LOCUS       sequence-195046-        7653 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of Rock1-GFP for the study of localisation 
            signaling..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7653)
  AUTHORS   Priya R, Gomez GA, Budnar S, Verma S, Cox HL, Hamilton NA, Yap AS
  TITLE     Feedback regulation through myosin II confers robustness on RhoA 
            signalling at E-cadherin junctions.
  JOURNAL   Nat Cell Biol. 2015 Oct;17(10):1282-93. doi: 10.1038/ncb3239. Epub 
            2015 Sep 14.
  PUBMED    26368311
REFERENCE   2  (bases 1 to 7653)
  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"; doi: "10.1038/ncb3239"; 
            journalName: "Nat Cell Biol"; date: "2015-10"; volume: "17"; issue: 
            "10"; pages: "1282-93"
COMMENT     Sequence Label: GFP-mROCK1-delta6
FEATURES             Location/Qualifiers
     source          1..7653
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      17..605
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     506..525
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        780..1083
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1084..1287
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1238..1258
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      1326..1335
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1332..1334,1335..1337,1338..2048)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1332..1334,1335..1337,1338..2048)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1377..1398)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1638..1657)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1985..2006
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    5160..5281
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5197..5216)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5251..5270
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(5288..5743)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5425..5446)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5637..5656
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        5770..5874
                     /gene="bla"
                     /label=AmpR promoter
     promoter        5876..6233
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        5876..6067
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(5899..5919)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      6084..6219
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             6268..7062
                     /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             6268..7062
                     /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(6322..6341)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6932..6951
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(7250..7269)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    7294..7341
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
ORIGIN
        1 agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa
       61 aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc
      121 cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt
      181 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc
      241 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac
      301 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca
      361 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg
      421 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag
      481 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt
      541 ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat
      601 ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc
      661 acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gccatgcatt
      721 agttattaat agtaatcaat tacggggtca ttagttcata gcccatatat ggagttccgc
      781 gttacataac ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg
      841 acgtcaataa tgacgtatgt tcccatagta acgccaatag ggactttcca ttgacgtcaa
      901 tgggtggagt atttacggta aactgcccac ttggcagtac atcaagtgta tcatatgcca
      961 agtacgcccc ctattgacgt caatgacggt aaatggcccg cctggcatta tgcccagtac
     1021 atgaccttat gggactttcc tacttggcag tacatctacg tattagtcat cgctattacc
