LOCUS       sequence-390802-        6694 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Used for the expression and purification of OPTN (WT) (SMC Internal 
            No. 1587).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6694)
  AUTHORS   Nguyen TN, Sawa-Makarska J, Khuu G, Lam WK, Adriaenssens E, 
            Fracchiolla D, Shoebridge S, Bernklau D, Padman BS, Skulsuppaisarn 
            M, Lindblom RSJ, Martens S, Lazarou M
  TITLE     Unconventional initiation of PINK1/Parkin mitophagy by Optineurin.
  JOURNAL   Mol Cell. 2023 May 18;83(10):1693-1709.e9. doi: 
            10.1016/j.molcel.2023.04.021.
  PUBMED    37207627
REFERENCE   2  (bases 1 to 6694)
  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: 
            "10.1016/j.molcel.2023.04"; journalName: "Mol Cell"; date: 
            "2023-05-18- 18"; volume: "83"; issue: "10"; pages: "1693-1709.e9"
COMMENT     Sequence Label: pGST2-GST-TEV-OPTN
FEATURES             Location/Qualifiers
     source          1..6694
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(196..201,202..217,218..224)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    232..248
                     /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             271..924
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     882..904
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             931..951
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             1275..1286
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(2757..2779)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2924..2942)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3010..3114
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3115..3183,3184..3975)
                     /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(3333..3352)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4146..4734
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4635..4654
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4888..4905
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        4978..5055
                     /gene="lacI"
                     /label=lacI promoter
     CDS             5056..6138
                     /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(5075..5094)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(6187..6192,6193..6210,6211..6217)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6225..6241
                     /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     6249..6265
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6249..6265
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(6261..6694,1..94)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(6281..6298)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6281..6297)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6290..6312)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 tggcgttgga attagcttat cgactgcacg gtgcaccaat gcttctggcg tcaggcagcc
       61 atcggaagct gtggtatggc tgtgcaggtc gtaaatcact gcataattcg tgtcgctcaa
      121 ggcgcactcc cgttctggat aatgtttttt gcgccgacat cataacggtt ctggcaaata
      181 ttctgaaatg agctgttgac aattaatcat cggctcgtat aatgtgtgga attgtgagcg
      241 gataacaatt tcacacagga aacagtattc atgtccccta tactaggtta ttggaaaatt
      301 aagggccttg tgcaacccac tcgacttctt ttggaatatc ttgaagaaaa atatgaagag
      361 catttgtatg agcgcgatga aggtgataaa tggcgaaaca aaaagtttga attgggtttg
      421 gagtttccca atcttcctta ttatattgat ggtgatgtta aattaacaca gtctatggcc
      481 atcatacgtt atatagctga caagcacaac atgttgggtg gttgtccaaa agagcgtgca
      541 gagatttcaa tgcttgaagg agcggttttg gatattagat acggtgtttc gagaattgca
      601 tatagtaaag actttgaaac tctcaaagtt gattttctta gcaagctacc tgaaatgctg
