LOCUS       sequence-430395-        5898 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For expression in E. coli of NAP1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5898)
  AUTHORS   Adriaenssens E, Nguyen TN, Sawa-Makarska J, Khuu G, Schuschnig M, 
            Shoebridge S, Watts EM, Csalyi KD, Padman BS, Lazarou M, Martens S
  TITLE     Control of mitophagy initiation and progression by the TBK1 adaptors
            NAP1 and SINTBAD
  JOURNAL   bioRxiv September 25, 202
REFERENCE   2  (bases 1 to 5898)
  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: "bioRxiv 
            September 25, 202"
COMMENT     Sequence Label: pGEX-4T1-MBP-TEV-NAP1 (WT)
FEATURES             Location/Qualifiers
     source          1..5898
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    22..38
                     /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     27..49
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             61..1161
                     /codon_start=1
                     /gene="malE (mutated)"
                     /product="maltose binding protein from E. coli"
                     /label=MBP
                     /note="This version of the gene does not encode a signal 
                     sequence, so MBP will remain in the cytosol."
                     /translation="MKIEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLE
                     EKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKL
                     IAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIA
                     ADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGE
                     TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFL
                     ENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAF
                     WYAVRTAVINAASGRQTVDEALKDAQT"
     primer_bind     1135..1158
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             1186..1206
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     primer_bind     complement(2452..2474)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2619..2637)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2705..2809
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2810..2878,2879..3670)
                     /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(3028..3047)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3841..4429
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4330..4349
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4583..4600
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        4673..4750
                     /gene="lacI"
                     /label=lacI promoter
     CDS             4751..5833
                     /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(4770..4789)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(5882..5887,5888..5898,1..7,8..14)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
ORIGIN
        1 cggctcgtat gttgtgtgga attgtgagcg gataacaatt tcacacagga aacagtattc
       61 atgaaaatcg aagaaggtaa actggtaatc tggattaacg gcgataaagg ctataacggt
      121 ctcgctgaag tcggtaagaa attcgagaaa gataccggaa ttaaagtcac cgttgagcat
      181 ccggataaac tggaagagaa attcccacag gttgcggcaa ctggcgatgg ccctgacatt
      241 atcttctggg cacacgaccg ctttggtggc tacgctcaat ctggcctgtt ggctgaaatc
      301 accccggaca aagcgttcca ggacaagctg tatccgttta cctgggatgc cgtacgttac
      361 aacggcaagc tgattgctta cccgatcgct gttgaagcgt tatcgctgat ttataacaaa
      421 gatctgctgc cgaacccgcc aaaaacctgg gaagagatcc cggcgctgga taaagaactg
      481 aaagcgaaag gtaagagcgc gctgatgttc aacctgcaag aaccgtactt cacctggccg
      541 ctgattgctg ctgacggggg ttatgcgttc aagtatgaaa acggcaagta cgacattaaa
      601 gacgtgggcg tggataacgc tggcgcgaaa gcgggtctga ccttcctggt tgacctgatt
      661 aaaaacaaac acatgaatgc agacaccgat tactccatcg cagaagctgc ctttaataaa
      721 ggcgaaacag cgatgaccat caacggcccg tgggcatggt ccaacatcga caccagcaaa
      781 gtgaattatg gtgtaacggt actgccgacc ttcaagggtc aaccatccaa accgttcgtt
      841 ggcgtgctga gcgcaggtat taacgccgcc agtccgaaca aagagctggc aaaagagttc
      901 ctcgaaaact atctgctgac tgatgaaggt ctggaagcgg ttaataaaga caaaccgctg
      961 ggtgccgtag cgctgaagtc ttacgaggaa gagttggcga aagatccacg tattgccgcc
     1021 actatggaaa acgcccagaa aggtgaaatc atgccgaaca tcccgcagat gtccgctttc
     1081 tggtatgccg tgcgtactgc ggtgatcaac gccgccagcg gtcgtcagac tgtcgatgaa
     1141 gccctgaaag acgcgcagac taattcgagc tcggctccgg aattcgaaaa cctgtatttt
     1201 cagggaggag gaatggatgc actggtagaa gatgatatct gtattctgaa tcatgaaaaa
     1261 gcccataaga gagatacagt gactccagtt tcaatatatt caggagatga atctgttgct
     1321 tcccattttg ctcttgtcac tgcatatgaa gacatcaaaa aacgacttaa ggattcagag
     1381 aaagagaact ctttgttaaa gaagagaata agatttttgg aagaaaagct aatagctcga
     1441 tttgaagaag aaacaagttc cgtgggacga gaacaagtaa ataaggccta tcatgcatat
     1501 cgagaggttt gcattgatag agataatttg aagagcaaac tggacaaaat gaataaagac
     1561 aactctgaat ctttgaaagt attgaatgag cagctacaat ctaaagaagt agaactcctc
     1621 cagctgagga cagaggtgga aactcagcag gtgatgagga atttaaatcc accttcatca
