LOCUS       sequence-369676-        5545 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses human ATG13 HORMA Y115D mutant in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5545)
  AUTHORS   Ren X, Nguyen TN, Lam WK, Buffalo CZ, Lazarou M, Yokom AL, Hurley JH
  TITLE     Structural basis for ATG9A recruitment to the ULK1 complex in 
            mitophagy initiation.
  JOURNAL   Sci Adv. 2023 Feb 15;9(7):eadg2997. doi: 10.1126/sciadv.adg2997. 
            Epub 2023 Feb 15.
  PUBMED    36791199
REFERENCE   2  (bases 1 to 5545)
  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: "Sci Adv."; 
            date: "2023-02-15"; volume: "9(7):eadg2997. doi"; pages: " 
            10.1126/sciadv.adg2997. Epub 2023 Feb 15"
COMMENT     Sequence Label: pCAG-OSF-ATG13 (2-197)-Y115D
FEATURES             Location/Qualifiers
     source          1..5545
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        4..383
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     intron          661..1677
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1601..1623
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1685..1704
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      1751..1760
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1766..1789
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             1826..1849
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             1856..1879
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             2409..2420
                     /codon_start=1
                     /product="WELQut protease recognition and cleavage site"
                     /label=WELQut site
                     /translation="WELQ"
     primer_bind     complement(2566..2585)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    2631..2686
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(2685..2704)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(3047..3063)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3047..3063)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3060..3082)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3071..3087
                     /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)."
     promoter        complement(join(3095..3101,3102..3119,3120..3125))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3140..3161
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        3220..3415
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      3266..3401
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    3421..3555
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3458..3477)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3512..3531
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3623..3640)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3794..4382)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3874..3893)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4553..5344,5345..5413))
                     /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     5176..5195
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5414..5518)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgacattg attattgact agttattaat agtaatcaat tacggggtca ttagttcata
       61 gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc
      121 ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag
      181 ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac
      241 atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg
      301 cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg
      361 tattagtcat cgctattacc atggtcgagg tgagccccac gttctgcttc actctcccca
      421 tctccccccc ctccccaccc ccaattttgt atttatttat tttttaatta ttttgtgcag
      481 cgatgggggc gggggggggg ggggggcgcg ccrggggggg sggggsgggg sgrggggsgg
      541 ggsggggsgr ggcggagagg tgcggcggca gccaatcaga gcggcgcgct ccgaaagttt
      601 ccttttatgg cgaggcggcg gcggcggcgg ccctataaaa agcgaagcgc gcggcgggcg
      661 ggagtcgctg cgcgctgcct tcgccccgtg ccccgctccg ccgccgcctc gcgccgcccg
      721 ccccggctct gactgaccgc gttactccca caggtgagcg ggcgggacgg cccttctcct
      781 ccgggctgta attagcgctt ggtttaatga cggcttgttt cttttctgtg gctgcgtgaa
      841 agccttgagg ggctccggga gggccctttg tgcgggggga gcggctcggg gggtgcgtgc
      901 gtgtgtgtgt gcgtggggag cgccgcgtgc ggctccgcgc tgcccggcgg ctgtgagcgc
      961 tgcgggcgcg gcgcggggct ttgtgcgctc cgcagtgtgc gcgaggggag cgcggccggg
     1021 ggcggtgccc cgcggtgcgg ggggggctgc gaggggaaca aaggctgcgt gcggggtgtg
     1081 tgcgtggggg ggtgagcagg gggtgtgggc gcgtcggtcg ggctgcaacc ccccctgcac
     1141 ccccctcccc gagttgctga gcacggcccg gcttcgggtg cggggctccg tacggggcgt
     1201 ggcgcggggc tcgccgtgcc gggcgggggg tggcggcagg tgggggtgcc gggcggggcg
     1261 gggccgcctc gggccgggga gggctcgggg gaggggcgcg gcggcccccg gagcgccggc
     1321 ggctgtcgag gcgcggcgag ccgcagccat tgccttttat ggtaatcgtg cgagagggcg
