LOCUS       sequence-351714-        7687 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of PKC-delta fused to EGFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7687)
  AUTHORS   Suzuki S, Nakamura A, Hatano Y, Yoshikawa M, Yoshii T, Sawada S, 
            Atsuta-Tsunoda K, Aoki K, Tsukiji S
  TITLE     A chemogenetic platform for controlling plasma membrane signaling 
            and synthetic signal oscillation.
  JOURNAL   Cell Chem Biol. 2022 Sep 15;29(9):1446-1464.e10. doi: 
            10.1016/j.chembiol.2022.06.005. Epub 2022 Jul 13.
  PUBMED    35835118
REFERENCE   2  (bases 1 to 7687)
  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: 
            "10.1016/j.chembiol.2022.06.005"; journalName: "Cell Chem Biol"; 
            date: "2022-09-15- 15"; volume: "29"; issue: "9"; pages: 
            "1446-1464.e10"
COMMENT     Sequence Label: pCAGGS-PKCdelta-EGFP
FEATURES             Location/Qualifiers
     source          1..7687
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     intron          117..1133
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1057..1079
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1141..1160
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      1211..1220
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(3317..3319,3320..3322,3323..4036)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(3317..3319,3320..3322,3323..4033)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3362..3383)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3623..3642)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3970..3991
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(4055..4075)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(4055..4073)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4094..4110)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4212..4231)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    4277..4332
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(4331..4350)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(4693..4709)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    4717..4733
                     /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(4741..4747,4748..4765,4766..4771))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4786..4807
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        4866..5061
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      4912..5047
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    5067..5201
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5104..5123)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5158..5177
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5269..5286)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5440..6028)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5520..5539)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6199..6990,6991..7059))
                     /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     6822..6841
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7060..7164)
                     /gene="bla"
                     /label=AmpR promoter
     enhancer        7195..7574
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 gggcgaggcg gagaggtgcg gcggcagcca atcagagcgg cgcgctccga aagtttcctt
       61 ttatggcgag gcggcggcgg cggcggccct ataaaaagcg aagcgcgcgg cgggcgggag
      121 tcgctgcgcg ctgccttcgc cccgtgcccc gctccgccgc cgcctcgcgc cgcccgcccc
      181 ggctctgact gaccgcgtta ctcccacagg tgagcgggcg ggacggccct tctcctccgg
      241 gctgtaatta gcgcttggtt taatgacggc ttgtttcttt tctgtggctg cgtgaaagcc
      301 ttgaggggct ccgggagggc cctttgtgcg gggggagcgg ctcggggggt gcgtgcgtgt
      361 gtgtgtgcgt ggggagcgcc gcgtgcggct ccgcgctgcc cggcggctgt gagcgctgcg
      421 ggcgcggcgc ggggctttgt gcgctccgca gtgtgcgcga ggggagcgcg gccgggggcg
      481 gtgccccgcg gtgcgggggg ggctgcgagg ggaacaaagg ctgcgtgcgg ggtgtgtgcg
      541 tgggggggtg agcagggggt gtgggcgcgt cggtcgggct gcaacccccc ctgcaccccc
      601 ctccccgagt tgctgagcac ggcccggctt cgggtgcggg gctccgtacg gggcgtggcg
      661 cggggctcgc cgtgccgggc ggggggtggc ggcaggtggg ggtgccgggc ggggcggggc
      721 cgcctcgggc cggggagggc tcgggggagg ggcgcggcgg cccccggagc gccggcggct
