LOCUS       sequence-195036-        7953 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 7953)
  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 7953)
  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-delta3
FEATURES             Location/Qualifiers
     source          1..7953
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      7..595
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     496..515
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        770..1073
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1074..1277
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1228..1248
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      1316..1325
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1322..1324,1325..1327,1328..2038)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1322..1324,1325..1327,1328..2038)
                     /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(1367..1388)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1628..1647)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1975..1996
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    5450..5571
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5487..5506)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5541..5560
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(5578..6033)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5715..5736)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5927..5946
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        6060..6164
                     /gene="bla"
                     /label=AmpR promoter
     promoter        6166..6523
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        6166..6357
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(6189..6209)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      6374..6509
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             6558..7352
                     /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             6558..7352
                     /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(6612..6631)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     7222..7241
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(7540..7559)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    7584..7631
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
ORIGIN
        1 atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc
       61 gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac
      121 tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca
      181 ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt
      241 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc
      301 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg
      361 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc
      421 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac
      481 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct
      541 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc
      601 cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt
      661 tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gccatgcatt agttattaat
      721 agtaatcaat tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac
      781 ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa
      841 tgacgtatgt tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt
      901 atttacggta aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc
      961 ctattgacgt caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttat
     1021 gggactttcc tacttggcag tacatctacg tattagtcat cgctattacc atggtgatgc
     1081 ggttttggca gtacatcaat gggcgtggat agcggtttga ctcacgggga tttccaagtc
     1141 tccaccccat tgacgtcaat gggagtttgt tttggcacca aaatcaacgg gactttccaa
     1201 aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg taggcgtgta cggtgggagg
     1261 tctatataag cagagctggt ttagtgaacc gtcagatccg ctagcgctac cggtcgccac
     1321 catggtgagc aagggcgagg agctgttcac cggggtggtg cccatcctgg tcgagctgga
     1381 cggcgacgta aacggccaca agttcagcgt gtccggcgag ggcgagggcg atgccaccta
