LOCUS       sequence-417185-        5745 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of ATG13 (450-517) mutant.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5745)
  AUTHORS   Chen M, Ren X, Nguyen TN, Khuu G, Cook ASI, Lazarou M, Hurley JH
  TITLE     Structure and activation of the human autophagy-initiating 
            ULK1C:PI3KC3-C1 supercomplex
  JOURNAL   bioRxiv 2023.06.01.543278
REFERENCE   2  (bases 1 to 5745)
  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 
            2023.06.01.543278"
COMMENT     Sequence Label: pCAG-GST-ATG13 (450-517)-A452D/F453D
FEATURES             Location/Qualifiers
     source          1..5745
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        4..383
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        385..661
                     /label=chicken beta-actin promoter
     intron          662..1678
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1602..1624
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1686..1705
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      1753..1762
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1759..2415
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MASPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNK
                     KFELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIR
                     YGVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVV
                     LYMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     2373..2395
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             2443..2463
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     primer_bind     complement(2766..2785)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    2831..2886
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(2885..2904)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(3247..3263)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3247..3263)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3260..3282)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3271..3287
                     /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(3295..3301,3302..3319,3320..3325))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3340..3361
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        3420..3615
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      3466..3601
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    3621..3755
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3658..3677)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3712..3731
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3823..3840)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3994..4582)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4074..4093)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4753..5544,5545..5613))
                     /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     5376..5395
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5614..5718)
                     /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 gggggsgcgc gccagggggg ggggggsggg gggagggggg
      541 gggsggggsg aggsggagag gtgcggcggc agccaatcag agcggcgcgc tccgaaagtt
      601 tccttttatg gcgaggcggc ggcggcggcg gccctataaa aagcgaagcg cgcggcgggc
      661 gggagtcgct gcgcgctgcc ttcgccccgt gccccgctcc gccgccgcct cgcgccgccc
      721 gccccggctc tgactgaccg cgttactccc acaggtgagc gggcgggacg gcccttctcc
      781 tccgggctgt aattagcgct tggtttaatg acggcttgtt tcttttctgt ggctgcgtga
      841 aagccttgag gggctccggg agggcccttt gtgcgggggg agcggctcgg ggggtgcgtg
      901 cgtgtgtgtg tgcgtgggga gcgccgcgtg cggctccgcg ctgcccggcg gctgtgagcg
      961 ctgcgggcgc ggcgcggggc tttgtgcgct ccgcagtgtg cgcgagggga gcgcggccgg
     1021 gggcggtgcc ccgcggtgcg gggggggctg cgaggggaac aaaggctgcg tgcggggtgt
     1081 gtgcgtgggg gggtgagcag ggggtgtggg cgcgtcggtc gggctgcaac cccccctgca
     1141 cccccctccc cgagttgctg agcacggccc ggcttcgggt gcggggctcc gtacggggcg
     1201 tggcgcgggg ctcgccgtgc cgggcggggg gtggcggcag gtgggggtgc cgggcggggc
     1261 ggggccgcct cgggccgggg rgggctcggg ggaggggcgc ggcggccccc ggagcgccgg
     1321 cggctgtcga ggcgcggcga gccgcagcca ttgcctttta tggtaatcgt gcgagagggc
     1381 gcagggactt cctttgtccc aaatctgtgc ggagccgaaa tctgggaggc gccgccgcac
     1441 cccctctagc gggcgcgggg cgaagcggtg cggcgccggc aggaaggaaa tgggcgggga
     1501 gggccttcgt gcgtcgccgc gccgccgtcc ccttctccct ctccagcctc ggggctgtcc
     1561 gcggggggac ggctgccttc gggggggacg gggcagggcg gggttcggct tctggcgtgt
     1621 gaccggcggc tctagagcct ctgctaacca tgttcatgcc ttcttctttt tcctacagct
     1681 cctgggcaac gtgctggtta ttgtgctgtc tcatcatttt ggcaaagaat tcctcgatcg
     1741 acggtatcga tagccaccat ggcatcccct atactaggtt attggaaaat taagggcctt
     1801 gtgcaaccca ctcgacttct tttggaatat cttgaagaaa aatatgaaga gcatttgtat
     1861 gagcgcgatg aaggtgataa atggcgaaac aaaaagtttg aattgggttt ggagtttccc
     1921 aatcttcctt attatattga tggtgatgtt aaattaacac agtctatggc catcatacgt
     1981 tatatagctg acaagcacaa catgttgggt ggttgtccaa aagagcgtgc agagatttca
     2041 atgcttgaag gagcggtttt ggatattaga tacggtgttt cgagaattgc atatagtaaa
     2101 gactttgaaa ctctcaaagt tgattttctt agcaagctac ctgaaatgct gaaaatgttc
     2161 gaagatcgtt tatgtcataa aacatattta aatggtgatc atgtaaccca tcctgacttc
     2221 atgttgtatg acgctcttga tgttgtttta tacatggacc caatgtgcct ggatgcgttc
     2281 ccaaaattag tttgttttaa aaaacgtatt gaagctatcc cacaaattga taagtacttg
     2341 aaatccagca agtatatagc atggcctttg cagggctggc aagccacgtt tggtggtggc
     2401 gaccatcctc caaaatcgga tgattacgat atcccaacga ccgaaaacct gtattttcag
     2461 ggtggtacca aacccgacga ttccaaggac gatatcctcc ctatggatct gggaaccttc
     2521 tacagggagt ttcagaatcc ccctcagctc agtagcctga gcattgacat cggtgcccag
     2581 tccatggctg aggacctcga ttctctgcct gaaaaactgg ctgtgcacga gaagaacgtc
     2641 agagaattcg acgcatttgt ggagaccctg cagtgactcg aggaattcac tcctcaggtg
     2701 caggctgcct atcagaaggt ggtggctggt gtggccaatg ccctggctca caaataccac
     2761 tgagatcttt ttccctctgc caaaaattat ggggacatca tgaagcccct tgagcatctg
     2821 acttctggct aataaaggaa atttattttc attgcaatag tgtgttggaa ttttttgtgt
     2881 ctctcactcg gaaggacata tgggagggca aatcatttaa aacatcagaa tgagtatttg
     2941 gtttagagtt tggcaacata tgcccatatg ctggctgcca tgaacaaagg ttggctataa
     3001 agaggtcatc agtatatgaa acagccccct gctgtccatt ccttattcca tagaaaagcc
     3061 ttgacttgag gttagatttt ttttatattt tgttttgtgt tatttttttc tttaacatcc
     3121 ctaaaatttt ccttacatgt tttactagcc agatttttcc tcctctcctg actactccca
     3181 gtcatagctg tccctcttct cttatggaga tccctcgacc tgcagcccaa gcttggcgta
     3241 atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat
     3301 acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt
     3361 aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agcggatccg
     3421 catctcaatt agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact
     3481 ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag
     3541 gccgaggccg cctcggcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc
     3601 ctaggctttt gcaaaaagct aacttgttta ttgcagctta taatggttac aaataaagca
     3661 atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt
     3721 ccaaactcat caatgtatct tatcatgtct ggatccgctg cattaatgaa tcggccaacg
     3781 cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct
     3841 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     3901 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     3961 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     4021 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     4081 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     4141 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     4201 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     4261 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     4321 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     4381 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt
     4441 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     4501 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     4561 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     4621 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     4681 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     4741 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     4801 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     4861 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     4921 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     4981 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     5041 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     5101 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     5161 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     5221 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     5281 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     5341 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     5401 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     5461 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     5521 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     5581 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     5641 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     5701 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgg
//