      781 gtcgaggcgc ggcgagccgc agccattgcc ttttatggta atcgtgcgag agggcgcagg
      841 gacttccttt gtcccaaatc tgtgcggagc cgaaatctgg gaggcgccgc cgcaccccct
      901 ctagcgggcg cggggcgaag cggtgcggcg ccggcaggaa ggaaatgggc ggggagggcc
      961 ttcgtgcgtc gccgcgccgc cgtccccttc tccctctcca gcctcggggc tgtccgcggg
     1021 gggacggctg ccttcggggg ggacggggca gggcggggtt cggcttctgg cgtgtgaccg
     1081 gcggctctag agcctctgct aaccatgttc atgccttctt ctttttccta cagctcctgg
     1141 gcaacgtgct ggttattgtg ctgtctcatc attttggcaa agaattcata gtagctagca
     1201 gttgctcgag gccaccatgg cgccgttcct gcgcatcgcc ttcaactcct atgagctggg
     1261 ctccctgcag gccgaggacg aggcgaacca gcccttctgt gccgtgaaga tgaaggaggc
     1321 gctcagcaca gagcgtggga aaacactggt gcagaagaag ccgaccatgt atcctgagtg
     1381 gaagtcgacg ttcgatgccc acatctatga ggggcgcgtc atccagattg tgctaatgcg
     1441 ggcagcagag gagccagtgt ctgaggtgac cgtgggtgtg tcggtgctgg ccgagcgctg
     1501 caagaagaac aatggcaagg ctgagttctg gctggacctg cagcctcagg ccaaggtgtt
     1561 gatgtctgtt cagtatttcc tggaggacgt ggattgcaaa cagtctatgc gcagtgagga
     1621 cgaggccaag ttcccaacga tgaaccgccg cggagccatc aaacaggcca aaatccacta
     1681 catcaagaac catgagttta tcgccacctt ctttgggcaa cccaccttct gttctgtgtg
     1741 caaagacttt gtctggggcc tcaacaagca aggctacaaa tgcaggcaat gtaacgctgc
     1801 catccacaag aaatgcatcg acaagatcat cggcagatgc actggcaccg cggccaacag
     1861 ccgggacact atattccaga aagaacgctt caacatcgac atgccgcacc gcttcaaggt
     1921 tcacaactac atgagcccca ccttctgtga ccactgcggc agcctgctct ggggactggt
     1981 gaagcaggga ttaaagtgtg aagactgcgg catgaatgtg caccataaat gccgggagaa
     2041 ggtggccaac ctctgcggca tcaaccagaa gcttttggct gaggccttga accaagtcac
     2101 ccagagagcc tcccggagat cagactcagc ctcctcagag cctgttggga tatatcaggg
     2161 tttcgagaag aagaccggag ttgctgggga ggacatgcaa gacaacagtg ggacctacgg
     2221 caagatctgg gagggcagca gcaagtgcaa catcaacaac ttcatcttcc acaaggtcct
     2281 gggcaaaggc agcttcggga aggtgctgct tggagagctg aagggcagag gagagtactt
     2341 tgccatcaag gccctcaaga aggatgtggt cctgatcgac gacgacgtgg agtgcaccat
     2401 ggttgagaag cgggtgctga cacttgccgc agagaatccc tttctcaccc acctcatctg
     2461 caccttccag accaaggacc acctgttctt tgtgatggag ttcctcaacg ggggggacct
     2521 gatgtaccac atccaggaca aaggccgctt tgaactctac cgtgccacgt tttatgccgc
     2581 tgagataatg tgtggactgc agtttctaca cagcaagggc atcatttaca gggacctcaa
     2641 actggacaat gtgctgttgg accgggatgg ccacatcaag attgccgact ttgggatgtg
     2701 caaagagaac atattcgggg agagccgggc cagcaccttc tgcggcaccc ctgactatat
     2761 cgcccctgag atcctacagg gcctgaagta cacattctct gtggactggt ggtctttcgg
     2821 ggtccttctg tacgagatgc tcattggcca gtcccccttc catggtgatg atgaggatga
     2881 actcttcgag tccatccgtg tggacacgcc acattatccc cgctggatca ccaaggagtc
     2941 caaggacatc ctggagaagc tctttgaaag ggaaccaacc aagaggctgg gagtgacggg
     3001 aaacatcaaa atccacccct tcttcaagac cataaactgg actctgctgg aaaagcggag
     3061 gttggagcca cccttcaggc ccaaagtgaa gtcacccaga gactacagta actttgacca
     3121 ggagttcctg aacgagaagg cgcgcctctc ctacagcgac aagaacctca tcgactccat
     3181 ggaccagtct gcattcgctg gcttctcctt tgtgaacccc aaattcgagc acctcctgga
     3241 agatgcggcc gctggtagtg gtgctggctc tggtgctggt agtggcgctg gttccggtgc
     3301 tggctcttcg gccgctatgg tgagcaaggg cgaggagctg ttcaccgggg tggtgcccat
     3361 cctggtcgag ctggacggcg acgtaaacgg ccacaagttc agcgtgtccg gcgagggcga
     3421 gggcgatgcc acctacggca agctgaccct gaagttcatc tgcaccaccg gcaagctgcc
     3481 cgtgccctgg cccaccctcg tgaccaccct gacctacggc gtgcagtgct tcagccgcta
     3541 ccccgaccac atgaagcagc acgacttctt caagtccgcc atgcccgaag gctacgtcca
     3601 ggagcgcacc atcttcttca aggacgacgg caactacaag acccgcgccg aggtgaagtt
     3661 cgagggcgac accctggtga accgcatcga gctgaagggc atcgacttca aggaggacgg
     3721 caacatcctg gggcacaagc tggagtacaa ctacaacagc cacaacgtct atatcatggc
     3781 cgacaagcag aagaacggca tcaaggtgaa cttcaagatc cgccacaaca tcgaggacgg
     3841 cagcgtgcag ctcgccgacc actaccagca gaacaccccc atcggcgacg gccccgtgct
     3901 gctgcccgac aaccactacc tgagcaccca gtccgccctg agcaaagacc ccaacgagaa
     3961 gcgcgatcac atggtcctgc tggagttcgt gaccgccgcc gggatcactc tcggcatgga
     4021 cgagctgtac aagtaggagc tccagctttt gttcccttta gtgagggtta attgcgcgct
