LOCUS       sequence-391275-        8771 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Used for the expression and purification of mCherry labelled TBK1 
            (SMC Internal No.: 1675).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8771)
  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 8771)
  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: pFastBac-GST-TEV-mCherry-TBK1
FEATURES             Location/Qualifiers
     source          1..8771
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(join(85..90,91..107,108..113))
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     polyA_signal    621..669
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     694..713
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     promoter        complement(805..914)
                     /label=p10 promoter
                     /note="baculovirus promoter for expression in insect cells"
     promoter        938..1029
                     /gene="polh from Autographa californica"
                     /label=polyhedrin promoter
                     /note="promoter for the baculovirus polyhedrin gene"
     primer_bind     954..971
                     /label=Polyhedrin-F
                     /note="Polyhedrin promoter, forward primer"
     CDS             1072..1728
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MASPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNK
                     KFELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIR
                     YGVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVV
                     LYMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     1686..1708
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             1735..1752
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             1762..1782
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             1789..2496
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(1934..1952)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2202..2221
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    4822..4956
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4859..4878)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4913..4932
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     mobile_element  4985..5150
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7L
                     /note="mini-Tn7 element (left end of the Tn7 transposon)"
     rep_origin      5334..5788
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5421..5440)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5630..5651
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5814..5918
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5919..5987,5988..6779)
                     /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     complement(6137..6156)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6950..7538
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7439..7458
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7692..7709
                     /label=L4440
                     /note="L4440 vector, forward primer"
     mobile_element  7843..8067
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7R
                     /note="mini-Tn7 element (right end of the Tn7 transposon)"
     CDS             complement(8134..8667)
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
ORIGIN
        1 aaaaccgcca ctgcgccgtt accaccgctg cgttcggtca aggttctgga ccagttgcgt
       61 gagcgcatac gctacttgca ttacagttta cgaaccgaac aggcttatgt caactgggtt
      121 cgtgccttca tccgtttcca cggtgtgcgt cacccggcaa ccttgggcag cagcgaagtc
      181 gaggcatttc tgtcctggct ggcgaacgag cgcaaggttt cggtctccac gcatcgtcag
      241 gcattggcgg ccttgctgtt cttctacggc aaggtgctgt gcacggatct gcccttgctt
      301 caggagatcg gtagacctcg gccgtcgcgg cgcttgccgg tggtgctgac cccggatgaa
      361 gtggttcgca tcctcggttt tctggaaggc gagcatcgtt tgttcgccca ggactctagc
      421 tatagttcta gtggttggct acgtacccgt agtggctatg gcagggcttg ccgccccgac
      481 gttggctgcg agccctgggc cttcacccga acttgggggt tggggtgggg aaaaggaaga
      541 aacgcgggcg tattggtccc aatggggtct cggtggggta tcgacagagt gccagccctg
      601 ggaccgaacc ccgcgtttat gaacaaacga cccaacaccc gtgcgtttta ttctgtcttt
      661 ttattgccgt catagcgcgg gttccttccg gtattgtctc cttccgtgtt tcagttagcc
      721 tcccccatct cccggtaccg catgctatgc atcagctgct agcaccatgg ctcgagatcc
      781 cgggtgatca agtcttcgtc gagtgattgt aaataaaatg taatttacag tatagtattt
      841 taattaatat acaaatgatt tgataataat tcttatttaa ctataatata ttgtgttggg
      901 ttgaattaaa ggtccgtata ctccggaata ttaatagatc atggagataa ttaaaatgat
      961 aaccatctcg caaataaata agtattttac tgttttcgta acagttttgt aataaaaaaa