     1381 cagggacttc ctttgtccca aatctgtgcg gagccgaaat ctgggaggcg ccgccgcacc
     1441 ccctctagcg ggcgcggggc gaagcggtgc ggcgccggca ggaaggaaat gggcggggag
     1501 ggccttcgtg cgtcgccgcg ccgccgtccc cttctccctc tccagcctcg gggctgtccg
     1561 cggggggacg gctgccttcg ggggggacgg ggcagggcgg ggttcggctt ctggcgtgtg
     1621 accggcggct ctagagcctc tgctaaccat gttcatgcct tcttcttttt cctacagctc
     1681 ctgggcaacg tgctggttat tgtgctgtct catcattttg gcaaagaatt cctcgatcga
     1741 cggtatcgat gccaccatgg ctagctggag ccacccgcag ttcgagaaag gtggaggtgc
     1801 ccgaggtgga tcgggaggtg gatcgtggag ccacccgcag ttcgaaaaag gattcgatta
     1861 caaggatgac gatgacaagg gtaccgagac cgacctcaac agccaggacc gtaaagacct
     1921 cgacaagttc attaagtttt tcgccctcaa gaccgtgcag gtgatcgtgc aggcccggct
     1981 gggagagaaa atctgcacac gcagctcctc ttcacccacc ggttctgact ggttcaacct
     2041 ggctattaag gatatccctg aagtgactca cgaagcaaag aaagcactcg ctggacagct
     2101 gccagcagtg ggtcgttcaa tgtgtgtcga gatctccctg aagacatctg aaggagactc
     2161 aatggagctc gaaatttggt gcctggagat gaacgaaaag tgtgataaag agatcaaagt
     2221 gagcgacacc gtctataatc gtctgtccct gctcctgaaa tctctcctgg ccatcactcg
     2281 ggtgacacct gcttaccgcc tcagtaggaa gcagggtcat gaatatgtga ttctgtacag
     2341 gatctatttc ggcgaggtcc agctctccgg actgggtgaa ggctttcaga ctgtgcgtgt
     2401 cggaaccgtg ggaactccag tcggcacaat taccctctct tgcgcatacc gtatcaatct
     2461 ggccttcatg tcataactcg aggaattcac tcctcaggtg caggctgcct atcagaaggt
     2521 ggtggctggt gtggccaatg ccctggctca caaataccac tgagatcttt ttccctctgc
     2581 caaaaattat ggggacatca tgaagcccct tgagcatctg acttctggct aataaaggaa
     2641 atttattttc attgcaatag tgtgttggaa ttttttgtgt ctctcactcg gaaggacata
     2701 tgggagggca aatcatttaa aacatcagaa tgagtatttg gtttagagtt tggcaacata
     2761 tgcccatatg ctggctgcca tgaacaaagg ttggctataa agaggtcatc agtatatgaa
     2821 acagccccct gctgtccatt ccttattcca tagaaaagcc ttgacttgag gttagatttt
     2881 ttttatattt tgttttgtgt tatttttttc tttaacatcc ctaaaatttt ccttacatgt
     2941 tttactagcc agatttttcc tcctctcctg actactccca gtcatagctg tccctcttct
     3001 cttatggaga tccctcgacc tgcagcccaa gcttggcgta atcatggtca tagctgtttc
     3061 ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt
     3121 gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc
     3181 ccgctttcca gtcgggaaac ctgtcgtgcc agcggatccg catctcaatt agtcagcaac
     3241 catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc
     3301 tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg cctcggcctc
     3361 tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt gcaaaaagct
     3421 aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca
     3481 aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct
     3541 tatcatgtct ggatccgctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc
     3601 gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
     3661 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
     3721 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     3781 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     3841 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     3901 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     3961 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     4021 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     4081 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     4141 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     4201 tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc
     4261 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     4321 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     4381 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     4441 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     4501 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     4561 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     4621 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
     4681 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
     4741 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     4801 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     4861 ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     4921 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     4981 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     5041 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     5101 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     5161 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     5221 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     5281 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     5341 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     5401 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     5461 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     5521 catttccccg aaaagtgcca cctgg
//
