LOCUS       sequence-9820-ex        6814 bp    DNA     circular SYN 14-APR-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6814)
  AUTHORS   Oancea E, Teruel MN, Quest AF, Meyer T
  TITLE     Green fluorescent protein (GFP)-tagged cysteine-rich domains from 
            protein kinase C as fluorescent indicators for diacylglycerol 
            signaling in living cells.
  JOURNAL   J Cell Biol. 1998 Feb 9. 140(3):485-98.
  PUBMED    9456311
REFERENCE   2  (bases 1 to 6814)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "J Cell 
            Biol. 1998 Feb 9. 140(3):485-98."
COMMENT     Sequence Label: GFP-N2-PKCgamma
FEATURES             Location/Qualifiers
     source          1..6814
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      2754..2763
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2760..2762,2763..2765,2766..3479)
                     /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(2760..2762,2763..2765,2766..3476)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2805..2826)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3066..3085)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3413..3434
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    3602..3723
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3639..3658)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3693..3712
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3730..4185)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3867..3888)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4079..4098
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4212..4316
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4318..4675
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4318..4509
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4341..4361)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4526..4661
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4710..5504
                     /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             4710..5504
                     /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(4764..4783)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5374..5393
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5692..5711)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5736..5783
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6112..6700
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6601..6620
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggactcag atctatggcg ggtctgggtc ctggcggggg cgactcagaa gggggacccc
      661 gacccctgtt ttgcagaaag ggggcgctga ggcagaaggt ggtccacgag gtgaagagcc
      721 acaagttcac cgctcgtttc ttcaagcagc caaccttctg cagtcactgt accgacttca
      781 tctggggcat tggaaagcag ggcctgcaat gtcaagtctg cagctttgtg gttcaccgcc
      841 gatgccacga atttgtgacc ttcgagtgtc caggcgctgg aaagggcccc cagacggacg
      901 accctcgcaa caagcacaag ttccgtctgc acagctacag cagtcccacc ttctgcgacc
      961 actgtggttc cctcctctac gggctggtgc accagggcat gaaatgttcc tgttgcgaaa
     1021 tgaatgtgca ccgacgctgt gtgcgcagcg tgccctccct ttgcggcgtg gaccatacag
     1081 agcgccgtgg acgtctgcaa ctggaaatcc gggctcccac atcagatgag atccatatta
     1141 ctgtgggtga ggcccggaac ctcattccta tggaccccaa tggcctgtct gatccctatg
     1201 tgaaactgaa gctcatcccg gaccctcgga acctgacaaa acagaagaca aagaccgtga
     1261 aagccacact gaatcccgtg tggaacgaga ccttcgtgtt caacctgaag ccgggggatg
     1321 tggagcgccg gctcagtgtg gaggtgtggg attgggatag gacatcccga aatgacttca
     1381 tgggtgccat gtcctttggt gtctcagagc tactcaaggc tcctgtggat ggatggtaca
     1441 agttactgaa ccaggaggag ggcgagtatt acaatgtacc ggtggccgat gctgacaact
     1501 gcagcctcct ccagaagttt gaggcctgta attacccctt ggaattgtat gagagagtgc
     1561 ggatgggccc ctcttcctct cccattcctt ctccatcccc cagtcccacg gactccaaga
     1621 gatgcttctt cggtgccagc ccaggacgcc tgcatatctc tgacttcagc ttcctcatgg
     1681 ttctagggaa aggcagtttt gggaaggtga tgctggcaga gcgcagagga tccgatgaac
     1741 tctatgccat caagatactg aaaaaagacg tcattgtcca ggatgatgat gtagactgca
     1801 cccttgtgga gaagcgtgtg ctggcattgg gaggccgagg tcctggaggc cggccacact
     1861 ttctcacaca acttcattcc acctttcaga ctccggaccg cctgtatttt gtgatggagt
     1921 acgtcactgg gggcgattta atgtaccaca ttcagcaact gggcaagttt aaggagcccc
     1981 acgcagcatt ctatgccgcg gaaatcgcca taggcctctt cttccttcac aaccagggca
     2041 tcatctacag ggacctcaag ttggataatg tgatgctgga tgctgaagga cacatcaaga
     2101 tcacagactt cggcatgtgt aaagagaatg tcttccctgg gtccacaacc cgcaccttct
     2161 gtgggacccc agactacata gcacctgaga tcattgccta tcagccctat gggaagtctg
     2221 tcgactggtg gtcctttgga gtcctgctgt atgagatgtt ggcaggacag ccaccctttg
     2281 atggggaaga tgaggaggag ctgtttcaag ccatcatgga acaaactgtc acctatccca
     2341 agtcactttc ccgggaagct gtggccatct gcaaggggtt cctgaccaag cacccaggaa
     2401 agcgcctggg ctcagggcca gatggggaac ccaccatccg ggctcatggc tttttccgtt
     2461 ggatcgattg ggagaggttg gagagactgg aaattgcgcc tccttttaga ccacgtccgt
     2521 gtggccgcag cggcgaaaac tttgacaagt tcttcacgcg ggcagcgcca gccttgaccc
     2581 cgccagaccg cttggtccta gccagcatcg accaagctga tttccagggc tttacttatg
     2641 tgaacccgga cttcgtgcac ccagatgccc gcagccccac aagccctgtg cctgtgcccg
