LOCUS       sequence-403652-        6025 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For construction of CFP-tagged human PCNA, the GFP coding sequence 
            in the pENeGFPCNAL2 vector was replaced by the ECFP coding sequence 
            from the pECFP-C1 vector (Clontech Laboratories, Inc., CA)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6025)
  AUTHORS   Zhang P, Rausch C, Hastert FD, Boneva B, Filatova A, Patil SJ, Nuber
            UA, Gao Y, Zhao X, Cardoso MC
  TITLE     Methyl-CpG binding domain protein 1 regulates localization and 
            activity of Tet1 in a CXXC3 domain-dependent manner.
  JOURNAL   Nucleic Acids Res. 2017 Apr 25. doi: 10.1093/nar/gkx281.
  PUBMED    28449087
REFERENCE   2  (bases 1 to 6025)
  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: "Nucleic 
            Acids Res. 2017 Apr 25. doi: 10.1093/nar/gkx281."
COMMENT     Sequence Label: pENeCFPCNAL2 pc922
FEATURES             Location/Qualifiers
     source          1..6025
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     66..85
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        66..84
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     enhancer        108..411
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        412..615
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     566..586
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     612..636
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     5'UTR           705..744
                     /label=HSV TK 5'-UTR
                     /note="5' untranslated region from the herpes simplex virus
                     thymidine kinase gene"
     CDS             780..800
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     regulatory      813..822
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(819..821,822..824,825..1535)
                     /codon_start=1
                     /product="enhanced CFP"
                     /label=ECFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(864..885)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1125..1144)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1472..1493
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     intron          2397..2969
                     /label=beta-globin intron
                     /note="intron from rabbit beta-globin gene"
     primer_bind     complement(3101..3120)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3166..3221
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3220..3239)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     rep_origin      3571..3706
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(3812..3829)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3983..4571)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4063..4082)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4742..5533,5534..5602))
                     /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     5365..5384
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5603..5707)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5775..5793
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5831..5853)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     5953..5972
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 agcccagtag taggttgagg ccgttgagca ccgccgccgc aaggaatggt ctggcttatc
       61 gaaattaata cgactcacta tagggagacc ggaattcgag ctcgccccgt tacataactt
      121 acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg
      181 acgtatgttc ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat
      241 ttacggtaaa ctgcccactt ggcagtacat caagtgtatc atatgccaag tacgccccct
      301 attgacgtca atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttatgg
      361 gactttccta cttggcagta catctacgta ttagtcatcg ctattaccat ggtgatgcgg
      421 ttttggcagt acatcaatgg gcgtggatag cggtttgact cacggggatt tccaagtctc
      481 caccccattg acgtcaatgg gagtttgttt tggcaccaaa atcaacggga ctttccaaaa
      541 tgtcgtaaca actccgcccc attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc
      601 tatataagca gagctcgttt agtgaaccgt cagatcgcct ggagacgcca tccacgctgt
      661 tttgacctcc atagaagaca ccgggaccga tccagcctcc gggggatctt ggtggcgtga
      721 aactcccgca cctcttcggc cagcgccttg tagaagcgcg tatggcttcg tggggatccc
      781 cgaagaagaa gcgcaaagta ctggtaccgg tcgccaccat ggtgagcaag ggcgaggagc
      841 tgttcaccgg ggtggtgccc atcctggtcg agctggacgg cgacgtaaac ggccacaagt
      901 tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg caagctgacc ctgaagttca
      961 tctgcaccac cggcaagctg cccgtgccct ggcccaccct cgtgaccacc ctgacctggg
     1021 gcgtgcagtg cttcagccgc taccccgacc acatgaagca gcacgacttc ttcaagtccg
     1081 ccatgcccga aggctacgtc caggagcgca ccatcttctt caaggacgac ggcaactaca
     1141 agacccgcgc cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc gagctgaagg
     1201 gcatcgactt caaggaggac ggcaacatcc tggggcacaa gctggagtac aactacatca
     1261 gccacaacgt ctatatcacc gccgacaagc agaagaacgg catcaaggcc aacttcaaga
     1321 tccgccacaa catcgaggac ggcagcgtgc agctcgccga ccactaccag cagaacaccc
     1381 ccatcggcga cggccccgtg ctgctgcccg acaaccacta cctgagcacc cagtccgccc
     1441 tgagcaaaga ccccaacgag aagcgcgatc acatggtcct gctggagttc gtgaccgccg
     1501 ccgggatcac tctcggcatg gacgagctgt acaagggcga agggcaaggg caagggcaag
