LOCUS       sequence-448033-        5506 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of recombinant protein for purification (co-expression 
            with ATG13).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5506)
  AUTHORS   Adriaenssens E, Schaar S, Cook ASI, Stuke JFM, Sawa-Makarska J, 
            Nguyen TN, Ren X, Schuschnig M, Romanov J, Khuu G, Uoselis L, 
            Lazarou M, Hummer G, Hurley JH, Martens S
  TITLE     Reconstitution of BNIP3/NIX-mitophagy initiation reveals 
            hierarchical flexibility of the autophagy machinery.
  JOURNAL   Nat Cell Biol. 2025 Jul 25. doi: 10.1038/s41556-025-01712-y.
  PUBMED    40715440
REFERENCE   2  (bases 1 to 5506)
  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: "Nat Cell 
            Biol. 2025 Jul 25. doi: 10.1038/s41556-025-01712-y."
COMMENT     Sequence Label: pCAG_ATG101
FEATURES             Location/Qualifiers
     source          1..5506
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        4..383
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        385..659
                     /label=chicken beta-actin promoter
     intron          660..1676
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1600..1622
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1684..1703
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     primer_bind     complement(2527..2546)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    2592..2647
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(2646..2665)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(3008..3024)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3008..3024)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3021..3043)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3032..3048
                     /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(3056..3062,3063..3080,3081..3086))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3101..3122
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        3181..3376
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      3227..3362
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    3382..3516
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3419..3438)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3473..3492
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3584..3601)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3755..4343)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3835..3854)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4514..5305,5306..5374))
                     /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     5137..5156
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5375..5479)
                     /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 gggcgcgcgc caggcggggc ggggcggggc gaggggcggg
      541 gcggggcgag gcggagaggt gcggcggcag ccaatcagag cggcgcgctc cgaaagtttc
      601 cttttatggc gaggcggcgg cggcggcggc cctataaaaa gcgaagcgcg cggcgggcgg
      661 gagtcgctgc gcgctgcctt cgccccgtgc cccgctccgc cgccgcctcg cgccgcccgc
      721 cccggctctg actgaccgcg ttactcccac aggtgagcgg gcgggacggc ccttctcctc
      781 cgggctgtaa ttagcgcttg gtttaatgac ggcttgtttc ttttctgtgg ctgcgtgaaa
      841 gccttgaggg gctccgggag ggccctttgt gcggggggag cggctcgggg ggtgcgtgcg
      901 tgtgtgtgtg cgtggggagc gccgcgtgcg gctccgcgct gcccggcggc tgtgagcgct
      961 gcgggcgcgg cgcggggctt tgtgcgctcc gcagtgtgcg cgaggggagc gcggccgggg
     1021 gcggtgcccc gcggtgcggg gggggctgcg aggggaacaa aggctgcgtg cggggtgtgt
     1081 gcgtgggggg gtgagcaggg ggtgtgggcg cgtcggtcgg gctgcaaccc cccctgcacc
     1141 cccctccccg agttgctgag cacggcccgg cttcgggtgc ggggctccgt acggggcgtg
     1201 gcgcggggct cgccgtgccg ggcggggggt ggcggcaggt gggggtgccg ggcggggcgg
     1261 ggccgcctcg ggccggggag ggctcggggg aggggcgcgg cggcccccgg agcgccggcg
     1321 gctgtcgagg cgcggcgagc cgcagccatt gccttttatg gtaatcgtgc gagagggcgc
     1381 agggacttcc tttgtcccaa atctgtgcgg agccgaaatc tgggaggcgc cgccgcaccc
     1441 cctctagcgg gcgcggggcg aagcggtgcg gcgccggcag gaaggaaatg ggcggggagg
     1501 gccttcgtgc gtcgccgcgc cgccgtcccc ttctccctct ccagcctcgg ggctgtccgc
     1561 ggggggacgg ctgccttcgg gggggacggg gcagggcggg gttcggcttc tggcgtgtga
     1621 ccggcggctc tagagcctct gctaaccatg ttcatgcctt cttctttttc ctacagctcc
     1681 tgggcaacgt gctggttatt gtgctgtctc atcattttgg caaagaattc ctcgatcgac
     1741 ggtatcgatg ccaccatgaa ttgccggtca gaggtcctgg aagtgagcgt ggaaggtaga
