LOCUS       sequence-477203-        9299 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Overexpression of human PARK2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9299)
  AUTHORS   Goldman, C., Kareva, T., Sarrafha, L., Schuldt, B.R., Sahasrabudhe, 
            A., Ahfeldt, T.,, Blanchard, J.W
  TITLE     Genetically encoded and modular subcellular organelle probes reveal 
            dysfunction in lysosomes and mitochondria driven by PRKN knockout
  JOURNAL   iScience 28 (2025)
REFERENCE   2  (bases 1 to 9299)
  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: "iScience 28
            (2025)"
COMMENT     Sequence Label: pFUW PRKN-P2A-PuroR
FEATURES             Location/Qualifiers
     source          1..9299
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(47..66)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        238..617
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        619..817
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     772..792
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             835..1015
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1062..1187
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1680..1913
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2098..2142
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    2440..2557
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2622..3833
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     3670..3689
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             5283..5339
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             5340..5939
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(5340..5359)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5836..5856
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    5965..6553
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(6018..6038)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(6436..6447)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(6556..6572)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6557..6573)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             7078..7258
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(7284..7301)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    7290..7401
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      complement(7547..8135)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(7627..7646)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(8306..9097,9098..9166))
                     /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     8929..8948
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(9167..9271)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgacggat cgggagatct cccgatcccc tatggtgcac tctcagtaca atctgctctg
       61 atgccgcata gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt
      121 gcgcgagcaa aatttaagct acaacaaggc aaggcttgac cgacaattgc atgaagaatc
      181 tgcttagggt taggcgtttt gcgctgcttc gcgatgtacg ggccagatat acgcgttgac
      241 attgattatt gactagttat taatagtaat caattacggg gtcattagtt catagcccat
      301 atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg
      361 acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt
      421 tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag
      481 tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc
      541 attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag
      601 tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt ggatagcggt
      661 ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc
      721 accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg
      781 gcggtaggcg tgtacggtgg gaggtctata taagcagcgc gttttgcctg tactgggtct
      841 ctctggttag accagatctg agcctgggag ctctctggct aactagggaa cccactgctt
      901 aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac
      961 tctggtaact agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtggc
     1021 gcccgaacag ggacttgaaa gcgaaaggga aaccagagga gctctctcga cgcaggactc
     1081 ggcttgctga agcgcgcacg gcaagaggcg aggggcggcg actggtgagt acgccaaaaa
     1141 ttttgactag cggaggctag aaggagagag atgggtgcga gagcgtcagt attaagcggg
     1201 ggagaattag atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag aaaaaatata
     1261 aattaaaaca tatagtatgg gcaagcaggg agctagaacg attcgcagtt aatcctggcc
     1321 tgttagaaac atcagaaggc tgtagacaaa tactgggaca gctacaacca tcccttcaga
     1381 caggatcaga agaacttaga tcattatata atacagtagc aaccctctat tgtgtgcatc
     1441 aaaggataga gataaaagac accaaggaag ctttagacaa gatagaggaa gagcaaaaca
     1501 aaagtaagac caccgcacag caagcggccg ctgatcttca gacctggagg aggagatatg
     1561 agggacaatt ggagaagtga attatataaa tataaagtag taaaaattga accattagga