     1681 aactgggagg tggaaaagtt gagctgtgac ctgaagatcc atggtttgga acaagagctg
     1741 gaactgatga ggaaagaatg tagcgatctc aaaatagaac tacagaaagc caaacaaacg
     1801 gatccatatc aggaagacaa tctgaagagc agagatctcc aaaaactaag catttcaagt
     1861 gataatatgc agcatgcata ctgggaactg aagagagaaa tgtctaattt acatctggtg
     1921 actcaagtac aagctgaact actaagaaaa ctgaaaacct caactgcaat caagaaagcc
     1981 tgtgcccctg taggatgcag tgaagacctt ggaagagaca gcacaaaact gcacttgatg
     2041 aattttactg caacatacac aagacatccc cctctcttac caaatggcaa agctctttgt
     2101 cataccacat cttccccttt accaggagat gtaaaggttt tatcagagaa agcaatcctc
     2161 caatcatgga cagacaatga gagatccatt cctaatgatg gtacatgctt tcaggaacac
     2221 agttcttatg gcagaaattc tctggaagac aattcctggg tatttccaag tcctcctaaa
     2281 tcaagtgaga cagcatttgg ggaaactaaa actaaaactt tgcctttacc caaccttcca
     2341 ccactgcatt acttggatca acataatcag aactgccttt ataagaatta agcggccgca
     2401 tcgtgactga ctgacgatct gcctcgcgcg tttcggtgat gacggtgaaa acctctgaca
     2461 catgcagctc ccggagacgg tcacagcttg tctgtaagcg gatgccggga gcagacaagc
     2521 ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc gcagccatga cccagtcacg
     2581 tagcgatagc ggagtgtata attcttgaag acgaaagggc ctcgtgatac gcctattttt
     2641 ataggttaat gtcatgataa taatggtttc ttagacgtca ggtggcactt ttcggggaaa
     2701 tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat
     2761 gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca
     2821 acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca
     2881 cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta
     2941 catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt
     3001 tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc gtgttgacgc
     3061 cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc
     3121 accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc
     3181 cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa
     3241 ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga
     3301 accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgcagcaat
     3361 ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca
     3421 attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc
     3481 ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat
     3541 tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag
     3601 tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa
     3661 gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca
     3721 tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc
     3781 ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
     3841 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
     3901 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
     3961 cagcagagcg cagataccaa atactgtcct tctagtgtag ccgtagttag gccaccactt
     4021 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
     4081 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
     4141 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
     4201 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
     4261 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
     4321 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
     4381 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
     4441 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
     4501 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
     4561 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcctgat
     4621 gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcataaatt ccgacaccat
     4681 cgaatggcgc aaaacctttc gcggtatggc atgatagcgc ccggaagaga gtcaattcag
     4741 ggtggtgaat gtgaaaccag taacgttata cgatgtcgca gagtatgccg gtgtctctta
     4801 tcagaccgtt tcccgcgtgg tgaaccaggc cagccacgtt tctgcgaaaa cgcgggaaaa
     4861 agtggaagcg gcgatggcgg agctgaatta cattcccaac cgcgtggcac aacaactggc
     4921 gggcaaacag tcgttgctga ttggcgttgc cacctccagt ctggccctgc acgcgccgtc
     4981 gcaaattgtc gcggcgatta aatctcgcgc cgatcaactg ggtgccagcg tggtggtgtc
     5041 gatggtagaa cgaagcggcg tcgaagcctg taaagcggcg gtgcacaatc ttctcgcgca
     5101 acgcgtcagt gggctgatca ttaactatcc gctggatgac caggatgcca ttgctgtgga
     5161 agctgcctgc actaatgttc cggcgttatt tcttgatgtc tctgaccaga cacccatcaa
     5221 cagtattatt ttctcccatg aagacggtac gcgactgggc gtggagcatc tggtcgcatt
     5281 gggtcaccag caaatcgcgc tgttagcggg cccattaagt tctgtctcgg cgcgtctgcg
     5341 tctggctggc tggcataaat atctcactcg caatcaaatt cagccgatag cggaacggga
     5401 aggcgactgg agtgccatgt ccggttttca acaaaccatg caaatgctga atgagggcat
     5461 cgttcccact gcgatgctgg ttgccaacga tcagatggcg ctgggcgcaa tgcgcgccat
     5521 taccgagtcc gggctgcgcg ttggtgcgga tatctcggta gtgggatacg acgataccga
     5581 agacagctca tgttatatcc cgccgttaac caccatcaaa caggattttc gcctgctggg
     5641 gcaaaccagc gtggaccgct tgctgcaact ctctcagggc caggcggtga agggcaatca
     5701 gctgttgccc gtctcactgg tgaaaagaaa aaccaccctg gcgcccaata cgcaaaccgc
     5761 ctctccccgc gcgttggccg attcattaat gcagctggca cgacaggttt cccgactgga
     5821 aagcgggcag tgagcgcaac gcaattaatg taagttagct cactcattag gcaccccagg
     5881 ctttacactt tatgcttc
//