     4081 tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgcta attcactcct
     4141 caggtgcagg ctgcctatca gaaggtggtg gctggtgtgg ccaatgccct ggctcacaaa
     4201 taccactgag atctttttcc ctctgccaaa aattatgggg acatcatgaa gccccttgag
     4261 catctgactt ctggctaata aaggaaattt attttcattg caatagtgtg ttggaatttt
     4321 ttgtgtctct cactcggaag gacatatggg agggcaaatc atttaaaaca tcagaatgag
     4381 tatttggttt agagtttggc aacatatgcc catatgctgg ctgccatgaa caaaggttgg
     4441 ctataaagag gtcatcagta tatgaaacag ccccctgctg tccattcctt attccataga
     4501 aaagccttga cttgaggtta gatttttttt atattttgtt ttgtgttatt tttttcttta
     4561 acatccctaa aattttcctt acatgtttta ctagccagat ttttcctcct ctcctgacta
     4621 ctcccagtca tagctgtccc tcttctctta tggagatccc tcgacctgca gcccaagctt
     4681 ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca
     4741 caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact
     4801 cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagcg
     4861 gatccgcatc tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc
     4921 ctaactccgc ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat
     4981 gcagaggccg aggccgcctc ggcctctgag ctattccaga agtagtgagg aggctttttt
     5041 ggaggcctag gcttttgcaa aaagctaact tgtttattgc agcttataat ggttacaaat
     5101 aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg
     5161 gtttgtccaa actcatcaat gtatcttatc atgtctggat ccgctgcatt aatgaatcgg
     5221 ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga
     5281 ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat
     5341 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca
     5401 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc
     5461 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata
     5521 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc
     5581 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc
     5641 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga
     5701 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc
     5761 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag
     5821 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag
     5881 aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag
     5941 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca
     6001 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga
     6061 cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat
     6121 cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga
     6181 gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg
     6241 tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga
     6301 gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc
     6361 agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac
     6421 tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc
     6481 agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc
     6541 gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc
     6601 catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt
     6661 ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc
     6721 atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg
     6781 tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag
     6841 cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat
     6901 cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc
     6961 atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa
     7021 aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta
     7081 ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa
     7141 aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg ggtcgacatt
     7201 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
     7261 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
     7321 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
     7381 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
     7441 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
     7501 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
     7561 tcgctattac catggtcgag gtgagcccca cgttctgctt cactctcccc atctcccccc
     7621 cctccccacc cccaattttg tatttattta ttttttaatt attttgtgca gcgatggggg
     7681 cgggggg
//
