LOCUS       sequence-347292-        7727 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 7727)
  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 7727)
  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..7727
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        4..383
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        386..661
                     /label=chicken beta-actin promoter
     intron          662..1670
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1594..1616
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     regulatory      1748..1757
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(3854..3856,3857..3859,3860..4573)
                     /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(3854..3856,3857..3859,3860..4570)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3899..3920)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4160..4179)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4507..4528
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(4592..4612)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(4592..4610)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4631..4647)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4749..4768)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    4814..4869
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(4868..4887)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(5230..5246)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    5254..5270
                     /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(5278..5284,5285..5302,5303..5308))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5323..5344
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        5403..5598
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      5449..5584
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    5604..5738
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5641..5660)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5695..5714
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5806..5823)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5977..6565)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6057..6076)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6736..7527,7528..7596))
                     /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     7359..7378
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7597..7701)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgacattg attattgact agttattaat agtaatcaat tacggggtca ttagttcata
       61 gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc
      121 ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag
      181 ggactttcca ttgacgtcaa tgggtggact atttacggta aactgcccac ttggcagtac
      241 atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg
      301 cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg
      361 tattagtcat cgctattacc atgggtcgag gtgagcccca cgttctgctt cactctcccc
      421 atctcccccc cctccccacc cccaattttg tatttattta ttttttaatt attttgtgca
      481 gcgatggggg cggggggggg gggggcgcgc gccaggcggg gcggggcggg ggcgagggcg
      541 gggcggggcg aggcggagag gtgcggcggc agccaatcag agcggcgcgc tccgaaagtt
      601 tccttttatg gcgaggcggc ggcggcggcg gccctataaa aagcgaagcg cgcggcgggc
      661 gggagtcgct gcgttgcctt cgccccgtgc cccgctccgc gccgcctcgc gccgcccgcc
      721 ccggctctga ctgaccgcgt tactcccaca ggtgagcggg cgggacggcc cttctcctcc
      781 gggctgtaat tagcgcttgg tttaatgacg gctcgtttct tttctgtggc tgcgtgaaag
      841 ccttaaaggg ctccgggagg gccctttgtg cgggggggag cggctcgggg ggtgcgtgcg
      901 tgtgtgtgtg cgtggggagc gccgcgtgcg gcccgcgctg cccggcggct gtgagcgctg
      961 cgggcgcggc gcggggcttt gtgcgctccg cgtgtgcgcg aggggagcgc ggccgggggc
     1021 ggtgccccgc ggtgcggggg ggctgcgagg ggaacaaagg ctgcgtgcgg ggtgtgtgcg
     1081 tgggggggtg agcagggggt gtgggcgcgg cggtcgggct gtaacccccc cctgcacccc
     1141 cctccccgag ttgctgagca cggcccggct tcgggtgcgg ggctccgtgc ggggcgtggc
     1201 gcggggctcg ccgtgccggg cggggggtgg cggcaggtgg gggtgccggg cggggcgggg
     1261 ccgcctcggg ccggggaggg ctcgggggag gggcgcggcg gccccggagc gccggcggct
     1321 gtcgaggcgc ggcgagccgc agccattgcc ttttatggta atcgtgcgag agggcgcagg
     1381 gacttccttt gtcccaaatc tggcggagcc gaaatctggg aggcgccgcc gcaccccctc
     1441 tagcgggcgc gggcgaagcg gtgcggcgcc ggcaggaagg aaatgggcgg ggagggcctt
     1501 cgtgcgtcgc cgcgccgccg tccccttctc catctccagc ctcggggctg ccgcaggggg
     1561 acggctgcct tcggggggga cggggcaggg cggggttcgg cttctggcgt gtgaccggcg
     1621 gctctagagc ctctgctaac catgttcatg ccttcttctt tttcctacag ctcctgggca
     1681 acgtgctggt tgttgtgctg tctcatcatt ttggcaaaga attcatagta gctagcagtt
     1741 gctcgaggcc accatggcgc cgttcctgcg catcgccttc aactcctatg agctgggctc
     1801 cctgcaggcc gaggacgagg cgaaccagcc cttctgtgcc gtgaagatga aggaggcgct
     1861 cagcacagag cgtgggaaaa cactggtgca gaagaagccg accatgtatc ctgagtggaa
     1921 gtcgacgttc gacgcccaca tctatgaggg gcgcgtcatc cagattgtgc taatgcgggc
     1981 agcagaggag ccagtgtctg aggtgaccgt gggtgtgtcg gtgctggccg agcgctgcaa
     2041 gaagaacaat ggcaaggctg agttctggct ggacctgcag cctcaggcca aggtgttgat
     2101 gtctgttcag tatttcctgg aggacgtgga ttgcaaacag tctatgcgca gtgaggacga
     2161 ggccaagttc ccaacgatga accgccgcgg agccatcaaa caggccaaaa tccactacat
     2221 caagaaccat gagtttatcg ccaccttctt tgggcaaccc accttctgtt ctgtgtgcaa
     2281 agactttgtc tggggcctca acaagcaagg ctacaaatgc aggcaatgta acgctgccat
     2341 ccacaagaaa tgcatcgaca agatcatcgg cagatgcact ggcaccgcgg ccaacagccg
     2401 ggacactata ttccagaaag aacgcttcaa catcgacatg ccgcaccgct tcaaggttca
     2461 caactacatg agccccacct tctgtgacca ctgcggcagc ctgctctggg gactggtgaa
     2521 gcagggatta aagtgtgaag actgcggcat gaatgtgcac cataaatgcc gggagaaggt
     2581 ggccaacctc tgcggcatca accagaagct tttggctgag gccttgaacc aagtcaccca
     2641 gagagcctcc cggagatcag actcagcctc ctcagagcct gttgggatat atcagggttt
     2701 cgagaagaag accggagttg ctggggagga catgcaagac aacagtggga cctacggcaa