     2701 tcatggaatt ctgcagtcga cggtaccgcg ggcccgggat ccaccggccg gtcgccacca
     2761 tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc catcctggtc gagctggacg
     2821 gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg
     2881 gcaagctgac cctgaagttc atctgcacca ccggcaagct gcccgtgccc tggcccaccc
     2941 tcgtgaccac cctgacctac ggcgtgcagt gcttcagccg ctaccccgac cacatgaagc
     3001 agcacgactt cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc accatcttct
     3061 tcaaggacga cggcaactac aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg
     3121 tgaaccgcat cgagctgaag ggcatcgact tcaaggagga cggcaacatc ctggggcaca
     3181 agctggagta caactacaac agccacaacg tctatatcat ggccgacaag cagaagaacg
     3241 gcatcaaggt gaacttcaag atccgccaca acatcgagga cggcagcgtg cagctcgccg
     3301 accactacca gcagaacacc cccatcggcg acggccccgt gctgctgccc gacaaccact
     3361 acctgagcac ccagtccgcc ctgagcaaag accccaacga gaagcgcgat cacatggtcc
     3421 tgctggagtt cgtgaccgcc gccgggatca ctctcggcat ggacgagctg tacaagtaaa
     3481 gcggccgcga ctctagatca taatcagcca taccacattt gtagaggttt tacttgcttt
     3541 aaaaaacctc ccacacctcc ccctgaacct gaaacataaa atgaatgcaa ttgttgttgt
     3601 taacttgttt attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac
     3661 aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc
     3721 ttaaggcgta aattgtaagc gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa
     3781 tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat cccttataaa tcaaaagaat
     3841 agaccgagat agggttgagt gttgttccag tttggaacaa gagtccacta ttaaagaacg
     3901 tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac
     3961 catcacccta atcaagtttt ttggggtcga ggtgccgtaa agcactaaat cggaacccta
     4021 aagggagccc ccgatttaga gcttgacggg gaaagccggc gaacgtggcg agaaaggaag
     4081 ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg
     4141 taaccaccac acccgccgcg cttaatgcgc cgctacaggg cgcgtcaggt ggcacttttc
     4201 ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc
     4261 cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtcctg
     4321 aggcggaaag aaccagctgt ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc
     4381 cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca ggtgtggaaa
     4441 gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac
     4501 catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc
     4561 tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg cctcggcctc
     4621 tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt gcaaagatcg
     4681 atcaagagac aggatgagga tcgtttcgca tgattgaaca agatggattg cacgcaggtt
     4741 ctccggccgc ttgggtggag aggctattcg gctatgactg ggcacaacag acaatcggct
     4801 gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg cccggttctt tttgtcaaga
     4861 ccgacctgtc cggtgccctg aatgaactgc aagacgaggc agcgcggcta tcgtggctgg
     4921 ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg ggaagggact
     4981 ggctgctatt gggcgaagtg ccggggcagg atctcctgtc atctcacctt gctcctgccg
     5041 agaaagtatc catcatggct gatgcaatgc ggcggctgca tacgcttgat ccggctacct
     5101 gcccattcga ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg atggaagccg
     5161 gtcttgtcga tcaggatgat ctggacgaag agcatcaggg gctcgcgcca gccgaactgt
     5221 tcgccaggct caaggcgagc atgcccgacg gcgaggatct cgtcgtgacc catggcgatg
     5281 cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc tggattcatc gactgtggcc
     5341 ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc tacccgtgat attgctgaag
     5401 agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt
     5461 cgcagcgcat cgccttctat cgccttcttg acgagttctt ctgagcggga ctctggggtt
     5521 cgaaatgacc gaccaagcga cgcccaacct gccatcacga gatttcgatt ccaccgccgc
     5581 cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca
     5641 gcgcggggat ctcatgctgg agttcttcgc ccaccctagg gggaggctaa ctgaaacacg
     5701 gaaggagaca ataccggaag gaacccgcgc tatgacggca ataaaaagac agaataaaac
     5761 gcacggtgtt gggtcgtttg ttcataaacg cggggttcgg tcccagggct ggcactctgt
     5821 cgatacccca ccgagacccc attggggcca atacgcccgc gtttcttcct tttccccacc
     5881 ccacccccca agttcgggtg aaggcccagg gctcgcagcc aacgtcgggg cggcaggccc
     5941 tgccatagcc tcaggttact catatatact ttagattgat ttaaaacttc atttttaatt
     6001 taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga
     6061 gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc
     6121 tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt
     6181 ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc
     6241 gcagatacca aatactgtcc ttctagtgta gccgtagtta ggccaccact tcaagaactc
     6301 tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg
     6361 cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg
     6421 gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga
     6481 actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc
     6541 ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg
     6601 gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg
     6661 atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt
     6721 tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc
     6781 tgattctgtg gataaccgta ttaccgccat gcat
//
