LOCUS Exported 4952 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION To measure Rac or Cdc42 activity by binding and FRET analysis. ACCESSION . VERSION . KEYWORDS Ypet-PBD SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 4952) AUTHORS Petrie RJ, Gavara N, Chadwick RS, Yamada KM TITLE Nonpolarized signaling reveals two distinct modes of 3D cell migration. JOURNAL J Cell Biol. 2012 Apr 30;197(3):439-55. doi: 10.1083/jcb.201201124. PUBMED 22547408 REFERENCE 2 (bases 1 to 4952) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..4952 /organism="synthetic DNA construct" /mol_type="other DNA" rep_origin 10..598 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 499..518 /label=pBR322ori-F /note="pBR322 origin, forward primer" enhancer 773..1076 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 1077..1280 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 1231..1251 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" regulatory 1318..1327 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 1324..2040 /codon_start=1 /product="yellow fluorescent protein optimized for FRET with CyPet (Nguyen and Daugherty, 2005)" /label=YPet /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KLLCTTGKLPVPWPTLVTTLGYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALFKDPNEKRDHMVLL EFLTAAGITEGMNELYK" primer_bind complement(1630..1649) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" polyA_signal 2452..2573 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(2489..2508) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 2543..2562 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" rep_origin complement(2580..3035) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(2717..2738) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 2929..2948 /label=F1ori-R /note="F1 origin, reverse primer" promoter 3062..3166 /gene="bla" /label=AmpR promoter promoter 3168..3525 /label=SV40 promoter /note="SV40 enhancer and early promoter" primer_bind complement(3191..3211) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" rep_origin 3376..3511 /label=SV40 ori /note="SV40 origin of replication" primer_bind 3438..3457 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" CDS 3560..4354 /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(3614..3633) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 4224..4243 /label=Neo-F /note="Neomycin resistance gene, forward primer" primer_bind complement(4542..4561) /label=TK-pA-R /note="Thymidine kinase polyA, reverse primer" polyA_signal 4586..4633 /label=HSV TK poly(A) signal /note="herpes simplex virus thymidine kinase polyadenylation signal (Cole and Stacy, 1985)" ORIGIN 1 aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc 61 accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt 121 aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc cgtagttagg 181 ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc 241 agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt 301 accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga 361 gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct 421 tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg 481 cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca 541 cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa 601 cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt 661 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgccatgc attagttatt 721 aatagtaatc aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat 781 aacttacggt aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa 841 taatgacgta tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg 901 agtatttacg gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc 961 cccctattga cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct 1021 tatgggactt tcctacttgg cagtacatct acgtattagt catcgctatt accatggtga 1081 tgcggttttg gcagtacatc aatgggcgtg gatagcggtt tgactcacgg ggatttccaa 1141 gtctccaccc cattgacgtc aatgggagtt tgttttggca ccaaaatcaa cgggactttc 1201 caaaatgtcg taacaactcc gccccattga cgcaaatggg cggtaggcgt gtacggtggg 1261 aggtctatat aagcagagct ggtttagtga accgtcagat ccgctagcgc taccggtgcc 1321 accatggtga gcaaaggcga agagctgttc accggcgtgg tgcccatcct ggtggagctg 1381 gacggcgacg tgaacggcca