     1801 caggtggaag aggctatgct ggccgtgctg catacagtgc tgctgcacag atcaactggc
     1861 aagttccatt acaagaaaga aggaacatat tccatcggaa ccgtgggtac tcaggacgtc
     1921 gattgcgact tcattgattt tacttacgtg cgcgtcagct ccgaggaact ggacagagca
     1981 ctccgtaagg tggtcggaga attcaaagat gccctgagga acagtggcgg agacggtctg
     2041 ggccagatga gcctcgagtt ctatcagaag aaaaagtcta gatggccatt ttcagatgaa
     2101 tgtatcccct gggaagtgtg gaccgtgaag gtccacgtgg tcgccctggc tactgagcag
     2161 gaacggcaga tctgccgcga gaaagtgggc gaaaagctct gtgagaagat catcaacatc
     2221 gtggaagtca tgaaccgtca tgagtacctg cccaagatgc ctacacagag cgaggtggat
     2281 aatgtcttcg acaccggtct gcgggatgtg cagccctacc tctataagat ctcctttcag
     2341 attacagacg ctctgggcac ctctgtcacc actacaatgc gcaggctcat caaagacacc
     2401 ctggcactct aacccgggtg atatcttaat taattaactc gaggaattca ctcctcaggt
     2461 gcaggctgcc tatcagaagg tggtggctgg tgtggccaat gccctggctc acaaatacca
     2521 ctgagatctt tttccctctg ccaaaaatta tggggacatc atgaagcccc ttgagcatct
     2581 gacttctggc taataaagga aatttatttt cattgcaata gtgtgttgga attttttgtg
     2641 tctctcactc ggaaggacat atgggagggc aaatcattta aaacatcaga atgagtattt
     2701 ggtttagagt ttggcaacat atgcccatat gctggctgcc atgaacaaag gttggctata
     2761 aagaggtcat cagtatatga aacagccccc tgctgtccat tccttattcc atagaaaagc
     2821 cttgacttga ggttagattt tttttatatt ttgttttgtg ttattttttt ctttaacatc
     2881 cctaaaattt tccttacatg ttttactagc cagatttttc ctcctctcct gactactccc
     2941 agtcatagct gtccctcttc tcttatggag atccctcgac ctgcagccca agcttggcgt
     3001 aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca
     3061 tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat
     3121 taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagcggatcc
     3181 gcatctcaat tagtcagcaa ccatagtccc gcccctaact ccgcccatcc cgcccctaac
     3241 tccgcccagt tccgcccatt ctccgcccca tggctgacta atttttttta tttatgcaga
     3301 ggccgaggcc gcctcggcct ctgagctatt ccagaagtag tgaggaggct tttttggagg
     3361 cctaggcttt tgcaaaaagc taacttgttt attgcagctt ataatggtta caaataaagc
     3421 aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg
     3481 tccaaactca tcaatgtatc ttatcatgtc tggatccgct gcattaatga atcggccaac
     3541 gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc
     3601 tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt
     3661 tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg
     3721 ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg
     3781 agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat
     3841 accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta
     3901 ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct
     3961 gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc
     4021 ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa
     4081 gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg
     4141 taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag
     4201 tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt
     4261 gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta
     4321 cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc
     4381 agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca
     4441 cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa
     4501 cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat
     4561 ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct
     4621 taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt
     4681 tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat
     4741 ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta
     4801 atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg
     4861 gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt
     4921 tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg
     4981 cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg
     5041 taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc
     5101 ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa
     5161 ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac
     5221 cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt
     5281 ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg
     5341 gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa
     5401 gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata
     5461 aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgg
//