     1561 ggccgggccg cggctacgcg tatcgatcca tgttcgaggc gcgcctggtc cagggctcca
     1621 tcctcaagaa ggtgttggag gcactcaagg acctcatcaa cgaggcctgc tgggatatta
     1681 gctccagcgg tgtaaacctg cagagcatgg actcgtccca cgtctctttg gtgcagctca
     1741 ccctgcggtc tgagggcttc gacacctacc gctgcgaccg caacctggcc atgggcgtga
     1801 acctcaccag tatgtccaaa atactaaaat gcgccggcaa tgaagatatc attacactaa
     1861 gggccgaaga taacgcggat accttggcgc tagtatttga agcaccaaac caggagaaag
     1921 tttcagacta tgaaatgaag ttgatggatt tagatgttga acaacttgga attccagaac
     1981 aggagtacag ctgtgtagta aagatgcctt ctggtgaatt tgcacgtata tgccgagatc
     2041 tcagccatat tggagatgct gttgtaattt cctgtgcaaa agacggagtg aaattttctg
     2101 caagtggaga acttggaaat ggaaacatta aattgtcaca gacaagtaat gtcgataaag
     2161 aggaggaagc tgttaccata gagatgaatg aaccagttca actaactttt gcactgaggt
     2221 acctgaactt ctttacaaaa gccactccac tctcttcaac ggtgacactc agtatgtctg
     2281 cagatgtacc ccttgttgta gagtataaaa ttgcggatat gggacactta aaatactact
     2341 tggctcccaa gatcgaggat gaagaaggat cttagtctag agctgagaac ttcagggtga
     2401 gtttggggac ccttgattgt tctttctttt tcgctattgt aaaattcatg ttatatggag
     2461 ggggcaaagt tttcagggtg ttgtttagaa tgggaagatg tcccttgtat caccatggac
     2521 cctcatgata attttgtttc tttcactttc tactctgttg acaaccattg tctcctctta
     2581 ttttcttttc attttctgta actttttcgt taaactttag cttgcatttg taacgaattt
     2641 ttaaattcac ttttgtttat ttgtcagatt gtaagtactt tctctaatca cttttttttc
     2701 aaggcaatca gggtatatta tattgtactt cagcacagtt ttagagaaca attgttataa
     2761 ttaaatgata aggtagaata tttctgcata taaattctgg ctggcgtgga aatattctta
     2821 ttggtagaaa caactacacc ctggtcatca tcctgccttt ctctttatgg ttacaatgat
     2881 atacactgtt tgagatgagg ataaaatact ctgagtccaa accgggcccc tctgctaacc
     2941 atgttcatgc cttcttctct ttcctacagc tcctgggcaa cgtgctggtt gttgtgctgt
     3001 ctcatcattt tggcaaagaa ttcactcctc aggtgcaggc tgcctatcag aaggtggtgg
     3061 ctggtgtggc caatgccctg gctcacaaat accactgaga tctttttccc tctgccaaaa
     3121 attatgggga catcatgaag ccccttgagc atctgacttc tggctaataa aggaaattta
     3181 ttttcattgc aatagtgtgt tggaattttt tgtgtctctc actcggaagg acatatggga
     3241 gggcaaatca tttaaaacat cagaatgagt atttggttta gagtttggca acatatgccc
     3301 atatgctggc tgccatgaac aaaggttggc tataaagagg tcatcagtat atgaaacagc
     3361 cccctgctgt ccattcctta ttccatagaa aagccttgac ttgaggttag atttttttta
     3421 tattttgttt tgtgttattt ttttctttaa catccctaaa attttcctta catgttttac
     3481 tagccagatt tttcctcctc tcctgactac tcccagtcat agctgtccct cttctcttat
     3541 ggagatcccg gagctttttg caaaagccta ggcctccaaa aaagcctctt cactacttct
     3601 ggaatagctc agaggccgag gcggcctcgg cctctgcata aataaaaaaa attagtcagc
     3661 catggggcgg agaatgggcg gaactgggcg gagttagggg cgggatgggc ggagttaggg
     3721 gcgggactat ggttgctgac taattgctgc attaatgaat cggccaacgc gcggggagag
     3781 gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg
     3841 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat
     3901 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     3961 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     4021 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     4081 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     4141 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     4201 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     4261 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     4321 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     4381 cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct
     4441 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     4501 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     4561 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa
     4621 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt
     4681 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca
     4741 gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca
     4801 tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc
     4861 ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa
     4921 accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc
     4981 agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca
     5041 acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat
     5101 tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag
     5161 cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac
     5221 tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt
     5281 ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt
     5341 gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc
     5401 tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat
     5461 ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca
     5521 gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga
     5581 cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg
     5641 gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg
     5701 ttccgcgcac atttccccga aaagtgccac ctgacgtcta agaaaccatt attatcatga
     5761 cattaaccta taaaaatagg cgtatcacga ggccctttcg tctcgcgcgt ttcggtgatg
     5821 acggtgaaaa cctctgacac atgcagctcc cggagacggt cacagcttgt ctgtaagcgg
     5881 atgccgggag cagacaagcc cgtcagggcg cgtcagcggg tgttggcggg tgtcggggct
     5941 ggcttaacta tgcggcatca gagcagattg tactgagagt gcaccattcg acgctctccc
     6001 ttatgcgact cctgcattag gaagc
//