     1081 atggtgatgc ggttttggca gtacatcaat gggcgtggat agcggtttga ctcacgggga
     1141 tttccaagtc tccaccccat tgacgtcaat gggagtttgt tttggcacca aaatcaacgg
     1201 gactttccaa aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg taggcgtgta
     1261 cggtgggagg tctatataag cagagctggt ttagtgaacc gtcagatccg ctagcgctac
     1321 cggtcgccac catggtgagc aagggcgagg agctgttcac cggggtggtg cccatcctgg
     1381 tcgagctgga cggcgacgta aacggccaca agttcagcgt gtccggcgag ggcgagggcg
     1441 atgccaccta cggcaagctg accctgaagt tcatctgcac caccggcaag ctgcccgtgc
     1501 cctggcccac cctcgtgacc accctgacct acggcgtgca gtgcttcagc cgctaccccg
     1561 accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc
     1621 gcaccatctt cttcaaggac gacggcaact acaagacccg cgccgaggtg aagttcgagg
     1681 gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag gacggcaaca
     1741 tcctggggca caagctggag tacaactaca acagccacaa cgtctatatc atggccgaca
     1801 agcagaagaa cggcatcaag gtgaacttca agatccgcca caacatcgag gacggcagcg
     1861 tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc gtgctgctgc
     1921 ccgacaacca ctacctgagc acccagtccg ccctgagcaa agaccccaac gagaagcgcg
     1981 atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactctcggc atggacgagc
     2041 tgtacaagtc cggactcaga tctcgagctc aaaaaggaga tgaagaaaca tttcctattc
     2101 caaaagcttt tgttggcaat cagctacctt ttgtaggatt tacatattat agcaatcgta
     2161 gatacttacc ttcagcaaat gccagtgaga acagatctag ctccaatgtg gataaaagct
     2221 tgcaggaaag tttacagaaa acaatctaca agcttgaaga gcaactgcat aatgaaatgc
     2281 aattaaaaga tgaaatggag cagaaatgca gaacttcaaa cttaaaacta gacaagataa
     2341 tgaaagaatt ggatgaagag gggaatcaaa gaagaaatct agaatctgca gtgtctcaaa
     2401 ttgagaaaga aaagatgttg ctacaacata gaattaatga gtaccaaaga aaagtggaac
     2461 aggaaaatga gaagagaaga aatatagaaa atgaagtttc tacactaaag gaccagttag
     2521 aagacttacg gaaagcaagt cagacctcac agcttgctaa tgagaagctg acacagttac
     2581 agaagcagtt agaagaagcg aatgacttac ttaggacaga atcagacaca gctgtaagat
     2641 taaggaagag tcacacagag atgagcaagt caattagtca gctagagtcc ctgaacagag
     2701 aattacagga aagaaacaga attttagaga attccaagtc acaagcagac aaggattact
     2761 accagctgca agctgtattg gaagctgagc gaagagacag agggcatgat tctgagatga
     2821 ttggagacct tcaagcacga attacatctt tacaagaaga agtgaaacat ctcaaacata
     2881 atcttgaaag agtggaagga gaaagaaaag aggctcaaga catgcttaat cattcagaaa
     2941 aggaaaagaa taatttggag atagatttga attacaagct caagtctata caacagcggt
     3001 tagaacaaga agtaaatgaa cataaagtca ccaaagcacg cctaactgac aagcaccaat
     3061 ctatcgaaga ggcaaagtct gttgccatgt gtgaaatgga aaaaaagctg aaggaagaga
     3121 gagaagctcg agagaaggca gaaaatagag ttgttgagac tgagaaacaa tgttccatgc
     3181 tagatgttga cctgaaacag tctcagcaga agctagaaca tttgaccgaa aataaagaaa
     3241 ggatggagga tgaagttaag aatctcgccc tgcagctgga acaggaatca aataaacggc
     3301 tcttgttaca gaatgaattg aagactcaag catttgaagc agacaattta aaaggtttag
     3361 aaaaacagat gaaacaggaa ataaatactt tattggaagc aaagagattg ttagaatttg
     3421 agttagctca gcttacaaag caatataggg gaaatgaagg gcaaatgcgg gagttacaag
     3481 atcaacttga agctgagcag tatttctcta cactttataa aacccaagta aaggagctta
     3541 aagaagaaat tgaagaaaaa aaccgagaaa atttaaggaa aatacaggaa ctgcaaagtg
     3601 aaaaagaaac tctctctact cagttggatt tagcagaaac aaaagctgag tctgagcagc
     3661 tggcgcgagg gattctggaa gaacagtatt ttgagttgac tcaggaaagc aagaaagctg
     3721 cttcaagaaa tagacaagag attacagata aagatcatac tgttagtcgg cttgaagaaa
     3781 caaacagtgt gctaaccaaa gatattgaaa tgctaagaaa agaaaatgaa gagctaaatg
     3841 aaagaatgag gacagcagaa gaagaatata aattgaagaa ggaggaagaa ataaataatc
     3901 ttaaggctgc ttttgaaaag aatatcagta ccgaacggac ccttaaaaca caggctgtta
     3961 acaagttggc agaaataatg aatcgcaaag attttaaaat tgatcgaaag aaagctaata
     4021 cacaagattt aagaaaaaaa gaaaaggaaa atagaaagct acaactggaa cttaaccaag
     4081 agagagaaaa atttaaccaa atggtggtga aacaccagaa ggagctgaat gacatgcaag
     4141 cgcaattggt agaagaatgt acacatagaa atgaacttca aatgcagttg gctagcaaag
     4201 agagtgatat tgagcagttg cgggcaaaac ttttggacct ttcggattca actagtgttg
     4261 ctagttttcc tagtgcagat gaaactgatg gaaaccttcc agagtcaaga attgaaggtt
     4321 ggctttctgt accaaataga ggaaatatca aaagatatgg ctggaagaaa cagtatgttg
