LOCUS       sequence-448491-        7189 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express mEGFP-fusion protein of ROCK2 fragment (941-1388).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7189)
  AUTHORS   Chen X, Fansler MM, Janjos U, Ule J, Mayr C
  TITLE     The FXR1 network acts as signaling scaffold for actomyosin 
            remodeling.
  JOURNAL   Cell. 5 August 2024. doi: 10.1016/j.cell.2024.07.015.
  PUBMED    37961296
REFERENCE   2  (bases 1 to 7189)
  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: "Cell. 5 
            August 2024. doi: 10.1016/j.cell.2024.07.015."
COMMENT     Sequence Label: pcDNA3puro-mGFP-ROCK2-C (941-1388)
FEATURES             Location/Qualifiers
     source          1..7189
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(905..907,908..910,911..1621)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(950..971)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1211..1230)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1558..1579
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(3029..3046)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3035..3259
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3305..3733
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3392..3411)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3602..3623
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3742..3762)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3747..4076
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3927..4062
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3989..4008
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4089..4688
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(4089..4108)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4585..4605
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4862..4983
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4899..4918)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4953..4972
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5032..5048)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5032..5048)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5045..5067)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5056..5072
                     /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(5080..5086,5087..5104,5105..5110))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5125..5146
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5263..5280)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5434..6022)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5514..5533)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6193..6984,6985..7053))
                     /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     6816..6835
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7054..7158)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 aaccatggtg agcaagggcg aggagctgtt caccggggtg gtgcccatcc tggtcgagct
      961 ggacggcgac gtaaacggcc acaagttcag cgtgtccggc gagggcgagg gcgatgccac
     1021 ctacggcaag ctgaccctga agttcatctg caccaccggc aagctgcccg tgccctggcc
     1081 caccctcgtg accaccctga cctacggcgt gcagtgcttc agccgctacc ccgaccacat
     1141 gaagcagcac gacttcttca agtccgccat gcccgaaggc tacgtccagg agcgcaccat
     1201 cttcttcaag gacgacggca actacaagac ccgcgccgag gtgaagttcg agggcgacac
     1261 cctggtgaac cgcatcgagc tgaagggcat cgacttcaag gaggacggca acatcctggg
     1321 gcacaagctg gagtacaact acaacagcca caacgtctat atcatggccg acaagcagaa
     1381 gaacggcatc aaggtgaact tcaagatccg ccacaacatc gaggacggca gcgtgcagct
     1441 cgccgaccac taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa
     1501 ccactacctg agcacccagt ccaagctgag caaagacccc aacgagaagc gcgatcacat
     1561 ggtcctgctg gagttcgtga ccgccgccgg gatcactctc ggcatggacg agctgtacaa
     1621 gaaagagaag atcatgaaag agctggagat caaagagatg atggctagac acaaacagga
     1681 acttacggaa aaagatgcta caattgcttc tcttgaggaa actaatagga cactaactag
     1741 tgatgttgcc aatcttgcaa atgagaaaga agaattaaat aacaaattga aagatgttca
     1801 agagcaactg tcaagattga aagatgaaga aataagcgca gcagctatta aagcacagtt
     1861 tgagaagcag ctattaacag aaagaacact caaaactcaa gctgtgaata agttggctga
     1921 gatcatgaat cgaaaagaac ctgtcaagcg tggtaatgac acagatgtgc ggagaaaaga
     1981 gaaggagaat agaaagctac atatggagct taaatctgaa cgtgagaaat tgacccagca
     2041 gatgatcaag tatcagaaag aactgaatga aatgcaggca caaatagctg aagagagcca
     2101 gattcgaatt gaactgcaga tgacattgga cagtaaagac agtgacattg agcagctgcg
     2161 gtcacaactc caagccttgc atattggtct ggatagttcc agtataggca gtggaccagg
     2221 ggatgctgag gcagatgatg ggtttccaga atcaagatta gaaggatggc tttcattgcc
     2281 tgtacgaaac aacactaaga aatttggatg ggttaaaaag tatgtgattg taagcagtaa
     2341 gaagattctt ttctatgaca gtgaacaaga taaagaacaa tccaatcctt acatggtttt
     2401 agatatagac aagttatttc atgtccgacc agttacacag acagatgtgt atagagcaga
     2461 tgctaaagaa attccaagga tattccagat tctgtatgcc aatgaaggag aaagtaagaa
     2521 ggaacaagaa tttccagtgg agccagttgg agaaaaatct aattatattt gccacaaggg
     2581 acatgagttt attcctactc tttatcattt cccaaccaac tgtgaggctt gtatgaagcc
     2641 cctgtggcac atgtttaagc ctcctcctgc tttggagtgc cgccgttgcc atattaagtg
