LOCUS       sequence-204185-        4946 bp    DNA     circular SYN 15-APR-2025
DEFINITION  To measure Rho activity by binding and FRET analysis.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4946)
  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 4946)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: "10.1083/jcb"; 
            journalName: "J Cell Biol"; date: "2012-04-30- 30"; volume: "197"; 
            issue: "3"; pages: "439-55"
COMMENT     Sequence Label: Ypet-RBD
FEATURES             Location/Qualifiers
     source          1..4946
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      14..602
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     503..522
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        777..1080
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1081..1284
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1235..1255
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      1322..1331
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1328..1330,1331..1333,1334..2044)
                     /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(1634..1653)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     polyA_signal    2450..2571
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2487..2506)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2541..2560
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2578..3033)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2715..2736)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2927..2946
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3060..3164
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3166..3523
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3166..3357
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3189..3209)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3374..3509
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3558..4352
                     /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             3558..4352
                     /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(3612..3631)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4222..4241
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4540..4559)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4584..4631
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
ORIGIN
        1 tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa
       61 aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga
      121 aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt
      181 taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt
      241 taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat
      301 agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct
      361 tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca
      421 cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag
      481 agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc
      541 gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga
      601 aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca
      661 tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc atgcattagt
      721 tattaatagt aatcaattac ggggtcatta gttcatagcc catatatgga gttccgcgtt
      781 acataactta cggtaaatgg cccgcctggc tgaccgccca acgacccccg cccattgacg
      841 tcaataatga cgtatgttcc catagtaacg ccaataggga ctttccattg acgtcaatgg
      901 gtggagtatt tacggtaaac tgcccacttg gcagtacatc aagtgtatca tatgccaagt
      961 acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc ccagtacatg
     1021 accttatggg actttcctac ttggcagtac atctacgtat tagtcatcgc tattaccatg
     1081 gtgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt
     1141 ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac
     1201 tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg
     1261 tgggaggtct atataagcag agctggttta gtgaaccgtc agatccgcta gcgctaccgg
     1321 tgccaccatg gtgagcaaag gcgaagagct gttcaccggc gtggtgccca tcctggtgga
     1381 gctggacggc gacgtgaacg gccacaagtt cagcgtgagc ggcgagggcg agggcgacgc
     1441 cacctacggc aagctgaccc tgaagctgct gtgcaccacc ggcaagctgc ccgtgccctg
     1501 gcccaccctg gtgaccaccc tgggctacgg cgtgcagtgc ttcgcccggt accccgacca
     1561 catgaagcag cacgacttct tcaagagcgc catgcccgag ggctacgtgc aggagcggac
     1621 catcttcttc aaggacgacg gcaactacaa gacccgggcc gaggtgaagt tcgagggcga
     1681 caccctggtg aaccggatcg agctgaaggg catcgacttc aaggaggacg gcaacatcct
     1741 gggccacaag ctggagtaca actacaacag ccacaacgtg tacatcaccg ccgacaagca
     1801 gaagaacggc atcaaggcca acttcaagat ccggcacaac atcgaggacg gcggcgtgca
     1861 gctggccgac cactaccagc agaacacccc catcggcgac ggccccgtgc tgctgcccga
     1921 caaccactac ctgagctacc agagcgccct gttcaaggac cccaacgaga agcgggacca
     1981 catggtgctg ctggagttcc tgaccgccgc cggcatcacc gagggcatga acgagctcta
     2041 taaggctcga gctcaagctt cgaattctat cctggaggac ctcaatatgc tctacatccg
     2101 gcagatggca ctcagcctgg aggacacaga gctgcagagg aaactagatc atgagatccg
     2161 gatgagggat ggggcctgca agctgctggc agcctgctcc cagcgagagc aggctctgga
     2221 agccaccaag agcctgctgg tgtgcaacag ccgtattctc agctacatgg gtgagctgca
     2281 gcggcgaaag gaggcccagg tgctggagaa gacaggctga ggatccaccg gatctagata
     2341 actgatcata atcagccata ccacatttgt agaggtttta cttgctttaa aaaacctccc
     2401 acacctcccc ctgaacctga aacataaaat gaatgcaatt gttgttgtta acttgtttat
     2461 tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt
     2521 tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt aaggcgtaaa
     2581 ttgtaagcgt taatattttg ttaaaattcg cgttaaattt ttgttaaatc agctcatttt
     2641 ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag accgagatag
     2701 ggttgagtgt tgttccagtt tggaacaaga gtccactatt aaagaacgtg gactccaacg
     2761 tcaaagggcg aaaaaccgtc tatcagggcg atggcccact acgtgaacca tcaccctaat
     2821 caagtttttt ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa gggagccccc
     2881 gatttagagc ttgacgggga aagccggcga acgtggcgag aaaggaaggg aagaaagcga
     2941 aaggagcggg cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta accaccacac
     3001 ccgccgcgct taatgcgccg ctacagggcg cgtcaggtgg cacttttcgg ggaaatgtgc
     3061 gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac
     3121 aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtcctgag gcggaaagaa
     3181 ccagctgtgg aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc cagcaggcag
     3241 aagtatgcaa agcatgcatc tcaattagtc agcaaccagg tgtggaaagt ccccaggctc
     3301 cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca tagtcccgcc
     3361 cctaactccg cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg
     3421 ctgactaatt ttttttattt atgcagaggc cgaggccgcc tcggcctctg agctattcca
     3481 gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaagatcgat caagagacag
     3541 gatgaggatc gtttcgcatg attgaacaag atggattgca cgcaggttct ccggccgctt
     3601 gggtggagag gctattcggc tatgactggg cacaacagac aatcggctgc tctgatgccg
     3661 ccgtgttccg gctgtcagcg caggggcgcc cggttctttt tgtcaagacc gacctgtccg
     3721 gtgccctgaa tgaactgcaa gacgaggcag cgcggctatc gtggctggcc acgacgggcg
     3781 ttccttgcgc agctgtgctc gacgttgtca ctgaagcggg aagggactgg ctgctattgg
     3841 gcgaagtgcc ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca
     3901 tcatggctga tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc ccattcgacc
     3961 accaagcgaa acatcgcatc gagcgagcac gtactcggat ggaagccggt cttgtcgatc
     4021 aggatgatct ggacgaagag catcaggggc tcgcgccagc cgaactgttc gccaggctca
     4081 aggcgagcat gcccgacggc gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga
     4141 atatcatggt ggaaaatggc cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg
     4201 cggaccgcta tcaggacata gcgttggcta cccgtgatat tgctgaagag cttggcggcg
     4261 aatgggctga ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg
     4321 ccttctatcg ccttcttgac gagttcttct gagcgggact ctggggttcg aaatgaccga
     4381 ccaagcgacg cccaacctgc catcacgaga tttcgattcc accgccgcct tctatgaaag
     4441 gttgggcttc ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct
     4501 catgctggag ttcttcgccc accctagggg gaggctaact gaaacacgga aggagacaat
     4561 accggaagga acccgcgcta tgacggcaat aaaaagacag aataaaacgc acggtgttgg
     4621 gtcgtttgtt cataaacgcg gggttcggtc ccagggctgg cactctgtcg ataccccacc
     4681 gagaccccat tggggccaat acgcccgcgt ttcttccttt tccccacccc accccccaag
     4741 ttcgggtgaa ggcccagggc tcgcagccaa cgtcggggcg gcaggccctg ccatagcctc
     4801 aggttactca tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta
     4861 ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca
     4921 ctgagcgtca gaccccgtag aaaaga
//