     1021 cctataaata ttccggatta ttcataccgt cccaccatcg ggcgcggatc catggcatcc
     1081 cctatactag gttattggaa aattaagggc cttgtgcaac ccactcgact tcttttggaa
     1141 tatcttgaag aaaaatatga agagcatttg tatgagcgcg atgaaggtga taaatggcga
     1201 aacaaaaagt ttgaattggg tttggagttt cccaatcttc cttattatat tgatggtgat
     1261 gttaaattaa cacagtctat ggccatcata cgttatatag ctgacaagca caacatgttg
     1321 ggtggttgtc caaaagagcg tgcagagatt tcaatgcttg aaggagcggt tttggatatt
     1381 agatacggtg tttcgagaat tgcatatagt aaagactttg aaactctcaa agttgatttt
     1441 cttagcaagc tacctgaaat gctgaaaatg ttcgaagatc gtttatgtca taaaacatat
     1501 ttaaatggtg atcatgtaac ccatcctgac ttcatgttgt atgacgctct tgatgttgtt
     1561 ttatacatgg acccaatgtg cctggatgcg ttcccaaaat tagtttgttt taaaaaacgt
     1621 attgaagcta tcccacaaat tgataagtac ttgaaatcca gcaagtatat agcatggcct
     1681 ttgcagggct ggcaagccac gtttggtggt ggcgaccatc ctccaaaatc ggatctggtt
     1741 ccgcgtggat caggatcagg agagaattta tactttcagg gagcgcgcat ggtgagcaag
     1801 ggcgaggagg ataacatggc catcatcaag gagttcatgc gcttcaaggt gcacatggag
     1861 ggctccgtga acggccacga gttcgagatc gagggcgagg gcgagggccg cccctacgag
     1921 ggcacccaga ccgccaagct gaaggtgacc aagggtggcc ccctgccctt cgcctgggac
     1981 atcctgtccc ctcagttcat gtacggctcc aaggcctacg tgaagcaccc cgccgacatc
     2041 cccgactact tgaagctgtc cttccccgag ggcttcaagt gggagcgcgt gatgaacttc
     2101 gaggacggcg gcgtggtgac cgtgacccag gactcctccc tgcaggacgg cgagttcatc
     2161 tacaaggtga agctgcgcgg caccaacttc ccctccgacg gccccgtaat gcagaagaag
     2221 accatgggct gggaggcctc ctccgagcgg atgtaccccg aggacggcgc cctgaagggc
     2281 gagatcaagc agaggctgaa gctgaaggac ggcggccact acgacgctga ggtcaagacc
     2341 acctacaagg ccaagaagcc cgtgcagctg cccggcgcct acaacgtcaa catcaagttg
     2401 gacatcacct cccacaacga ggactacacc atcgtggaac agtacgaacg cgccgagggc
     2461 cgccactcca ccggcggcat ggacgagctg tacaaggcgc gcatgcaatc cacatccaac
     2521 cacctctggc tcctctcgga cattctcgga caaggcgcta ccgctaacgt gttcagggga
     2581 aggcacaaga agaccggtga cctgttcgcc atcaaggtgt tcaacaacat ctcattcctg
     2641 cgccctgtgg acgtccagat gcgtgagttc gaagtgctga agaagctgaa ccacaagaac
     2701 atcgtcaagc tgttcgccat cgaggaagaa accactaccc gccacaaggt gctgatcatg
     2761 gagttctgcc cctgcggttc cctgtacact gtcctggaag agccttccaa cgcttacggt
     2821 ctgcccgaga gcgagttcct gatcgtcctg cgtgacgtgg tcggtggcat gaaccacctg
     2881 agggaaaacg gcatcgtgca cagagacatc aagccaggaa acatcatgcg cgtgatcggc
     2941 gaggacggac agagcgtcta caagctgacc gacttcggcg ctgccagaga gctggaagac
     3001 gacgaacagt tcgtctctct gtacggaact gaagagtacc tgcaccctga catgtacgag
     3061 agggccgtgc tgagaaagga ccaccagaag aagtacggtg ctaccgtgga cctgtggtct
     3121 atcggcgtca ccttctacca cgctgccact ggatcactgc ctttccgccc cttcgaaggt
     3181 cctcgccgta acaaggaggt catgtacaag atcatcactg gcaagccctc tggcgccatc
     3241 tcaggagtcc agaaggctga aaacggtccc atcgactgga gcggcgacat gccagtgagc
     3301 tgctctctgt cccgcggtct gcaagtgctg ctgaccccag tcctggccaa catcctggaa
     3361 gctgaccagg agaagtgctg gggtttcgac cagttcttcg ctgaaacctc cgacatcctg
     3421 cacagaatgg tcatccacgt cttcagcctg caacagatga ctgcccacaa gatttacatc
     3481 cactcttaca acactgctac tatcttccac gaactggtct acaagcagac caagatcatc
     3541 tccagcaacc aggaactgat ctacgaggga aggagactgg tgctggagcc cggtcgcctg
     3601 gctcagcact tcccaaagac taccgaagag aaccctatct tcgtggtgtc ccgcgagcca
     3661 ctgaacacta tcggcctgat ctacgagaag atcagcctgc ccaaggtcca cccacgttac
     3721 gacctggacg gcgacgctag catggctaag gccatcaccg gagtggtctg ctacgcttgc
     3781 cgcatcgctt caactctgct gctgtaccag gaactgatgc gcaagggaat ccgttggctg
     3841 atcgagctga tcaaggacga ctacaacgaa actgtgcaca agaagaccga ggtggtcatc
     3901 actctggact tctgcatccg taacatcgaa aagaccgtga aggtctacga gaagctgatg
     3961 aagatcaacc tggaggctgc cgaactggga gagatcagcg acatccacac taagctgctg
     4021 cgcctgtctt catcccaggg cactatcgaa acctccctgc aagacatcga ctcccgtctg
     4081 agcccaggag gttcactggc tgacgcttgg gctcaccagg aaggcaccca ccctaaggac
     4141 aggaacgtgg agaagctgca agtcctgctg aactgcatga ctgaaatcta ctaccagttc
     4201 aagaaggaca aggccgagcg ccgtctggct tacaacgaag agcagatcca caagttcgac
     4261 aagcagaagc tgtactacca cgccaccaag gctatgactc acttcaccga cgaatgcgtc
     4321 aagaagtacg aggctttcct gaacaagagc gaagagtgga tcaggaagat gctgcacctg
     4381 agaaagcagc tgctgtccct gaccaaccag tgcttcgaca tcgaagagga agtgagcaag
     4441 taccaggagt acaccaacga gctgcaagag actctgcctc agaagatgtt cactgccagc
     4501 tctggtatca agcacactat gaccccaatc tacccttcat ccaacaccct ggtggaaatg
     4561 actctgggta tgaagaagct gaaggaggaa atggagggcg tggtcaagga actggctgag
     4621 aacaaccaca tcttggaaag attcggtagc ctgacgatgg atggtggact gagaaacgtg
     4681 gactgtctgt aaaagcttgt cgagaagtac tagaggatca taatcagcca taccacattt
     4741 gtagaggttt tacttgcttt aaaaaacctc ccacacctcc ccctgaacct gaaacataaa