     1621 gtagcaccca ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc agtgggaata
     1681 ggagctttgt tccttgggtt cttgggagca gcaggaagca ctatgggcgc agcgtcaatg
     1741 acgctgacgg tacaggccag acaattattg tctggtatag tgcagcagca gaacaatttg
     1801 ctgagggcta ttgaggcgca acagcatctg ttgcaactca cagtctgggg catcaagcag
     1861 ctccaggcaa gaatcctggc tgtggaaaga tacctaaagg atcaacagct cctggggatt
     1921 tggggttgct ctggaaaact catttgcacc actgctgtgc cttggaatgc tagttggagt
     1981 aataaatctc tggaacagat ttggaatcac acgacctgga tggagtggga cagagaaatt
     2041 aacaattaca caagcttaat acactcctta attgaagaat cgcaaaacca gcaagaaaag
     2101 aatgaacaag aattattgga attagataaa tgggcaagtt tgtggaattg gtttaacata
     2161 acaaattggc tgtggtatat aaaattattc ataatgatag taggaggctt ggtaggttta
     2221 agaatagttt ttgctgtact ttctatagtg aatagagtta ggcagggata ttcaccatta
     2281 tcgtttcaga cccacctccc aaccccgagg ggacccgaca ggcccgaagg aatagaagaa
     2341 gaaggtggag agagagacag agacagatcc attcgattag tgaacggatc ggcactgcgt
     2401 gcgccaattc tgcagacaaa tggcagtatt catccacaat tttaaaagaa aaggggggat
     2461 tggggggtac agtgcagggg aaagaatagt agacataata gcaacagaca tacaaactaa
     2521 agaattacaa aaacaaatta caaaaattca aaattttcgg gtttattaca gggacagcag
     2581 agatccagtt tggttaatta acccgtgtcg gctccagatc tggcctccgc gccgggtttt
     2641 ggcgcctccc gcgggcgccc ccctcctcac ggcgagcgct gccacgtcag acgaagggcg
     2701 cagcgagcgt cctgatcctt ccgcccggac gctcaggaca gcggcccgct gctcataaga
     2761 ctcggcctta gaaccccagt atcagcagaa ggacatttta ggacgggact tgggtgactc
     2821 tagggcactg gttttctttc cagagagcgg aacaggcgag gaaaagtagt cccttctcgg
     2881 cgattctgcg gagggatctc cgtggggcgg tgaacgccga tgattatata aggacgcgcc
     2941 gggtgtggca cagctagttc cgtcgcagcc gggatttggg tcgcggttct tgtttgtgga
     3001 tcgctgtgat cgtcacttgg tgagtagcgg gctgctgggc tggccggggc tttcgtggcc
     3061 gccgggccgc tcggtgggac ggaagcgtgt ggagagaccg ccaagggctg tagtctgggt
     3121 ccgcgagcaa ggttgccctg aactgggggt tggggggagc gcagcaaaat ggcggctgtt
     3181 cccgagtctt gaatggaaga cgcttgtgag gcgggctgtg aggtcgttga aacaaggtgg
     3241 ggggcatggt gggcggcaag aacccaaggt cttgaggcct tcgctaatgc gggaaagctc
     3301 ttattcgggt gagatgggct ggggcaccat ctggggaccc tgacgtgaag tttgtcactg
     3361 actggagaac tcggtttgtc gtctgttgcg ggggcggcag ttatggcggt gccgttgggc
     3421 agtgcacccg tacctttggg agcgcgcgcc ctcgtcgtgt cgtgacgtca cccgttctgt
     3481 tggcttataa tgcagggtgg ggccacctgc cggtaggtgt gcggtaggct tttctccgtc
     3541 gcaggacgca gggttcgggc ctagggtagg ctctcctgaa tcgacaggcg ccggacctct
     3601 ggtgagggga gggataagtg aggcgtcagt ttctttggtc ggttttatgt acctatcttc
     3661 ttaagtagct gaagctccgg ttttgaacta tgcgctcggg gttggcgagt gtgttttgtg
     3721 aagtttttta ggcacctttt gaaatgtaat catttgggtc aatatgtaat tttcagtgtt
     3781 agactagtaa attgtccgct aaattctggc cgtttttggc ttttttgtta gacgaagctt
     3841 gggcccggga attaattatt aagatgatag tgtttgtcag gttcaactcc agccatggtt
     3901 tcccagtgga ggtcgattct gacaccagca tcttccagct caaggaggtg gttgctaagc
     3961 gacagggggt tccggctgac cagttgcgtg tgattttcgc agggaaggag ctgaggaatg
     4021 actggactgt gcagaattgt gacctggatc agcagagcat tgttcacatt gtgcagagac
     4081 cgtggagaaa aggtcaagaa atgaatgcaa ctggaggcga cgaccccaga aacgcggcgg
     4141 gaggctgtga gcgggagccc cagagcttga ctcgggtgga cctcagcagc tcagtcctcc
     4201 caggagactc tgtggggctg gctgtcattc tgcacactga cagcaggaag gactcaccac
     4261 cagctggaag tccagcaggt agatcaatct acaacagctt ttatgtgtat tgcaaaggcc
     4321 cctgtcaaag agtgcagccg ggaaaactca gggtacagtg cagcacctgc aggcaggcaa
     4381 cgctcacctt gacccagggt ccatcttgct gggatgatgt tttaattcca aaccggatga
     4441 gtggtgaatg ccaatcccca cactgccctg ggactagtgc agaatttttc tttaaatgtg
     4501 gagcacaccc cacctctgac aaggaaacat cagtagcttt gcacctgatc gcaacaaata
     4561 gtcggaacat cacttgcatt acgtgcacag acgtcaggag ccccgtcctg gttttccagt
     4621 gcaactcccg ccacgtgatt tgcttagact gtttccactt atactgtgtg acaagactca
     4681 atgatcggca gtttgttcac gaccctcaac ttggctactc cctgccttgt gtggctggct
     4741 gtcccaactc cttgattaaa gagctccatc acttcaggat tctgggagaa gagcagtaca
     4801 accggtacca gcagtatggt gcagaggagt gtgtcctgca gatggggggc gtgttatgcc
     4861 cccgccctgg ctgtggagcg gggctgctgc cggagcctga ccagaggaaa gtcacctgcg
     4921 aagggggcaa tggcctgggc tgtgggtttg ccttctgccg ggaatgtaaa gaagcgtacc
     4981 atgaagggga gtgcagtgcc gtatttgaag cctcaggaac aactactcag gcctacagag
     5041 tcgatgaaag agccgccgag caggctcgtt gggaagcagc ctccaaagaa accatcaaga
     5101 aaaccaccaa gccctgtccc cgctgccatg taccagtgga aaaaaatgga ggctgcatgc
     5161 acatgaagtg tccgcagccc cagtgcaggc tcgagtggtg ctggaactgt ggctgcgagt
     5221 ggaaccgcgt ctgcatgggg gaccactggt tcgacgtgga tctgcagtct agaggatctg
     5281 gtgccacgaa tttctcacta ctcaagcagg ccggtgatgt cgaggaaaac cctggtccta
