LOCUS       sequence-261375-        6118 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6118)
  AUTHORS   Beilina A, Van Der Brug M, Ahmad R, Kesavapany S, Miller DW, Petsko 
            GA, Cookson MR
  TITLE     Mutations in PTEN-induced putative kinase 1 associated with 
            recessive parkinsonism have differential effects on protein 
            stability.
  JOURNAL   Proc Natl Acad Sci U S A. 2005 Apr 19. 102(16):5703-8.
  PUBMED    15824318
REFERENCE   2  (bases 1 to 6118)
  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: "Proc Natl 
            Acad Sci U S A. 2005 Apr 19. 102(16):5703-8."
COMMENT     Sequence Label: pGEX5X.1 PINK1 WT
FEATURES             Location/Qualifiers
     source          1..6118
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(183..188,189..204,205..211)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    219..235
                     /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)."
     CDS             258..911
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     869..891
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             921..932
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             948..2102
                     /codon_start=1
                     /product="kinase domain from human PTEN-induced putative 
                     kinase 1, a mitochondria serine/threonine protein kinase"
                     /label=PINK1 kinase domain
                     /translation="EEKQAESRRAVSACQEIQAIFTQKSKPGPDPLDTRRLQGFRLEEY
                     LIGQSIGKGCSAAVYEATMPTLPQNLEVTKSTGLLPGRGPGTSAPGEGQERAPGAPAFP
                     LAIKMMWNISAGSSSEAILNTMSQELVPASRVALAGEYGAVTYRKSKRGPKQLAPHPNI
                     IRVLRAFTSSVPLLPGALVDYPDVLPSRLHPEGLGHGRTLFLVMKNYPCTLRQYLCVNT
                     PSPRLAAMMLLQLLEGVDHLVQQGIAHRDLKSDNILVELDPDGCPWLVIADFGCCLADE
                     SIGLQLPFSSWYVDRGGNGCLMAPEVSTARPGPRAVIDYSKADAWAVGAIAYEIFGLVN
                     PFYGQGKAHLESRSYQEAQLPALPESVPPDVRQLVRALLQREASK"
     primer_bind     complement(2168..2190)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2335..2353)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2421..2525
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2526..2594,2595..3386)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(2744..2763)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3557..4145
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4046..4065
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4299..4316
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        4389..4466
                     /gene="lacI"
                     /label=lacI promoter
     CDS             4467..5549
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(4486..4505)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(5598..5603,5604..5621,5622..5628)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5636..5652
                     /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)."
     primer_bind     5660..5676
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5660..5676
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(5672..6118,1..81)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(5692..5709)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5692..5708)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5701..5723)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 agcttatcga ctgcacggtg caccaatgct tctggcgtca ggcagccatc ggaagctgtg
       61 gtatggctgt gcaggtcgta aatcactgca taattcgtgt cgctcaaggc gcactcccgt
      121 tctggataat gttttttgcg ccgacatcat aacggttctg gcaaatattc tgaaatgagc
      181 tgttgacaat taatcatcgg ctcgtataat gtgtggaatt gtgagcggat aacaatttca
      241 cacaggaaac agtattcatg tcccctatac taggttattg gaaaattaag ggccttgtgc
      301 aacccactcg acttcttttg gaatatcttg aagaaaaata tgaagagcat ttgtatgagc
      361 gcgatgaagg tgataaatgg cgaaacaaaa agtttgaatt gggtttggag tttcccaatc
      421 ttccttatta tattgatggt gatgttaaat taacacagtc tatggccatc atacgttata
      481 tagctgacaa gcacaacatg ttgggtggtt gtccaaaaga gcgtgcagag atttcaatgc
      541 ttgaaggagc ggttttggat attagatacg gtgtttcgag aattgcatat agtaaagact
      601 ttgaaactct caaagttgat tttcttagca agctacctga aatgctgaaa atgttcgaag
      661 atcgtttatg tcataaaaca tatttaaatg gtgatcatgt aacccatcct gacttcatgt
      721 tgtatgacgc tcttgatgtt gttttataca tggacccaat gtgcctggat gcgttcccaa
      781 aattagtttg ttttaaaaaa cgtattgaag ctatcccaca aattgataag tacttgaaat
      841 ccagcaagta tatagcatgg cctttgcagg gctggcaagc cacgtttggt ggtggcgacc
      901 atcctccaaa atcggatctg atcgaaggtc gtgggatccc cgaattcgag gaaaaacagg
      961 cggagagccg gcgggcggtc tcggcctgtc aggagatcca ggcaattttt acccagaaaa
     1021 gcaagccggg gcctgacccg ttggacacga gacgcttgca gggctttcgg ctggaggagt
     1081 atctgatagg gcagtccatt ggtaagggct gcagtgctgc tgtgtatgaa gccaccatgc
     1141 ctacattgcc ccagaacctg gaggtgacaa agagcaccgg gttgcttcca gggagaggcc
     1201 caggtaccag tgcaccagga gaagggcagg agcgagctcc gggggcccct gccttcccct
     1261 tggccatcaa gatgatgtgg aacatctcgg caggttcctc cagcgaagcc atcttgaaca
     1321 caatgagcca ggagctggtc ccagcgagcc gagtggcctt ggctggggag tatggagcag
     1381 tcacttacag aaaatccaag agaggtccca agcaactagc ccctcacccc aacatcatcc
     1441 gggttctccg cgccttcacc tcttccgtgc cgctgctgcc aggggccctg gtcgactacc
     1501 ctgatgtgct gccctcacgc ctccaccctg aaggcctggg ccatggccgg acgctgttcc
     1561 tcgttatgaa gaactatccc tgtaccctgc gccagtacct ttgtgtgaac acacccagcc