caagttcagc gtgagcggcg agggcgaggg cgacgccacc 1441 tacggcaagc tgaccctgaa gctgctgtgc accaccggca agctgcccgt gccctggccc 1501 accctggtga ccaccctggg ctacggcgtg cagtgcttcg cccggtaccc cgaccacatg 1561 aagcagcacg acttcttcaa gagcgccatg cccgagggct acgtgcagga gcggaccatc 1621 ttcttcaagg acgacggcaa ctacaagacc cgggccgagg tgaagttcga gggcgacacc 1681 ctggtgaacc ggatcgagct gaagggcatc gacttcaagg aggacggcaa catcctgggc 1741 cacaagctgg agtacaacta caacagccac aacgtgtaca tcaccgccga caagcagaag 1801 aacggcatca aggccaactt caagatccgg cacaacatcg aggacggcgg cgtgcagctg 1861 gccgaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct gcccgacaac 1921 cactacctga gctaccagag cgccctgttc aaggacccca acgagaagcg ggaccacatg 1981 gtgctgctgg agttcctgac cgccgccggc atcaccgagg gcatgaacga gctctataag 2041 gctcgagctc aagcttcgaa ttccaataaa aagaaagaga aagagcggcc agagatttct 2101 ctcccttcag attttgaaca cacaattcat gtcggttttg atgctgtcac aggggagttt 2161 acgggaatgc cagagcagtg ggcccgcttg cttcagacat caaatatcac taagtcggag 2221 cagaagaaaa acccgcaggc tgttctggat gtgttggagt tttacaactc gaagaagaca 2281 tccaacagcc agaaatacat gagctttaca gataagtcag ctggatccac cggatctaga 2341 taactgatca taatcagcca taccacattt gtagaggttt tacttgcttt aaaaaacctc 2401 ccacacctcc ccctgaacct gaaacataaa atgaatgcaa ttgttgttgt taacttgttt 2461 attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca 2521 tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttaaggcgta 2581 aattgtaagc gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt 2641 ttttaaccaa taggccgaaa tcggcaaaat cccttataaa tcaaaagaat agaccgagat 2701 agggttgagt gttgttccag tttggaacaa gagtccacta ttaaagaacg tggactccaa 2761 cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac catcacccta 2821 atcaagtttt ttggggtcga ggtgccgtaa agcactaaat cggaacccta aagggagccc 2881 ccgatttaga gcttgacggg gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc 2941 gaaaggagcg ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg taaccaccac 3001 acccgccgcg cttaatgcgc cgctacaggg cgcgtcaggt ggcacttttc ggggaaatgt 3061 gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag 3121 acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtcctg aggcggaaag 3181 aaccagctgt ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc 3241 agaagtatgc aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc 3301 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg 3361 cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat 3421 ggctgactaa ttttttttat ttatgcagag gccgaggccg cctcggcctc tgagctattc 3481 cagaagtagt gaggaggctt ttttggaggc ctaggctttt gcaaagatcg atcaagagac 3541 aggatgagga tcgtttcgca tgattgaaca agatggattg cacgcaggtt ctccggccgc 3601 ttgggtggag aggctattcg gctatgactg ggcacaacag acaatcggct gctctgatgc 3661 cgccgtgttc cggctgtcag cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc 3721 cggtgccctg aatgaactgc aagacgaggc agcgcggcta tcgtggctgg ccacgacggg 3781 cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg ggaagggact ggctgctatt 3841 gggcgaagtg ccggggcagg atctcctgtc atctcacctt gctcctgccg agaaagtatc 3901 catcatggct gatgcaatgc ggcggctgca tacgcttgat ccggctacct gcccattcga 3961 ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg atggaagccg gtcttgtcga 4021 tcaggatgat ctggacgaag agcatcaggg gctcgcgcca gccgaactgt tcgccaggct 4081 caaggcgagc atgcccgacg gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc 4141 gaatatcatg gtggaaaatg gccgcttttc tggattcatc gactgtggcc ggctgggtgt 4201 ggcggaccgc tatcaggaca tagcgttggc tacccgtgat attgctgaag agcttggcgg 4261 cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat 4321 cgccttctat cgccttcttg acgagttctt ctgagcggga ctctggggtt cgaaatgacc 4381 gaccaagcga cgcccaacct gccatcacga gatttcgatt ccaccgccgc cttctatgaa 4441 aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat 4501 ctcatgctgg agttcttcgc ccaccctagg gggaggctaa ctgaaacacg gaaggagaca 4561 ataccggaag gaacccgcgc tatgacggca ataaaaagac agaataaaac gcacggtgtt 4621 gggtcgtttg ttcataaacg cggggttcgg tcccagggct ggcactctgt cgatacccca 4681 ccgagacccc attggggcca atacgcccgc gtttcttcct tttccccacc ccacccccca 4741 agttcgggtg aaggcccagg gctcgcagcc aacgtcgggg cggcaggccc tgccatagcc 4801 tcaggttact catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc 4861 taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc 4921 cactgagcgt cagaccccgt agaaaagatc aa //