LOCUS       sequence-197846-        6472 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Used as a RhoA-GTP sensor.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6472)
  AUTHORS   O'Neill PR, Castillo-Badillo JA, Meshik X, Kalyanaraman V, Melgarejo
            K, Gautam N
  TITLE     Membrane Flow Drives an Adhesion-Independent Amoeboid Cell Migration
            Mode.
  JOURNAL   Dev Cell. 2018 Jun 15. pii: S1534-5807(18)30422-2. doi: 
            10.1016/j.devcel.2018.05.029.
  PUBMED    29937389
REFERENCE   2  (bases 1 to 6472)
  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: "Dev Cell. 
            2018 Jun 15. pii: S1534-5807(18)30422-2. doi: 
            10.1016/j.devcel.2018.05.029."
COMMENT     Sequence Label: Venus rGBD
FEATURES             Location/Qualifiers
     source          1..6472
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    1..122
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(38..57)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     92..111
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(171..187)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(171..187)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(184..206)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    195..211
                     /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        complement(join(219..225,226..243,244..249))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    264..285
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(402..419)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(573..1161)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(653..672)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1332..2123,2124..2192))
                     /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     1955..1974
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2193..2297)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(2372..2391)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        2563..2942
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2943..3146
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     3097..3117
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     3191..3210
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3191..3209
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(3245..3247,3248..3250,3251..3961)
                     /codon_start=1
                     /product="Venus YFP with monomerizing A206K mutation (Nagai
                     et al., 2002; Kremers et al., 2006)"
                     /label=mVenus
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KLICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK
                     ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3290..3311)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3551..3570)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3898..3919
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(4391..4408)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4397..4621
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4667..5095
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4754..4773)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4964..4985
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5104..5124)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5109..5438
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5289..5424
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5351..5370
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             5505..6299
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(5559..5578)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6169..6188
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
ORIGIN
        1 aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca
       61 aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct
      121 tatcatgtct gtataccgtc gacctctagc tagagcttgg cgtaatcatg gtcatagctg
      181 tttcctgtgt gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata
      241 aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca
      301 ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc
      361 gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg
      421 cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta
      481 tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc
      541 aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag
      601 catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac
      661 caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc
      721 ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt
      781 aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc
      841 gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga
      901 cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta
      961 ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta
     1021 tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga
     1081 tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg
     1141 cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag
     1201 tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc
     1261 tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact
     1321 tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt
     1381 cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta
     1441 ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta
     1501 tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc
     1561 gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat
     1621 agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt
     1681 atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg
     1741 tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca
     1801 gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta
     1861 agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg
     1921 cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact
     1981 ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg
     2041 ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt
     2101 actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga
     2161 ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc
     2221 atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa
     2281 caaatagggg ttccgcgcac atttccccga aaagtgccac ctgacgtcga cggatcggga
     2341 gatctcccga tcccctatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa
     2401 gccagtatct gctccctgct tgtgtgttgg aggtcgctga gtagtgcgcg agcaaaattt
     2461 aagctacaac aaggcaaggc ttgaccgaca attgcatgaa gaatctgctt agggttaggc
     2521 gttttgcgct gcttcgcgat gtacgggcca gatatacgcg ttgacattga ttattgacta
     2581 gttattaata gtaatcaatt acggggtcat tagttcatag cccatatatg gagttccgcg
     2641 ttacataact tacggtaaat ggcccgcctg gctgaccgcc caacgacccc cgcccattga
     2701 cgtcaataat gacgtatgtt cccatagtaa cgccaatagg gactttccat tgacgtcaat
     2761 gggtggagta tttacggtaa actgcccact tggcagtaca tcaagtgtat catatgccaa
     2821 gtacgccccc tattgacgtc aatgacggta aatggcccgc ctggcattat gcccagtaca
     2881 tgaccttatg ggactttcct acttggcagt acatctacgt attagtcatc gctattacca
     2941 tggtgatgcg gttttggcag tacatcaatg ggcgtggata gcggtttgac tcacggggat
     3001 ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg
     3061 actttccaaa atgtcgtaac aactccgccc cattgacgca aatgggcggt aggcgtgtac
     3121 ggtgggaggt ctatataagc agagctctct ggctaactag agaacccact gcttactggc
     3181 ttatcgaaat taatacgact cactataggg agacccaagc tggctagcgt ttaaacttaa
     3241 gcttatggtg agcaagggcg aggagctgtt caccggggtg gtgcccatcc tggtcgagct
     3301 ggacggcgac gtaaacggcc acaagttcag cgtgtccggc gagggcgagg gcgatgccac
     3361 ctacggcaag ctgaccctga agctgatctg caccaccggc aagctgcccg tgccctggcc
     3421 caccctcgtg accaccctgg gctacggcct gcagtgcttc gcccgctacc ccgaccacat
     3481 gaagcagcac gacttcttca agtccgccat gcccgaaggc tacgtccagg agcgcaccat
     3541 cttcttcaag gacgacggca actacaagac ccgcgccgag gtgaagttcg agggcgacac
     3601 cctggtgaac cgcatcgagc tgaagggcat cgacttcaag gaggacggca acatcctggg
     3661 gcacaagctg gagtacaact acaacagcca caacgtctat atcaccgccg acaagcagaa
     3721 gaacggcatc aaggccaact tcaagatccg ccacaacatc gaggacggcg gcgtgcagct
     3781 cgccgaccac taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa
     3841 ccactacctg agctaccagt ccaaactgag caaagacccc aacgagaagc gcgatcacat
     3901 ggtcctgctg gagttcgtga ccgccgccgg gatcactctc ggcatggacg agctgtacaa
     3961 gggtaccatc ctggaggacc tcaatatgct ctacatccgg cagatggcac tcagcctgga
     4021 ggacacagag ctgcagagga aactagatca tgagatccgg atgagggatg gggcctgcaa
     4081 gctgctggca gcctgctccc agcgagagca ggctctggaa gccaccaaga gcctgctggt
     4141 gtgcaacagc cgtattctca gctacatggg tgagctgcag cggcgaaagg aggcccaggt
     4201 gctggagaag acaggctagg aattccgatt tccaaagcgg gaaaggaaat caaggagtac
     4261 gtggaggccc aagcaggaaa cgatcctttt ctcaaaggca tccctgagga caagaatccc
     4321 ttcaaggaga aaggtggctg tctgataagc tgactcgagt ctagagggcc cgtttaaacc
     4381 cgctgatcag cctcgactgt gccttctagt tgccagccat ctgttgtttg cccctccccc
     4441 gtgccttcct tgaccctgga aggtgccact cccactgtcc tttcctaata aaatgaggaa
     4501 attgcatcgc attgtctgag taggtgtcat tctattctgg ggggtggggt ggggcaggac
     4561 agcaaggggg aggattggga agacaatagc aggcatgctg gggatgcggt gggctctatg
     4621 gcttctgagg cggaaagaac cagctggggc tctagggggt atccccacgc gccctgtagc
     4681 ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc
     4741 gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt
     4801 ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac
     4861 ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag
     4921 acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa
     4981 actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg
     5041 atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattaattc
     5101 tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca ggcagaagta
     5161 tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag
     5221 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa
     5281 ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc catggctgac
     5341 taattttttt tatttatgca gaggccgagg ccgcctctgc ctctgagcta ttccagaagt
     5401 agtgaggagg cttttttgga ggcctaggct tttgcaaaaa gctcccggga gcttgtatat
     5461 ccattttcgg atctgatcaa gagacaggat gaggatcgtt tcgcatgatt gaacaagatg
     5521 gattgcacgc aggttctccg gccgcttggg tggagaggct attcggctat gactgggcac
     5581 aacagacaat cggctgctct gatgccgccg tgttccggct gtcagcgcag gggcgcccgg
     5641 ttctttttgt caagaccgac ctgtccggtg ccctgaatga actgcaggac gaggcagcgc
     5701 ggctatcgtg gctggccacg acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg
     5761 aagcgggaag ggactggctg ctattgggcg aagtgccggg gcaggatctc ctgtcatctc
     5821 accttgctcc tgccgagaaa gtatccatca tggctgatgc aatgcggcgg ctgcatacgc
     5881 ttgatccggc tacctgccca ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta
     5941 ctcggatgga agccggtctt gtcgatcagg atgatctgga cgaagagcat caggggctcg
     6001 cgccagccga actgttcgcc aggctcaagg cgcgcatgcc cgacggcgag gatctcgtcg
     6061 tgacccatgg cgatgcctgc ttgccgaata tcatggtgga aaatggccgc ttttctggat
     6121 tcatcgactg tggccggctg ggtgtggcgg accgctatca ggacatagcg ttggctaccc
     6181 gtgatattgc tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg ctttacggta
     6241 tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct tcttgacgag ttcttctgag
     6301 cgggactctg gggttcgaaa tgaccgacca agcgacgccc aacctgccat cacgagattt
     6361 cgattccacc gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg
     6421 ctggatgatc ctccagcgcg gggatctcat gctggagttc ttcgcccacc cc
//