     1621 cccgcctcgc cgccatgatg ctgctgcagc tgctggaagg cgtggaccat ctggttcaac
     1681 agggcatcgc gcacagagac ctgaaatccg acaacatcct tgtggagctg gacccagacg
     1741 gctgcccctg gctggtgatc gcagattttg gctgctgcct ggctgatgag agcatcggcc
     1801 tgcagttgcc cttcagcagc tggtacgtgg atcggggcgg aaacggctgt ctgatggccc
     1861 cagaggtgtc cacggcccgt cctggcccca gggcagtgat tgactacagc aaggctgatg
     1921 cctgggcagt gggagccatc gcctatgaaa tcttcgggct tgtcaatccc ttctacggcc
     1981 agggcaaggc ccaccttgaa agccgcagct accaagaggc tcagctacct gcactgcccg
     2041 agtcagtgcc tccagacgtg agacagttgg tgagggcact gctccagcga gaggccagca
     2101 agctcgagcg gccgcatcgt gactgactga cgatctgcct cgcgcgtttc ggtgatgacg
     2161 gtgaaaacct ctgacacatg cagctcccgg agacggtcac agcttgtctg taagcggatg
     2221 ccgggagcag acaagcccgt cagggcgcgt cagcgggtgt tggcgggtgt cggggcgcag
     2281 ccatgaccca gtcacgtagc gatagcggag tgtataattc ttgaagacga aagggcctcg
     2341 tgatacgcct atttttatag gttaatgtca tgataataat ggtttcttag acgtcaggtg
     2401 gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa
     2461 atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga
     2521 agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc
     2581 ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg
     2641 gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc
     2701 gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat
     2761 tatcccgtgt tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg
     2821 acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag
     2881 aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa
     2941 cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc
     3001 gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca
     3061 cgatgcctgc agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc
     3121 tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc
     3181 tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg
     3241 ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta
     3301 tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag
     3361 gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga
     3421 ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc
     3481 tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa
     3541 agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa
     3601 aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc
     3661 cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgtccttcta gtgtagccgt
     3721 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc
     3781 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac
     3841 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca
     3901 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg
     3961 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag
     4021 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt
     4081 ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat
     4141 ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc
     4201 acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt
     4261 gagctgatac cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag
     4321 cggaagagcg cctgatgcgg tattttctcc ttacgcatct gtgcggtatt tcacaccgca
     4381 taaattccga caccatcgaa tggcgcaaaa cctttcgcgg tatggcatga tagcgcccgg
     4441 aagagagtca attcagggtg gtgaatgtga aaccagtaac gttatacgat gtcgcagagt
     4501 atgccggtgt ctcttatcag accgtttccc gcgtggtgaa ccaggccagc cacgtttctg
     4561 cgaaaacgcg ggaaaaagtg gaagcggcga tggcggagct gaattacatt cccaaccgcg
     4621 tggcacaaca actggcgggc aaacagtcgt tgctgattgg cgttgccacc tccagtctgg
     4681 ccctgcacgc gccgtcgcaa attgtcgcgg cgattaaatc tcgcgccgat caactgggtg
     4741 ccagcgtggt ggtgtcgatg gtagaacgaa gcggcgtcga agcctgtaaa gcggcggtgc
     4801 acaatcttct cgcgcaacgc gtcagtgggc tgatcattaa ctatccgctg gatgaccagg
     4861 atgccattgc tgtggaagct gcctgcacta atgttccggc gttatttctt gatgtctctg
     4921 accagacacc catcaacagt attattttct cccatgaaga cggtacgcga ctgggcgtgg
     4981 agcatctggt cgcattgggt caccagcaaa tcgcgctgtt agcgggccca ttaagttctg
     5041 tctcggcgcg tctgcgtctg gctggctggc ataaatatct cactcgcaat caaattcagc
     5101 cgatagcgga acgggaaggc gactggagtg ccatgtccgg ttttcaacaa accatgcaaa
     5161 tgctgaatga gggcatcgtt cccactgcga tgctggttgc caacgatcag atggcgctgg
     5221 gcgcaatgcg cgccattacc gagtccgggc tgcgcgttgg tgcggatatc tcggtagtgg
     5281 gatacgacga taccgaagac agctcatgtt atatcccgcc gttaaccacc atcaaacagg
     5341 attttcgcct gctggggcaa accagcgtgg accgcttgct gcaactctct cagggccagg
     5401 cggtgaaggg caatcagctg ttgcccgtct cactggtgaa aagaaaaacc accctggcgc
     5461 ccaatacgca aaccgcctct ccccgcgcgt tggccgattc attaatgcag ctggcacgac
     5521 aggtttcccg actggaaagc gggcagtgag cgcaacgcaa ttaatgtaag ttagctcact
     5581 cattaggcac cccaggcttt acactttatg cttccggctc gtatgttgtg tggaattgtg
     5641 agcggataac aatttcacac aggaaacagc tatgaccatg attacggatt cactggccgt
     5701 cgttttacaa cgtcgtgact gggaaaaccc tggcgttacc caacttaatc gccttgcagc
     5761 acatccccct ttcgccagct ggcgtaatag cgaagaggcc cgcaccgatc gcccttccca
     5821 acagttgcgc agcctgaatg gcgaatggcg ctttgcctgg tttccggcac cagaagcggt
     5881 gccggaaagc tggctggagt gcgatcttcc tgaggccgat actgtcgtcg tcccctcaaa
     5941 ctggcagatg cacggttacg atgcgcccat ctacaccaac gtgacctatc ccattacggt
     6001 caatccgccg tttgttccca cggagaatcc gacgggttgt tactcgctca catttaatgt
     6061 tgatgaaagc tggctacagg aaggccagac gcgaattatt tttgatggcg ttggaatt
//