      661 aaaatgttcg aagatcgttt atgtcataaa acatatttaa atggtgatca tgtaacccat
      721 cctgacttca tgttgtatga cgctcttgat gttgttttat acatggaccc aatgtgcctg
      781 gatgcgttcc caaaattagt ttgttttaaa aaacgtattg aagctatccc acaaattgat
      841 aagtacttga aatccagcaa gtatatagca tggcctttgc agggctggca agccacgttt
      901 ggtggtggcg accatcctcc aaaaatcgat gaaaacctgt attttcaggg atcaggcatg
      961 tcccatcaac ctctcagctg cctcactgaa aaggaggaca gccccagtga aagcacagga
     1021 aatggacccc cccacctggc ccacccaaac ctggacacgt ttaccccgga ggagctgctg
     1081 cagcagatga aagagctcct gaccgagaac caccagctga aagaagccat gaagctaaat
     1141 aatcaagcca tgaaagggag atttgaggag ctttcggcct ggacagagaa acagaaggaa
     1201 gaacgccagt tttttgagat acagagcaaa gaagcaaaag agcgtctaat ggccttgagt
     1261 catgagaatg agaaattgaa ggaagagctt ggaaaactaa aagggaaatc agaaaggtca
     1321 tctgaggacc ccactgatga ctccaggctt cccagggccg aagcggagca ggaaaaggac
     1381 cagctcagga cccaggtggt gaggctacaa gcagagaagg cagacctgtt gggcatcgtg
     1441 tctgaactgc agctcaagct gaactccagc ggctcctcag aagattcctt tgttgaaatt
     1501 aggatggctg aaggagaagc agaagggtca gtaaaagaaa tcaagcatag tcctgggccc
     1561 acgagaacag tctccactgg cacggcattg tctaaatata ggagcagatc tgcagatggg
     1621 gccaagaatt acttcgaaca tgaggagtta actgtgagcc agctcctgct gtgcctaagg
     1681 gaagggaatc agaaggtgga gagacttgaa gttgcactca aggaggccaa agaaagagtt
     1741 tcagattttg aaaagaaaac aagtaatcgt tctgagattg aaacccagac agaggggagc
     1801 acagagaaag agaatgatga agagaaaggc ccggagactg ttggaagcga agtggaagca
     1861 ctgaacctcc aggtgacatc tctgtttaag gagcttcaag aggctcatac aaaactcagc
     1921 gaagctgagc taatgaagaa gagacttcaa gaaaagtgtc aggcccttga aaggaaaaat
     1981 tctgcaattc catcagagtt gaatgaaaag caagagcttg tttatactaa caaaaagtta
     2041 gagctacaag tggaaagcat gctatcagaa atcaaaatgg aacaggctaa aacagaggat
     2101 gaaaagtcca aattaactgt gctacagatg acacacaaca agcttcttca agaacataat
     2161 aatgcattga aaacaattga ggaactaaca agaaaagagt cagaaaaagt ggacagggca
     2221 gtgctgaagg aactgagtga aaaactggaa ctggcagaga aggctctggc ttccaaacag
     2281 ctgcaaatgg atgaaatgaa gcaaaccatt gccaagcagg aagaggacct ggaaaccatg
     2341 accatcctca gggctcagat ggaagtttac tgttctgatt ttcatgctga aagagcagcg
     2401 agagagaaaa ttcatgagga aaaggagcaa ctggcattgc agctggcagt tctgctgaaa
     2461 gagaatgatg ctttcgaaga cggaggcagg cagtccttga tggagatgca gagtcgtcat
     2521 ggggcgagaa caagtgactc tgaccagcag gcttaccttg ttcaaagagg agctgaggac
     2581 agggactggc ggcaacagcg gaatattccg attcattcct gccccaagtg tggagaggtt
     2641 ctgcctgaca tagacacgtt acagattcac gtgatggatt gcatcattta actcgagcgg
     2701 ccgcatcgtg actgactgac gatctgcctc gcgcgtttcg gtgatgacgg tgaaaacctc
     2761 tgacacatgc agctcccgga gacggtcaca gcttgtctgt aagcggatgc cgggagcaga
     2821 caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc ggggcgcagc catgacccag
     2881 tcacgtagcg atagcggagt gtataattct tgaagacgaa agggcctcgt gatacgccta
     2941 tttttatagg ttaatgtcat gataataatg gtttcttaga cgtcaggtgg cacttttcgg
     3001 ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg
     3061 ctaatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt
     3121 attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt
     3181 gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg
     3241 ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa
     3301 cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtgtt
     3361 gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag
     3421 tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt
     3481 gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga
     3541 ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt
     3601 tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgca
