LOCUS       sequence-195043-        5772 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 5772)
  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 5772)
  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-delta5
FEATURES             Location/Qualifiers
     source          1..5772
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        62..365
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        366..569
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     520..540
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      608..617
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(614..616,617..619,620..1330)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(614..616,617..619,620..1330)
                     /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(659..680)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(920..939)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1267..1288
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2561..2682
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2598..2617)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2652..2671
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2689..3144)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2826..2847)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3038..3057
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3171..3275
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3277..3634
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3277..3468
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3300..3320)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3485..3620
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3669..4463
                     /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             3669..4463
                     /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(3723..3742)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4333..4352
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4651..4670)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4695..4742
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5071..5659
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5560..5579
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ttagttatta atagtaatca attacggggt cattagttca tagcccatat atggagttcc
       61 gcgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac ccccgcccat
      121 tgacgtcaat aatgacgtat gttcccatag taacgccaat agggactttc cattgacgtc
      181 aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg tatcatatgc
      241 caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt
      301 acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc atcgctatta
      361 ccatggtgat gcggttttgg cagtacatca atgggcgtgg atagcggttt gactcacggg
      421 gatttccaag tctccacccc attgacgtca atgggagttt gttttggcac caaaatcaac
      481 gggactttcc aaaatgtcgt aacaactccg ccccattgac gcaaatgggc ggtaggcgtg
      541 tacggtggga ggtctatata agcagagctg gtttagtgaa ccgtcagatc cgctagcgct
      601 accggtcgcc accatggtga gcaagggcga ggagctgttc accggggtgg tgcccatcct
      661 ggtcgagctg gacggcgacg taaacggcca caagttcagc gtgtccggcg agggcgaggg
      721 cgatgccacc tacggcaagc tgaccctgaa gttcatctgc accaccggca agctgcccgt
      781 gccctggccc accctcgtga ccaccctgac ctacggcgtg cagtgcttca gccgctaccc
      841 cgaccacatg aagcagcacg acttcttcaa gtccgccatg cccgaaggct acgtccagga
      901 gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg tgaagttcga
      961 gggcgacacc ctggtgaacc gcatcgagct gaagggcatc gacttcaagg aggacggcaa
     1021 catcctgggg cacaagctgg agtacaacta caacagccac aacgtctata tcatggccga
     1081 caagcagaag aacggcatca aggtgaactt caagatccgc cacaacatcg aggacggcag
     1141 cgtgcagctc gccgaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct
     1201 gcccgacaac cactacctga gcacccagtc cgccctgagc aaagacccca acgagaagcg
     1261 cgatcacatg gtcctgctgg agttcgtgac cgccgccggg atcactctcg gcatggacga
     1321 gctgtacaag tccggactca gatctcgagc tcaaaaagga gatgaagaaa catttcctat
     1381 tccaaaagct tttgttggca atcagctacc ttttgtagga tttacatatt atagcaatcg
     1441 tagatactta ccttcagcaa atgccagtga gaacagatct agctccaatg tggataaaag
     1501 cttgcaggaa agtttacaga aaacaatcta caagcttgaa gagcaactgc ataatgaaat
     1561 gcaattaaaa gatgaaatgg agcagaaatg cagaacttca aacttaaaac tagacaagat
     1621 aatgaaagaa ttggatgaag aggggaatca aagaagaaat ctagaatctg cagtgtctca
     1681 aattgagaaa gaaaagatgt tgctacaaca tagaattaat gagtaccaaa gaaaagtgga
     1741 acaggaaaat gagaagagaa gaaatataga aaatgaagtt tctacactaa aggaccagtt
     1801 agaagactta cggaaagcaa gtcagacctc acagcttgct aatgagaagc tgacacagtt
     1861 acagaagcag ttagaagaag cgaatgactt acttaggaca gaatcagaca cagctgtaag