     4801 atgaatgcaa ttgttgttgt taacttgttt attgcagctt ataatggtta caaataaagc
     4861 aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg
     4921 tccaaactca tcaatgtatc ttatcatgtc tggatctgat cactgcttga gcctaggaga
     4981 tccgaaccag ataagtgaaa tctagttcca aactattttg tcatttttaa ttttcgtatt
     5041 agcttacgac gctacaccca gttcccatct attttgtcac tcttccctaa ataatcctta
     5101 aaaactccat ttccacccct cccagttccc aactattttg tccgcccaca gcggggcatt
     5161 tttcttcctg ttatgttttt aatcaaacat cctgccaact ccatgtgaca aaccgtcatc
     5221 ttcggctact ttttctctgt cacagaatga aaatttttct gtcatctctt cgttattaat
     5281 gtttgtaatt gactgaatat caacgcttat ttgcagcctg aatggcgaat gggacgcgcc
     5341 ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact
     5401 tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc
     5461 cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt
     5521 acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc
     5581 ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttcttaatag tggactcttg
     5641 ttccaaactg gaacaacact caaccctatc tcggtctatt cttttgattt ataagggatt
     5701 ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat
     5761 tttaacaaaa tattaacgct tacaatttag gtggcacttt tcggggaaat gtgcgcggaa
     5821 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac
     5881 cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg
     5941 tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc
     6001 tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg
     6061 atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga
     6121 gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc
     6181 aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag
     6241 aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga
     6301 gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg
     6361 cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga
     6421 atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt
     6481 tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact
     6541 ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt
     6601 ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg
     6661 ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta
     6721 tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac
     6781 tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat ttttaattta
     6841 aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt
     6901 tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt
     6961 tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt
     7021 gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc
     7081 agataccaaa tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg
     7141 tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg
     7201 ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt
     7261 cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac
     7321 tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg
     7381 acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg
     7441 gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat
     7501 ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt
     7561 tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg
     7621 attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa
     7681 cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga gcgcctgatg cggtattttc
     7741 tccttacgca tctgtgcggt atttcacacc gcatagacca gccgcgtaac ctggcaaaat
     7801 cggttacggt tgagtaataa atggatgccc tgcgtaagcg ggtgtgggcg gacaataaag
     7861 tcttaaactg aacaaaatag atctaaacta tgacaataaa gtcttaaact agacagaata
     7921 gttgtaaact gaaatcagtc cagttatgct gtgaaaaagc atactggact tttgttatgg
     7981 ctaaagcaaa ctcttcattt tctgaagtgc aaattgcccg tcgtattaaa gaggggcgtg
     8041 gccaagggca tggtaaagac tatattcgcg gcgttgtgac aatttaccga acaactccgc
     8101 ggccgggaag ccgatctcgg cttgaacgaa ttgttaggtg gcggtacttg ggtcgatatc
     8161 aaagtgcatc acttcttccc gtatgcccaa ctttgtatag agagccactg cgggatcgtc
     8221 accgtaatct gcttgcacgt agatcacata agcaccaagc gcgttggcct catgcttgag
     8281 gagattgatg agcgcggtgg caatgccctg cctccggtgc tcgccggaga ctgcgagatc
     8341 atagatatag atctcactac gcggctgctc aaacttgggc agaacgtaag ccgcgagagc
     8401 gccaacaacc gcttcttggt cgaaggcagc aagcgcgatg aatgtcttac tacggagcaa
     8461 gttcccgagg taatcggagt ccggctgatg ttgggagtag gtggctacgt ctccgaactc
     8521 acgaccgaaa agatcaagag cagcccgcat ggatttgact tggtcagggc cgagcctaca
     8581 tgtgcgaatg atgcccatac ttgagccacc taactttgtt ttagggcgac tgccctgctg
     8641 cgtaacatcg ttgctgctgc gtaacatcgt tgctgctcca taacatcaaa catcgaccca
     8701 cggcgtaacg cgcttgctgc ttggatgccc gaggcataga ctgtacaaaa aaacagtcat
     8761 aacaagccat g
//
