LOCUS       sequence-66618-e        6096 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  expresses mutant Parkin with YFP tag in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6096)
  AUTHORS   Lazarou M, Narendra DP, Jin SM, Tekle E, Banerjee S, Youle RJ
  TITLE     PINK1 drives Parkin self-association and HECT-like E3 activity 
            upstream of mitochondrial binding.
  JOURNAL   J Cell Biol. 2013 Jan 21;200(2):163-72. doi: 10.1083/jcb.201210111. 
            Epub 2013 Jan 14.
  PUBMED    23319602
REFERENCE   2  (bases 1 to 6096)
  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: 
            "10.1083/jcb.201210111"; journalName: "J Cell Biol"; date: 
            "2013-01-21- 21"; volume: "200"; issue: "2"; pages: "163-72"
COMMENT     Sequence Label: YFP-Parkin C431N
FEATURES             Location/Qualifiers
     source          1..6096
                     /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      607..616
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(613..615,616..618,619..1329)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(613..615,616..618,619..1329)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(658..679)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(919..938)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1266..1287
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2884..3005
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2921..2940)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2975..2994
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3012..3467)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3149..3170)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3361..3380
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3494..3598
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3600..3957
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3600..3791
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3623..3643)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3808..3943
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3992..4786
                     /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             3992..4786
                     /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(4046..4065)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4656..4675
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4974..4993)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5018..5065
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5394..5982
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5883..5902
                     /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 ccggtcgcca ccatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg
      661 gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc
      721 gatgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg
      781 ccctggccca ccctcgtgac caccctgacc tacggcgtgc agtgcttcag ccgctacccc
      841 gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag
      901 cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag
      961 ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac
     1021 atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat catggccgac
     1081 aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga ggacggcagc
     1141 gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg
     1201 cccgacaacc actacctgag cacccagtcc gccctgagca aagaccccaa cgagaagcgc
     1261 gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag
     1321 ctgtacaagt ccggactcag atctcgagct caagctttga tagtgtttgt caggttcaac
     1381 tccagccatg gtttcccagt ggaggtcgat tctgacacca gcatcttcca gctcaaggag
     1441 gtggttgcta agcgacaggg ggttccggct gaccagttgc gtgtgatttt cgcagggaag
     1501 gagctgagga atgactggac tgtgcagaat tgtgacctgg atcagcagag cattgttcac
     1561 attgtgcaga gaccgtggag aaaaggtcaa gaaatgaatg caactggagg cgacgacccc
     1621 agaaacgcgg cgggaggctg tgagcgggag ccccagagct tgactcgggt ggacctcagc
     1681 agctcagtcc tcccaggaga ctctgtgggg ctggctgtca ttctgcacac tgacagcagg
     1741 aaggactcac caccagctgg aagtccagca ggtagatcaa tctacaacag cttttatgtg
     1801 tattgcaaag gcccctgtca aagagtgcag ccgggaaaac tcagggtaca gtgcagcacc
     1861 tgcaggcagg caacgctcac cttgacccag ggtccatctt gctgggatga tgttttaatt
     1921 ccaaaccgga tgagtggtga atgccaatcc ccacactgcc ctgggactag tgcagaattt
     1981 ttctttaaat gtggagcaca ccccacctct gacaaggaaa catcagtagc tttgcacctg
     2041 atcgcaacaa atagtcggaa catcacttgc attacgtgca cagacgtcag gagccccgtc
     2101 ctggttttcc agtgcaactc ccgccacgtg atttgcttag actgtttcca cttatactgt
     2161 gtgacaagac tcaatgatcg gcagtttgtt cacgaccctc aacttggcta ctccctgcct
     2221 tgtgtggctg gctgtcccaa ctccttgatt aaagagctcc atcacttcag gattctggga