     5341 tgaccgagta caagcccacg gtgcgcctcg ccacccgcga cgacgtcccc agggccgtac
     5401 gcaccctcgc cgccgcgttc gccgactacc ccgccacgcg ccacaccgtc gatccggacc
     5461 gccacatcga gcgggtcacc gagctgcaag aactcttcct cacgcgcgtc gggctcgaca
     5521 tcggcaaggt gtgggtcgcg gacgacggcg ccgcggtggc ggtctggacc acgccggaga
     5581 gcgtcgaagc gggggcggtg ttcgccgaga tcggcccgcg catggccgag ttgagcggtt
     5641 cccggctggc cgcgcagcaa cagatggaag gcctcctggc gccgcaccgg cccaaggagc
     5701 ccgcgtggtt cctggccacc gtcggcgtct cgcccgacca ccagggcaag ggtctgggca
     5761 gcgccgtcgt gctccccgga gtggaggcgg ccgagcgcgc cggggtgccc gccttcctgg
     5821 agacctccgc gccccgcaac ctccccttct acgagcggct cggcttcacc gtcaccgccg
     5881 acgtcgaggt gcccgaagga ccgcgcacct ggtgcatgac ccgcaagccc ggtgcctgac
     5941 gaattcgata tcaagcttat cgataatcaa cctctggatt acaaaatttg tgaaagattg
     6001 actggtattc ttaactatgt tgctcctttt acgctatgtg gatacgctgc tttaatgcct
     6061 ttgtatcatg ctattgcttc ccgtatggct ttcattttct cctccttgta taaatcctgg
     6121 ttgctgtctc tttatgagga gttgtggccc gttgtcaggc aacgtggcgt ggtgtgcact
     6181 gtgtttgctg acgcaacccc cactggttgg ggcattgcca ccacctgtca gctcctttcc
     6241 gggactttcg ctttccccct ccctattgcc acggcggaac tcatcgccgc ctgccttgcc
     6301 cgctgctgga caggggctcg gctgttgggc actgacaatt ccgtggtgtt gtcggggaaa
     6361 tcatcgtcct ttccttggct gctcgcctgt gttgccacct ggattctgcg cgggacgtcc
     6421 ttctgctacg tcccttcggc cctcaatcca gcggaccttc cttcccgcgg cctgctgccg
     6481 gctctgcggc ctcttccgcg tcttcgcctt cgccctcaga cgagtcggat ctccctttgg
     6541 gccgcctccc cgcatcgata ccgtcgacct cgagacctag aaaaacatgg agcaatcaca
     6601 agtagcaata cagcagctac caatgctgat tgtgcctggc tagaagcaca agaggaggag
     6661 gaggtgggtt ttccagtcac acctcaggta cctttaagac caatgactta caaggcagct
     6721 gtagatctta gccacttttt aaaagaaaag gggggactgg aagggctaat tcactcccaa
     6781 cgaagacaag atatccttga tctgtggatc taccacacac aaggctactt ccctgattgg
     6841 cagaactaca caccagggcc agggatcaga tatccactga cctttggatg gtgctacaag
     6901 ctagtaccag ttgagcaaga gaaggtagaa gaagccaatg aaggagagaa cacccgcttg
     6961 ttacaccctg tgagcctgca tgggatggat gacccggaga gagaagtatt agagtggagg
     7021 tttgacagcc gcctagcatt tcatcacatg gcccgagagc tgcatccgga ctgtactggg
     7081 tctctctggt tagaccagat ctgagcctgg gagctctctg gctaactagg gaacccactg
     7141 cttaagcctc aataaagctt gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt
     7201 gactctggta actagagatc cctcagaccc ttttagtcag tgtggaaaat ctctagcagg
     7261 gcccgtttaa acccgctgat cagcctcgac tgtgccttct agttgccagc catctgttgt
     7321 ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc actcccactg tcctttccta
     7381 ataaaatgag gaaattgcat cgcattgtct gagtaggtgt cattctattc tggggggtgg
     7441 ggtggggcag gacagcaagg gggaggattg ggaagacaat agcaggcatg tgagcaaaag
     7501 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc
     7561 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
     7621 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
     7681 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
     7741 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
     7801 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
     7861 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
     7921 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
     7981 ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
     8041 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
     8101 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg
     8161 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa
     8221 aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta
     8281 tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag
     8341 cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga
     8401 tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac
     8461 cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc
     8521 ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta
     8581 gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac
     8641 gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat
     8701 gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa
     8761 gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg
     8821 tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag
     8881 aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc
     8941 cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct
     9001 caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat
     9061 cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg
     9121 ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc
     9181 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
     9241 tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgac
//