     4381 tggtaagcag caaaaagatc ttgttctata atgatgaaca agataaggaa caatccagtc
     4441 catctatggt gttagacata gataaattgt ttcatgtccg acctgtaacc caaggagatg
     4501 tgtacagagc agaaactgaa gaaattccta aaatattcca gatactatat gcaaatgaag
     4561 gtgaatgtag aaaagatata gaagtggaac cggttcaaca aggtgaaaaa actaatttcc
     4621 agaatcacaa aggccatgaa ttcattccta ccctctacca ctttcctgcc aactgtgagg
     4681 cctgtgccaa acctttgtgg catgtcttta agccaccacc tgccctagag tgtcgaagat
     4741 gccatgttaa gtgccacaga gaccatctag acaagaaaga ggacttgatt tccccgtgca
     4801 aagtaagtta cgatgtaaca tcagcaagag atatgctgct gttagcatgt tctcaggatg
     4861 aacaaaaaaa gtgggttact catctagtaa agaagattcc taagaaccca ccatctggct
     4921 ttgtccgtgc ctctcctcga acgctgtcaa caagatccac tgcaaatcag tctttccgca
     4981 aagtagtcaa aaatacatct ggaaaaacta gttaaggtac cgcgggcccg ggatccaccg
     5041 gatctagata actgatcata atcagccata ccacatttgt agaggtttta cttgctttaa
     5101 aaaacctccc acacctcccc ctgaacctga aacataaaat gaatgcaatt gttgttgtta
     5161 acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa
     5221 ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt
     5281 aacgcgtaaa ttgtaagcgt taatattttg ttaaaattcg cgttaaattt ttgttaaatc
     5341 agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag
     5401 accgagatag ggttgagtgt tgttccagtt tggaacaaga gtccactatt aaagaacgtg
     5461 gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg atggcccact acgtgaacca
     5521 tcaccctaat caagtttttt ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa
     5581 gggagccccc gatttagagc ttgacgggga aagccggcga acgtggcgag aaaggaaggg
     5641 aagaaagcga aaggagcggg cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta
     5701 accaccacac ccgccgcgct taatgcgccg ctacagggcg cgtcaggtgg cacttttcgg
     5761 ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg
     5821 ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtcctgag
     5881 gcggaaagaa ccagctgtgg aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc
     5941 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccagg tgtggaaagt
     6001 ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca
     6061 tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc gcccattctc
     6121 cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc tcggcctctg
     6181 agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaagatcgat
     6241 caagagacag gatgaggatc gtttcgcatg attgaacaag atggattgca cgcaggttct
     6301 ccggccgctt gggtggagag gctattcggc tatgactggg cacaacagac aatcggctgc
     6361 tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc cggttctttt tgtcaagacc
     6421 gacctgtccg gtgccctgaa tgaactgcaa gacgaggcag cgcggctatc gtggctggcc
     6481 acgacgggcg ttccttgcgc agctgtgctc gacgttgtca ctgaagcggg aagggactgg
     6541 ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag
     6601 aaagtatcca tcatggctga tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc
     6661 ccattcgacc accaagcgaa acatcgcatc gagcgagcac gtactcggat ggaagccggt
     6721 cttgtcgatc aggatgatct ggacgaagag catcaggggc tcgcgccagc cgaactgttc
     6781 gccaggctca aggcgagcat gcccgacggc gaggatctcg tcgtgaccca tggcgatgcc
     6841 tgcttgccga atatcatggt ggaaaatggc cgcttttctg gattcatcga ctgtggccgg
     6901 ctgggtgtgg cggaccgcta tcaggacata gcgttggcta cccgtgatat tgctgaagag
     6961 cttggcggcg aatgggctga ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg
     7021 cagcgcatcg ccttctatcg ccttcttgac gagttcttct gagcgggact ctggggttcg
     7081 aaatgaccga ccaagcgacg cccaacctgc catcacgaga tttcgattcc accgccgcct
     7141 tctatgaaag gttgggcttc ggaatcgttt tccgggacgc cggctggatg atcctccagc
     7201 gcggggatct catgctggag ttcttcgccc accctagggg gaggctaact gaaacacgga
     7261 aggagacaat accggaagga acccgcgcta tgacggcaat aaaaagacag aataaaacgc
     7321 acggtgttgg gtcgtttgtt cataaacgcg gggttcggtc ccagggctgg cactctgtcg
     7381 ataccccacc gagaccccat tggggccaat acgcccgcgt ttcttccttt tccccacccc
     7441 accccccaag ttcgggtgaa ggcccagggc tcgcagccaa cgtcggggcg gcaggccctg
     7501 ccatagcctc aggttactca tatatacttt agattgattt aaaacttcat ttttaattta
     7561 aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt
     7621 tttcgttcca ctgagcgtca gaccccgtag aaa
//