     2701 tcataaagat catatggaca aaaaggagga gattatagca ccttgcaaag tatattatga
     2761 tatttcaacg gcaaagaatc tgttattact agcaaattct acagaagagc agcagaagtg
     2821 ggttagtcgg ttggtgaaaa agatacctaa aaagccccca gctccagacc cttttgcccg
     2881 atcatctcct agaacttcaa tgaagataca gcaaaaccag tctattagac ggccaagtcg
     2941 acagcttgcc ccaaacaaac ctagctaaga tatccagcac agtggcggcc gctcgagtct
     3001 agagggcccg tttaaacccg ctgatcagcc tcgactgtgc cttctagttg ccagccatct
     3061 gttgtttgcc cctcccccgt gccttccttg accctggaag gtgccactcc cactgtcctt
     3121 tcctaataaa atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc tattctgggg
     3181 ggtggggtgg ggcaggacag caagggggag gattgggaag acaatagcag gcatgctggg
     3241 gatgcggtgg gctctatggc ttctgaggcg gaaagaacca gctggggctc tagggggtat
     3301 ccccacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg
     3361 accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc
     3421 gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga
     3481 tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt
     3541 gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat
     3601 agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta ttcttttgat
     3661 ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa
     3721 tttaacgcga attaattctg tggaatgtgt gtcagttagg gtgtggaaag tccccaggct
     3781 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc aggtgtggaa
     3841 agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa
     3901 ccatagtccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt
     3961 ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc gcctctgcct
     4021 ctgagctatt ccagaagtag tgaggaggct tttttggagg cctaggcttt tgcaaaaagc
     4081 tcccaaccat gaccgagtac aagcccacgg tgcgcctcgc cacccgcgac gacgtcccca
     4141 gggccgtacg caccctcgcc gccgcgttcg ccgactaccc cgccacgcgc cacaccgtcg
     4201 atccggaccg ccacatcgag cgggtcaccg agctgcaaga actcttcctc acgcgcgtcg
     4261 ggctcgacat cggcaaggtg tgggtcgcgg acgacggcgc cgcggtggcg gtctggacca
     4321 cgccggagag cgtcgaagcg ggggcggtgt tcgccgagat cggcccgcgc atggccgagt
     4381 tgagcggttc ccggctggcc gcgcagcaac agatggaagg cctcctggcg ccgcaccggc
     4441 ccaaggagcc cgcgtggttc ctggccaccg tcggcgtctc gcccgaccac cagggcaagg
     4501 gtctgggcag cgccgtcgtg ctccccggag tggaggcggc cgagcgcgcc ggggtgcccg
     4561 ccttcctgga gacctccgcg ccccgcaacc tccccttcta cgagcggctc ggcttcaccg
     4621 tcaccgccga cgtcgaggtg cccgaaggac cgcgcacctg gtgcatgacc cgcaagcccg
     4681 gtgcctgagc gggactctgg ggttcgaaat gaccgaccaa gcgacgccca acctgccatc
     4741 acgagatttc gattccaccg ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg
     4801 ggacgccggc tggatgatcc tccagcgcgg ggatctcatg ctggagttct tcgcccaccc
     4861 caacttgttt attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac
     4921 aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc
     4981 ttatcatgtc tgtataccgt cgacctctag ctagagcttg gcgtaatcat ggtcatagct
     5041 gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat
     5101 aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc
     5161 actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg
     5221 cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct
     5281 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     5341 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     5401 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     5461 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     5521 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     5581 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     5641 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     5701 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     5761 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     5821 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt
     5881 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     5941 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     6001 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     6061 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     6121 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     6181 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     6241 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     6301 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     6361 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     6421 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     6481 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     6541 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     6601 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     6661 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     6721 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     6781 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     6841 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     6901 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     6961 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     7021 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     7081 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     7141 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtc
//