     2761 gatctgggag ggcagcagca agtgcaacat caacaacttc atcttccaca aggtcctggg
     2821 caaaggcagc ttcgggaagg tgctgcttgg agagctgaag ggcagaggag agtactttgc
     2881 catcaaggcc ctcaagaagg atgtggtcct gatcgacgac gacgtggagt gcaccatggt
     2941 tgagaagcgg gtgctgacac ttgccgcaga gaatcccttt ctcacccacc tcatctgcac
     3001 cttccagacc aaggaccacc tgttctttgt gatggagttc ctcaacgggg gggacctgat
     3061 gtaccacatc caggacaaag gccgctttga actctaccgt gccacgtttt atgccgctga
     3121 gataatgtgt ggactgcagt ttctacacag caagggcatc atttacaggg acctcaaact
     3181 ggacaatgtg ctgttggacc gggatggcca catcaagatt gccgactttg ggatgtgcaa
     3241 agagaacata ttcggggaga gccgggccag caccttctgc ggcacccctg actatatcgc
     3301 ccctgagatc ctacagggcc tgaagtacac attctctgtg gactggtggt ctttcggggt
     3361 ccttctgtac gagatgctca ttggccagtc ccccttccat ggtgatgatg aggatgaact
     3421 cttcgagtcc atccgtgtgg acacgccaca ttatccccgc tggatcacca aggagtccaa
     3481 ggacatcctg gagaagctct ttgaaaggga accaaccaag aggctgggag tgacgggaaa
     3541 catcaaaatc caccccttct tcaagaccat aaactggact ctgctggaaa agcggaggtt
     3601 ggagccaccc ttcaggccca aagtgaagtc acccagagac tacagtaact ttgaccagga
     3661 gttcctgaac gagaaggcgc gcctctccta cagcgacaag aacctcatcg actccatgga
     3721 ccagtctgca ttcgctggct tctcctttgt gaaccccaaa ttcgagcacc tcctggaaga
     3781 tgcggccgct ggtagtggtg ctggctctgg tgctggtagt ggcgctggtt ccggtgctgg
     3841 ctcttcggcc gctatggtga gcaagggcga ggagctgttc accggggtgg tgcccatcct
     3901 ggtcgagctg gacggcgacg taaacggcca caagttcagc gtgtccggcg agggcgaggg
     3961 cgatgccacc tacggcaagc tgaccctgaa gttcatctgc accaccggca agctgcccgt
     4021 gccctggccc accctcgtga ccaccctgac ctacggcgtg cagtgcttca gccgctaccc
     4081 cgaccacatg aagcagcacg acttcttcaa gtccgccatg cccgaaggct acgtccagga
     4141 gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg tgaagttcga
     4201 gggcgacacc ctggtgaacc gcatcgagct gaagggcatc gacttcaagg aggacggcaa
     4261 catcctgggg cacaagctgg agtacaacta caacagccac aacgtctata tcatggccga
     4321 caagcagaag aacggcatca aggtgaactt caagatccgc cacaacatcg aggacggcag
     4381 cgtgcagctc gccgaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct
     4441 gcccgacaac cactacctga gcacccagtc cgccctgagc aaagacccca acgagaagcg
     4501 cgatcacatg gtcctgctgg agttcgtgac cgccgccggg atcactctcg gcatggacga
     4561 gctgtacaag taggagctcc agcttttgtt ccctttagtg agggttaatt gcgcgcttgg
     4621 cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctaatt cactcctcag
     4681 gtgcaggctg cctatcagaa ggtggtggct ggtgtggcca atgccctggc tcacaaatac
     4741 cactgagatc tttttccctc tgccaaaaat tatggggaca tcatgaagcc ccttgagcat
     4801 ctgacttctg gctaataaag gaaatttatt ttcattgcaa tagtgtgttg gaattttttg
     4861 tgtctctcac tcggaaggac atatgggagg gcaaatcatt taaaacatca gaatgagtat
     4921 ttggtttaga gtttggcaac atatgcccat atgctggctg ccatgaacaa aggttggcta
     4981 taaagaggtc atcagtatat gaaacagccc cctgctgtcc attccttatt ccatagaaaa
     5041 gccttgactt gaggttagat tttttttata ttttgttttg tgttattttt ttctttaaca
     5101 tccctaaaat tttccttaca tgttttacta gccagatttt tcctcctctc ctgactactc
     5161 ccagtcatag ctgtccctct tctcttatgg agatccctcg acctgcagcc caagcttggc
     5221 gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa
     5281 catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac
     5341 attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagcggat
     5401 ccgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta
     5461 actccgccca gttccgccca ttctccgccc catggctgac taattttttt tatttatgca
     5521 gaggccgagg ccgcctcggc ctctgagcta ttccagaagt agtgaggagg cttttttgga
     5581 ggcctaggct tttgcaaaaa gctaacttgt ttattgcagc ttataatggt tacaaataaa
     5641 gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt
     5701 tgtccaaact catcaatgta tcttatcatg tctggatccg ctgcattaat gaatcggcca
     5761 acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc
     5821 gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg
     5881 gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa
     5941 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     6001 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     6061 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     6121 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc aatgctcacg
     6181 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     6241 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     6301 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     6361 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaaggac
     6421 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     6481 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     6541 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     6601 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
     6661 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
     6721 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     6781 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     6841 cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga
     6901 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     6961 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     7021 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
     7081 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     7141 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     7201 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     7261 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     7321 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     7381 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     7441 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     7501 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     7561 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
     7621 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     7681 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctg
//