     3661 gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg
     3721 caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc
     3781 cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt
     3841 atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg
     3901 gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg
     3961 attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa
     4021 cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa
     4081 atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga
     4141 tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg
     4201 ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact
     4261 ggcttcagca gagcgcagat accaaatact gtccttctag tgtagccgta gttaggccac
     4321 cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg
     4381 gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg
     4441 gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga
     4501 acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc
     4561 gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg
     4621 agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc
     4681 tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc
     4741 agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt
     4801 cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg agctgatacc
     4861 gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc
     4921 ctgatgcggt attttctcct tacgcatctg tgcggtattt cacaccgcat aaattccgac
     4981 accatcgaat ggcgcaaaac ctttcgcggt atggcatgat agcgcccgga agagagtcaa
     5041 ttcagggtgg tgaatgtgaa accagtaacg ttatacgatg tcgcagagta tgccggtgtc
     5101 tcttatcaga ccgtttcccg cgtggtgaac caggccagcc acgtttctgc gaaaacgcgg
     5161 gaaaaagtgg aagcggcgat ggcggagctg aattacattc ccaaccgcgt ggcacaacaa
     5221 ctggcgggca aacagtcgtt gctgattggc gttgccacct ccagtctggc cctgcacgcg
     5281 ccgtcgcaaa ttgtcgcggc gattaaatct cgcgccgatc aactgggtgc cagcgtggtg
     5341 gtgtcgatgg tagaacgaag cggcgtcgaa gcctgtaaag cggcggtgca caatcttctc
     5401 gcgcaacgcg tcagtgggct gatcattaac tatccgctgg atgaccagga tgccattgct
     5461 gtggaagctg cctgcactaa tgttccggcg ttatttcttg atgtctctga ccagacaccc
     5521 atcaacagta ttattttctc ccatgaagac ggtacgcgac tgggcgtgga gcatctggtc
     5581 gcattgggtc accagcaaat cgcgctgtta gcgggcccat taagttctgt ctcggcgcgt
     5641 ctgcgtctgg ctggctggca taaatatctc actcgcaatc aaattcagcc gatagcggaa
     5701 cgggaaggcg actggagtgc catgtccggt tttcaacaaa ccatgcaaat gctgaatgag
     5761 ggcatcgttc ccactgcgat gctggttgcc aacgatcaga tggcgctggg cgcaatgcgc
     5821 gccattaccg agtccgggct gcgcgttggt gcggatatct cggtagtggg atacgacgat
     5881 accgaagaca gctcatgtta tatcccgccg ttaaccacca tcaaacagga ttttcgcctg
     5941 ctggggcaaa ccagcgtgga ccgcttgctg caactctctc agggccaggc ggtgaagggc
     6001 aatcagctgt tgcccgtctc actggtgaaa agaaaaacca ccctggcgcc caatacgcaa
     6061 accgcctctc cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca ggtttcccga
     6121 ctggaaagcg ggcagtgagc gcaacgcaat taatgtaagt tagctcactc attaggcacc
     6181 ccaggcttta cactttatgc ttccggctcg tatgttgtgt ggaattgtga gcggataaca
     6241 atttcacaca ggaaacagct atgaccatga ttacggattc actggccgtc gttttacaac
     6301 gtcgtgactg ggaaaaccct ggcgttaccc aacttaatcg ccttgcagca catccccctt
     6361 tcgccagctg gcgtaatagc gaagaggccc gcaccgatcg cccttcccaa cagttgcgca
     6421 gcctgaatgg cgaatggcgc tttgcctggt ttccggcacc agaagcggtg ccggaaagct
     6481 ggctggagtg cgatcttcct gaggccgata ctgtcgtcgt cccctcaaac tggcagatgc
     6541 acggttacga tgcgcccatc tacaccaacg tgacctatcc cattacggtc aatccgccgt
     6601 ttgttcccac ggagaatccg acgggttgtt actcgctcac atttaatgtt gatgaaagct
     6661 ggctacagga aggccagacg cgaattattt ttga
//