     2281 gaagagcagt acaaccggta ccagcagtat ggtgcagagg agtgtgtcct gcagatgggg
     2341 ggcgtgttat gcccccgccc tggctgtgga gcggggctgc tgccggagcc tgaccagagg
     2401 aaagtcacct gcgaaggggg caatggcctg ggctgtgggt ttgccttctg ccgggaatgt
     2461 aaagaagcgt accatgaagg ggagtgcagt gccgtatttg aagcctcagg aacaactact
     2521 caggcctaca gagtcgatga aagagccgcc gagcaggctc gttgggaagc agcctccaaa
     2581 gaaaccatca agaaaaccac caagccctgt ccccgctgcc atgtaccagt ggaaaaaaat
     2641 ggaggcaaca tgcacatgaa gtgtccgcag ccccagtgca ggctcgagtg gtgctggaac
     2701 tgtggctgcg agtggaaccg cgtctgcatg ggggaccact ggttcgacgt gtagggatcc
     2761 accggatcta gataactgat cataatcagc cataccacat ttgtagaggt tttacttgct
     2821 ttaaaaaacc tcccacacct ccccctgaac ctgaaacata aaatgaatgc aattgttgtt
     2881 gttaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc
     2941 acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta
     3001 tcttaacgcg taaattgtaa gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta
     3061 aatcagctca ttttttaacc aataggccga aatcggcaaa atcccttata aatcaaaaga
     3121 atagaccgag atagggttga gtgttgttcc agtttggaac aagagtccac tattaaagaa
     3181 cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga
     3241 accatcaccc taatcaagtt ttttggggtc gaggtgccgt aaagcactaa atcggaaccc
     3301 taaagggagc ccccgattta gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga
     3361 agggaagaaa gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg
     3421 cgtaaccacc acacccgccg cgcttaatgc gccgctacag ggcgcgtcag gtggcacttt
     3481 tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta
     3541 tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtcc
     3601 tgaggcggaa agaaccagct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc
     3661 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga
     3721 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca
     3781 accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat
     3841 tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc cgcctcggcc
     3901 tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt ttgcaaagat
     3961 cgatcaagag acaggatgag gatcgtttcg catgattgaa caagatggat tgcacgcagg
     4021 ttctccggcc gcttgggtgg agaggctatt cggctatgac tgggcacaac agacaatcgg
     4081 ctgctctgat gccgccgtgt tccggctgtc agcgcagggg cgcccggttc tttttgtcaa
     4141 gaccgacctg tccggtgccc tgaatgaact gcaagacgag gcagcgcggc tatcgtggct
     4201 ggccacgacg ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga
     4261 ctggctgcta ttgggcgaag tgccggggca ggatctcctg tcatctcacc ttgctcctgc
     4321 cgagaaagta tccatcatgg ctgatgcaat gcggcggctg catacgcttg atccggctac
     4381 ctgcccattc gaccaccaag cgaaacatcg catcgagcga gcacgtactc ggatggaagc
     4441 cggtcttgtc gatcaggatg atctggacga agagcatcag gggctcgcgc cagccgaact
     4501 gttcgccagg ctcaaggcga gcatgcccga cggcgaggat ctcgtcgtga cccatggcga
     4561 tgcctgcttg ccgaatatca tggtggaaaa tggccgcttt tctggattca tcgactgtgg
     4621 ccggctgggt gtggcggacc gctatcagga catagcgttg gctacccgtg atattgctga
     4681 agagcttggc ggcgaatggg ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga
     4741 ttcgcagcgc atcgccttct atcgccttct tgacgagttc ttctgagcgg gactctgggg
     4801 ttcgaaatga ccgaccaagc gacgcccaac ctgccatcac gagatttcga ttccaccgcc
     4861 gccttctatg aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc
     4921 cagcgcgggg atctcatgct ggagttcttc gcccacccta gggggaggct aactgaaaca
     4981 cggaaggaga caataccgga aggaacccgc gctatgacgg caataaaaag acagaataaa
     5041 acgcacggtg ttgggtcgtt tgttcataaa cgcggggttc ggtcccaggg ctggcactct
     5101 gtcgataccc caccgagacc ccattggggc caatacgccc gcgtttcttc cttttcccca
     5161 ccccaccccc caagttcggg tgaaggccca gggctcgcag ccaacgtcgg ggcggcaggc
     5221 cctgccatag cctcaggtta ctcatatata ctttagattg atttaaaact tcatttttaa
     5281 tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt
     5341 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat
     5401 cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg
     5461 gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga
     5521 gcgcagatac caaatactgt ccttctagtg tagccgtagt taggccacca cttcaagaac
     5581 tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt
     5641 ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag
     5701 cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc
     5761 gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag
     5821 gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca
     5881 gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt
     5941 cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc
     6001 tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc
     6061 cctgattctg tggataaccg tattaccgcc atgcat
//