     1441 cggcaagctg accctgaagt tcatctgcac caccggcaag ctgcccgtgc cctggcccac
     1501 cctcgtgacc accctgacct acggcgtgca gtgcttcagc cgctaccccg accacatgaa
     1561 gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc gcaccatctt
     1621 cttcaaggac gacggcaact acaagacccg cgccgaggtg aagttcgagg gcgacaccct
     1681 ggtgaaccgc atcgagctga agggcatcga cttcaaggag gacggcaaca tcctggggca
     1741 caagctggag tacaactaca acagccacaa cgtctatatc atggccgaca agcagaagaa
     1801 cggcatcaag gtgaacttca agatccgcca caacatcgag gacggcagcg tgcagctcgc
     1861 cgaccactac cagcagaaca cccccatcgg cgacggcccc gtgctgctgc ccgacaacca
     1921 ctacctgagc acccagtccg ccctgagcaa agaccccaac gagaagcgcg atcacatggt
     1981 cctgctggag ttcgtgaccg ccgccgggat cactctcggc atggacgagc tgtacaagtc
     2041 cggactcaga tctcgagctc aaatgtcgac tggggacagt tttgagactc ggtttgaaaa
     2101 aatcgacaac ctgctgcggg atcccaaatc ggaagtgaat tcagattgtt tgctggatgg
     2161 attggatgcc ctggtttatg atttggattt tcctgcctta agaaaaaaca aaaatattga
     2221 caactttcta agcagatata aagacacaat aaataaaatt agagatttac ggatgaaagc
     2281 agaagattat gaagtggtaa aggtaatcgg cagaggtgca tttggagaag ttcaattggt
     2341 gaggcataaa tccactagga aggtttatgc tatgaagctg ctcagcaaat ttgaaatgat
     2401 taagagatcg gattctgctt ttttctggga agaaagggac atcatggcgt ttgccaatag
     2461 tccttgggtt gttcagcttt tttatgcatt ccaagatgac cggtatctct acatggtgat
     2521 ggagtacatg cctggcggag accttgtcaa cctgatgagc aactatgacg tgcctgaaaa
     2581 atgggcaaga ttctatactg cagaagtagt tcttgcattg gatgcaatcc attccatggg
     2641 ttttattcat agagatgtaa agcctgataa catgctgctg gataagtctg gacatttgaa
     2701 gttagcagat tttggtactt gtatgaagat gaataaggaa ggtatggtac gatgtgacac
     2761 agcagttgga acacctgatt atatctcacc cgaagtatta aaatcccaag gcggtgatgg
     2821 ctattatgga cgagagtgtg actggtggtc agttggggta tttctatatg aaatgctcgt
     2881 aggtgacaca cctttctatg cagattcttt ggttgggact tacagtaaaa ttatgaacca
     2941 taaaaattca cttactttcc ctgatgataa tgacatatca aaagaggcaa aaaatctcat
     3001 ttgtgccttc cttacagaca gagaagtgag attgggcaga aatggtgtag aggaaatcaa
     3061 acgtcatctc ttcttcaaaa atgatcagtg ggcttgggaa acgctccgag acactgtagc
     3121 accagttgtg cctgatttaa gtagtgacat tgatactagt aattttgatg acttagaaga
     3181 agataaagga gatgaagaaa catttcctat tccaaaagct tttgttggca atcagctacc
     3241 ttttgtagga tttacatatt atagcaatcg tagatactta ccttcagcaa atgccagtga
     3301 gaacagatct agctccaatg tggataaaag cttgcaggaa agtttacaga aaacaatcta
     3361 caagcttgaa gagcaactgc ataatgaaat gcaattaaaa gatgaaatgg agcagaaatg
     3421 cagaacttca aacttaaaac tagacaagat aatgaaagaa ttggatgaag aggggaatca
     3481 aagaagaaat ctagaatctg cagtgtctca aattgagaaa gaaaagatgt tgctacaaca
     3541 tagaattaat gagtaccaaa gaaaagtgga acaggaaaat gagaagagaa gaaatataga
     3601 aaatgaagtt tctacactaa aggaccagtt agaagactta cggaaagcaa gtcagacctc
     3661 acagcttgct aatgagaagc tgacacagtt acagaagcag ttagaagaag cgaatgactt
     3721 acttaggaca gaatcagaca cagctgtaag attaaggaag agtcacacag agatgagcaa
     3781 gtcaattagt cagctagagt ccctgaacag agaattacag gaaagaaaca gaattttaga
     3841 gaattccaag tcacaagcag acaaggatta ctaccagctg caagctgtat tggaagctga
     3901 gcgaagagac agagggcatg attctgagat gattggagac cttcaagcac gaattacatc
     3961 tttacaagaa gaagtgaaac atctcaaaca taatcttgaa agagtggaag gagaaagaaa
     4021 agaggctcaa gacatgctta atcattcaga aaaggaaaag aataatttgg agatagattt
     4081 gaattacaag ctcaagtcta tacaacagcg gttagaacaa gaagtaaatg aacataaagt
     4141 caccaaagca cgcctaactg acaagcacca atctatcgaa gaggcaaagt ctgttgccat
     4201 gtgtgaaatg gaaaaaaagc tgaaggaaga gagagaagct cgagagaagg cagaaaatag
     4261 agttgttgag actgagaaac aatgttccat gctagatgtt gacctgaaac agtctcagca
     4321 gaagctagaa catttgaccg aaaataaaga aaggatggag gatgaagtta agaatctcgc
     4381 cctgcagctg gaacaggaat caaataaacg gctcttgtta cagaatgaat tgaagactca
     4441 agcatttgaa gcagacaatt taaaaggttt agaaaaacag atgaaacagg aaataaatac
     4501 tttattggaa gcaaagagat tgttagaatt tgagttagct cagcttacaa agcaatatag
     4561 gggaaatgaa gggcaaatgc gggagttaca agatcaactt gaagctgagc agtatttctc
     4621 tacactttat aaaacccaag taaaggagct taaagaagaa attgaagaaa aaaaccgaga
     4681 aaatttaagg aaaatacagg aactgcaaag tgaaaaagaa actctctcta ctcagttgga