     1921 attaaggaag agtcacacag agatgagcaa gtcaattagt cagctagagt ccctgaacag
     1981 agaattacag gaaagaaaca gaattttaga gaattccaag tcacaagcag acaaggatta
     2041 ctaccagctg caagctgtat tggaagctga gcgaagagac agagggcatg attctgagat
     2101 gattggagac cttcaagcac gaattacatc tttacaagaa gaagtgaaac atctcaaaca
     2161 taatcttgaa agagtggaag gagaaagaaa agaggctcaa gacatgctta atcattcaga
     2221 aaaggaaaag aataatttgg agatagattt gaattacaag ctcaagtcta tacaacagcg
     2281 gttagaacaa gaagtaaatg aacataaagt caccaaagca cgcctaactg acaagcacca
     2341 atctatcgaa gaggcaaagt ctgttgccat gtgtgaaatg gaaaaaaagc tgaaggaaga
     2401 gagagaagct cgataaggta ccgcgggccc gggatccacc ggatctagat aactgatcat
     2461 aatcagccat accacatttg tagaggtttt acttgcttta aaaaacctcc cacacctccc
     2521 cctgaacctg aaacataaaa tgaatgcaat tgttgttgtt aacttgttta ttgcagctta
     2581 taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact
     2641 gcattctagt tgtggtttgt ccaaactcat caatgtatct taacgcgtaa attgtaagcg
     2701 ttaatatttt gttaaaattc gcgttaaatt tttgttaaat cagctcattt tttaaccaat
     2761 aggccgaaat cggcaaaatc ccttataaat caaaagaata gaccgagata gggttgagtg
     2821 ttgttccagt ttggaacaag agtccactat taaagaacgt ggactccaac gtcaaagggc
     2881 gaaaaaccgt ctatcagggc gatggcccac tacgtgaacc atcaccctaa tcaagttttt
     2941 tggggtcgag gtgccgtaaa gcactaaatc ggaaccctaa agggagcccc cgatttagag
     3001 cttgacgggg aaagccggcg aacgtggcga gaaaggaagg gaagaaagcg aaaggagcgg
     3061 gcgctagggc gctggcaagt gtagcggtca cgctgcgcgt aaccaccaca cccgccgcgc
     3121 ttaatgcgcc gctacagggc gcgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc
     3181 ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct
     3241 gataaatgct tcaataatat tgaaaaagga agagtcctga ggcggaaaga accagctgtg
     3301 gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     3361 aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct ccccagcagg
     3421 cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc
     3481 gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat
     3541 tttttttatt tatgcagagg ccgaggccgc ctcggcctct gagctattcc agaagtagtg
     3601 aggaggcttt tttggaggcc taggcttttg caaagatcga tcaagagaca ggatgaggat
     3661 cgtttcgcat gattgaacaa gatggattgc acgcaggttc tccggccgct tgggtggaga
     3721 ggctattcgg ctatgactgg gcacaacaga caatcggctg ctctgatgcc gccgtgttcc
     3781 ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga
     3841 atgaactgca agacgaggca gcgcggctat cgtggctggc cacgacgggc gttccttgcg
     3901 cagctgtgct cgacgttgtc actgaagcgg gaagggactg gctgctattg ggcgaagtgc
     3961 cggggcagga tctcctgtca tctcaccttg ctcctgccga gaaagtatcc atcatggctg
     4021 atgcaatgcg gcggctgcat acgcttgatc cggctacctg cccattcgac caccaagcga
     4081 aacatcgcat cgagcgagca cgtactcgga tggaagccgg tcttgtcgat caggatgatc
     4141 tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc aaggcgagca
     4201 tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg
     4261 tggaaaatgg ccgcttttct ggattcatcg actgtggccg gctgggtgtg gcggaccgct
     4321 atcaggacat agcgttggct acccgtgata ttgctgaaga gcttggcggc gaatgggctg
     4381 accgcttcct cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc gccttctatc
     4441 gccttcttga cgagttcttc tgagcgggac tctggggttc gaaatgaccg accaagcgac
     4501 gcccaacctg ccatcacgag atttcgattc caccgccgcc ttctatgaaa ggttgggctt
     4561 cggaatcgtt ttccgggacg ccggctggat gatcctccag cgcggggatc tcatgctgga
     4621 gttcttcgcc caccctaggg ggaggctaac tgaaacacgg aaggagacaa taccggaagg
     4681 aacccgcgct atgacggcaa taaaaagaca gaataaaacg cacggtgttg ggtcgtttgt
     4741 tcataaacgc ggggttcggt cccagggctg gcactctgtc gataccccac cgagacccca
     4801 ttggggccaa tacgcccgcg tttcttcctt ttccccaccc caccccccaa gttcgggtga
     4861 aggcccaggg ctcgcagcca acgtcggggc ggcaggccct gccatagcct caggttactc
     4921 atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat
     4981 cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc
     5041 agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg
     5101 ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct
     5161 accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct
     5221 tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct
     5281 cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg
     5341 gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc
     5401 gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga
     5461 gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg
     5521 cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta
     5581 tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg
     5641 ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg
     5701 ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat
     5761 taccgccatg ca
//
