LOCUS       sequence-369641-        5546 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Expresses human ATG13 HORMA L151D mutant in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5546)
  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 5546)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 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)-L151D
FEATURES             Location/Qualifiers
     source          1..5546
                     /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      1752..1761
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1767..1790
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             1827..1850
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             1857..1880
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             2410..2421
                     /codon_start=1
                     /product="WELQut protease recognition and cleavage site"
                     /label=WELQut site
                     /translation="WELQ"
     primer_bind     complement(2567..2586)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    2632..2687
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(2686..2705)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(3048..3064)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3048..3064)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3061..3083)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3072..3088
                     /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(3096..3102,3103..3120,3121..3126))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3141..3162
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        3221..3416
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      3267..3402
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    3422..3556
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3459..3478)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3513..3532
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3624..3641)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3795..4383)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3875..3894)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4554..5345,5346..5414))
                     /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     5177..5196
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5415..5519)
                     /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 ggggnsgcgc gccrggsggg gsggggsggg gsgrggggsg
      541 gggsggggcg aggcggagag 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 agggctcggg 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 tgccaccatg gctagctgga gccacccgca gttcgagaaa ggtggaggtg
     1801 cccgaggtgg atcgggaggt ggatcgtgga gccacccgca gttcgaaaaa ggattcgatt
     1861 acaaggatga cgatgacaag ggtaccgaga ccgacctcaa cagccaggac cgtaaagacc
     1921 tcgacaagtt cattaagttt ttcgccctca agaccgtgca ggtgatcgtg caggcccggc
     1981 tgggagagaa aatctgcaca cgcagctcct cttcacccac cggttctgac tggttcaacc
     2041 tggctattaa ggatatccct gaagtgactc acgaagcaaa gaaagcactc gctggacagc
     2101 tgccagcagt gggtcgttca atgtgtgtcg agatctccct gaagacatct gaaggagact
     2161 caatggagct cgaaatttgg tgcctggaga tgaacgaaaa gtgtgataaa gagatcaaag
     2221 tgagctacac cgtctataat cgtctgtccc tgctcctgaa atctctcctg gccatcactc
     2281 gggtgacacc tgcttaccgc ctcagtagga agcagggtca tgaatatgtg attgattaca
     2341 ggatctattt cggcgaggtc cagctctccg gactgggtga aggctttcag actgtgcgtg
     2401 tcggaaccgt gggaactcca gtcggcacaa ttaccctctc ttgcgcatac cgtatcaatc
     2461 tggccttcat gtcataactc gaggaattca ctcctcaggt gcaggctgcc tatcagaagg
     2521 tggtggctgg tgtggccaat gccctggctc acaaatacca ctgagatctt tttccctctg
     2581 ccaaaaatta tggggacatc atgaagcccc ttgagcatct gacttctggc taataaagga
     2641 aatttatttt cattgcaata gtgtgttgga attttttgtg tctctcactc ggaaggacat
     2701 atgggagggc aaatcattta aaacatcaga atgagtattt ggtttagagt ttggcaacat
     2761 atgcccatat gctggctgcc atgaacaaag gttggctata aagaggtcat cagtatatga
     2821 aacagccccc tgctgtccat tccttattcc atagaaaagc cttgacttga ggttagattt
     2881 tttttatatt ttgttttgtg ttattttttt ctttaacatc cctaaaattt tccttacatg
     2941 ttttactagc cagatttttc ctcctctcct gactactccc agtcatagct gtccctcttc
     3001 tcttatggag atccctcgac ctgcagccca agcttggcgt aatcatggtc atagctgttt
     3061 cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag
     3121 tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg
     3181 cccgctttcc agtcgggaaa cctgtcgtgc cagcggatcc gcatctcaat tagtcagcaa
     3241 ccatagtccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt
     3301 ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc gcctcggcct
     3361 ctgagctatt ccagaagtag tgaggaggct tttttggagg cctaggcttt tgcaaaaagc
     3421 taacttgttt attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac
     3481 aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc
     3541 ttatcatgtc tggatccgct gcattaatga atcggccaac gcgcggggag aggcggtttg
     3601 cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg
     3661 cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat
     3721 aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc
     3781 gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc
     3841 tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga
     3901 agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt
     3961 ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg
     4021 taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc
     4081 gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg
     4141 gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc
     4201 ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg
     4261 ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc
     4321 gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct
     4381 caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt
     4441 taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa
     4501 aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa
     4561 tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc
     4621 tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct
     4681 gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca
     4741 gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt
     4801 aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt
     4861 gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc
     4921 ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc
     4981 tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt
     5041 atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact
     5101 ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc
     5161 ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt
     5221 ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg
     5281 atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct
     5341 gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa
     5401 tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt
     5461 ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc
     5521 acatttcccc gaaaagtgcc acctgg
//