     4741 tttagcagaa acaaaagctg agtctgagca gctggcgcga gggattctgg aagaacagta
     4801 ttttgagttg actcaggaaa gcaagaaagc tgcttcaaga aatagacaag agattacaga
     4861 taaagatcat actgttagtc ggcttgaaga aacaaacagt gtgctaacca aagatattga
     4921 aatgctaaga aaagaaaatg aagagctaaa tgaaagaatg aggacagcag aagaagaata
     4981 taaattgaag aaggaggaag aaataaataa tcttaaggct gcttttgaaa agaatatcag
     5041 taccgaacgg acccttaaaa cacaggctgt taacaagttg gcagaaataa tgaatcgcaa
     5101 agattttaaa attgatcgaa agaaagctaa tacacaagat ttaagaaaaa aagaaaagga
     5161 aaatagaaag ctacaactgg aacttaacca agagagagaa aaatttaacc aaatggtggt
     5221 gaaacaccag aaggagctga atgacatgca agcgcaattg gtagaagaat gtacacatag
     5281 aaatgaactt caaatgcagt tgtaaggtac cgcgggcccg ggatccaccg gatctagata
     5341 actgatcata atcagccata ccacatttgt agaggtttta cttgctttaa aaaacctccc
     5401 acacctcccc ctgaacctga aacataaaat gaatgcaatt gttgttgtta acttgtttat
     5461 tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt
     5521 tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt aacgcgtaaa
     5581 ttgtaagcgt taatattttg ttaaaattcg cgttaaattt ttgttaaatc agctcatttt
     5641 ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag accgagatag
     5701 ggttgagtgt tgttccagtt tggaacaaga gtccactatt aaagaacgtg gactccaacg
     5761 tcaaagggcg aaaaaccgtc tatcagggcg atggcccact acgtgaacca tcaccctaat
     5821 caagtttttt ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa gggagccccc
     5881 gatttagagc ttgacgggga aagccggcga acgtggcgag aaaggaaggg aagaaagcga
     5941 aaggagcggg cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta accaccacac
     6001 ccgccgcgct taatgcgccg ctacagggcg cgtcaggtgg cacttttcgg ggaaatgtgc
     6061 gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac
     6121 aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtcctgag gcggaaagaa
     6181 ccagctgtgg aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc cagcaggcag
     6241 aagtatgcaa agcatgcatc tcaattagtc agcaaccagg tgtggaaagt ccccaggctc
     6301 cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca tagtcccgcc
     6361 cctaactccg cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg
     6421 ctgactaatt ttttttattt atgcagaggc cgaggccgcc tcggcctctg agctattcca
     6481 gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaagatcgat caagagacag
     6541 gatgaggatc gtttcgcatg attgaacaag atggattgca cgcaggttct ccggccgctt
     6601 gggtggagag gctattcggc tatgactggg cacaacagac aatcggctgc tctgatgccg
     6661 ccgtgttccg gctgtcagcg caggggcgcc cggttctttt tgtcaagacc gacctgtccg
     6721 gtgccctgaa tgaactgcaa gacgaggcag cgcggctatc gtggctggcc acgacgggcg
     6781 ttccttgcgc agctgtgctc gacgttgtca ctgaagcggg aagggactgg ctgctattgg
     6841 gcgaagtgcc ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca
     6901 tcatggctga tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc ccattcgacc
     6961 accaagcgaa acatcgcatc gagcgagcac gtactcggat ggaagccggt cttgtcgatc
     7021 aggatgatct ggacgaagag catcaggggc tcgcgccagc cgaactgttc gccaggctca
     7081 aggcgagcat gcccgacggc gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga
     7141 atatcatggt ggaaaatggc cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg
     7201 cggaccgcta tcaggacata gcgttggcta cccgtgatat tgctgaagag cttggcggcg
     7261 aatgggctga ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg
     7321 ccttctatcg ccttcttgac gagttcttct gagcgggact ctggggttcg aaatgaccga
     7381 ccaagcgacg cccaacctgc catcacgaga tttcgattcc accgccgcct tctatgaaag
     7441 gttgggcttc ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct
     7501 catgctggag ttcttcgccc accctagggg gaggctaact gaaacacgga aggagacaat
     7561 accggaagga acccgcgcta tgacggcaat aaaaagacag aataaaacgc acggtgttgg
     7621 gtcgtttgtt cataaacgcg gggttcggtc ccagggctgg cactctgtcg ataccccacc
     7681 gagaccccat tggggccaat acgcccgcgt ttcttccttt tccccacccc accccccaag
     7741 ttcgggtgaa ggcccagggc tcgcagccaa cgtcggggcg gcaggccctg ccatagcctc
     7801 aggttactca tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta
     7861 ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca
     7921 ctgagcgtca gaccccgtag aaaagatcaa agg
//
