taacatacca ccctaaagat tataaattta 5461 atgaattatt aaaatacgta caacaattgt ctgtaaatca acaacgcaca gaatctagcg 5521 cttaataaat gtactaataa caatgtatcg tgttttaata cgccggacca gtgaacagag 5581 gtgcgtctgg tgcaaactcc tttactttga acaccaggga aacttcaagg agaatttcct 5641 ccgcatgctt tcagcaaaaa acccctcaag acccgtttag aggccccaag gggttatgct 5701 agttacatat gggcatatgt tgccaaactc taaaccaaat actcattctg atgttttaaa 5761 tgatttgccc tcccatatgt ccttccgagt gagagacaca aaaaattcca acacactatt 5821 gcaatgaaaa taaatttcct ttattagcca gaagtcagat gctcaagggg cttcatgatg 5881 tccccataat ttttggcaga gggaaaaaga tcgatctcag tggtatttgt gagccagggc 5941 attggccaca ccagccacca ccttctgata ggcagcctgc acctgaggtt aatcacttag 6001 tgatggtgat ggtgatggtg gtgctcgagt tatcatagca gcacacacct gcggctcttc 6061 ttcggttctg gaggctccag ggcagccaat attgcttcgt caaatacatt ctttaggcct 6121 ttctgtgtaa gtgcagaaca ctccacatac ttgacagcct tcaggtcacg ggccagcttt 6181 tcagcagtct ctggagtgat aggcttctgt ttgttcttgg caagtttctc aatagtagag 6241 gggtcatctc tgagatcaat ttgagtccca acaagcaaga aaggagtctt tggacagtgg 6301 tgagttatct caggcaccca cttttctttc acgttttcaa atgaagatgg agagaccact 6361 gaaaaacaga ctagaaatac atctgtttgt ggataactca gcggtcgtaa tctgtcataa 6421 tcctcttgcc ctgcagtatc aaaaagtcca agagtatatg gttctccacc aatcataact 6481 gtgactgcat agttgtcaaa aacagtcggt acatattccg atggaaattt gtttgttgtg 6541 taggatatca ggagacatgt tttaccaaca gcaccatcgc ccacaacaac acacttaatt 6601 gtctgcatga attccttggt gcagatcaac ttcagggtca gcttgccgta ggtggcatcg 6661 ccctcgccct cgccggacac gctgaacttg tggccgttta cgtcgccgtc cagctcgacc 6721 aggatgggca ccaccccggt gaacagctcc tcgcccttgc tcaccatgcc accgctgcca 6781 cccttgtaca gctcgtccat gccgagagtg atcccggcgg cggtcacgaa ctccagcagg 6841 accatgtgat cgcgcttctc gttggggtct ttgctcaggg cggactggta gctcaggtag 6901 tggttgtcgg gcagcagcac ggggccgtcg ccgatggggg tgttctgctg gtagtggtcg 6961 gcgagctgca cgccgccgtc ctcgatgttg tggcggatct tgaagttggc cttgatgccg 7021 ttcttctgct tgtcggcg //