LOCUS       sequence-400832-        7133 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid for the expression of GFP-Atg32 lacking the PsR domain in S.
            cerevisiae..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7133)
  AUTHORS   Xia X, Katzenell S, Reinhart EF, Bauer KM, Pellegrini M, Ragusa MJ
  TITLE     A pseudo-receiver domain in Atg32 is required for mitophagy.
  JOURNAL   Autophagy. 2018;14(9):1620-1628. doi: 10.1080/15548627.2018.1472838.
            Epub 2018 Aug 1.
  PUBMED    29909755
REFERENCE   2  (bases 1 to 7133)
  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: 
            "Autophagy."; date: "2018"; volume: "14(9):1620-1628. doi"; pages: "
            10.1080/15548627.2018.1472838. Epub 2018 Aug 1"
COMMENT     Sequence Label: GFP-Atg32-deltaPsR pXP721
FEATURES             Location/Qualifiers
     source          1..7133
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(171..962,963..1031))
                     /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     794..813
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1032..1136)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     1204..1222
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(1260..1282)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     1382..1401
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        1425..1866
                     /gene="S. cerevisiae GAL1"
                     /label=GAL1 promoter
                     /note="inducible promoter, regulated by Gal4"
     protein_bind    1425..1542
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     primer_bind     1837..1860
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     CDS             join(1896..1898,1899..2609)
                     /codon_start=1
                     /product="enhanced GFP (EGFP) derivative that exhibits 
                     robust folding and enhanced photostability (Zhong et al., 
                     2019)"
                     /label=SiriusGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(1938..1959)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     2546..2567
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2619..2639
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     terminator      3813..4060
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(3829..3847)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     protein_bind    4106..4139
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    5581..5614
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     complement(6143..6159)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6143..6159)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6156..6178)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6167..6183
                     /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(6191..6197,6198..6215,6216..6221))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6236..6257
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6374..6391)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6545..7133)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6625..6644)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
       61 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
      121 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
      181 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
      241 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
      301 atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
      361 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
      421 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
      481 attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt
      541 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
      601 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
      661 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt
      721 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
      781 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
      841 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
      901 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
      961 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
     1021 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
     1081 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
     1141 tttccccgaa aagtgccacc tgacgtctaa gaaaccatta ttatcatgac attaacctat
     1201 aaaaataggc gtatcacgag gccctttcgt ctcgcgcgtt tcggtgatga cggtgaaaac
     1261 ctctgacaca tgcagctccc ggagacggtc acagcttgtc tgtaagcgga tgccgggagc
     1321 agacaagccc gtcagggcgc gtcagcgggt gttggcgggt gtcggggctg gcttaactat
     1381 gcggcatcag agcagattgt actgagagtg caccaggcat atgacggatt agaagccgcc
     1441 gagcgggtga cagccctccg aaggaagact ctcctccgtg cgtcctcgtc ttcaccggtc
     1501 gcgttcctga aacgcagatg tgcctcgcgc cgcactgctc cgaacaataa agattctaca
     1561 atactagctt ttatggttat gaagaggaaa aattggcagt aacctggccc cacaaacctt
     1621 caaatgaacg aatcaaatta acaaccatag gatgataatg cgattagttt tttagcctta
     1681 tttctggggt aattaatcag cgaagcgatg atttttgatc tattaacaga tatataaatg
     1741 caaaaactgc ataaccactt taactaatac tttcaacatt ttcggtttgt attacttctt
     1801 attcaaatgt aataaaagta tcaacaaaaa attgttaata tacctctata ctttaacgtc
     1861 aaggagaaaa aaccactagt aacacaatgt cttctgtgag caagggcgag gagctgttca
     1921 ccggggtggt gcccatcctg gtcgagctgg acggcgacgt aaacggccac aagttcagcg
     1981 tgcgcggcga gggcgagggc gatgccacca acggcaagct gaccctgaag ttcatctgca
     2041 ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac caccctgacc tacggcgtgc
     2101 agtgcttcag ccgctacccc gaccacatga agcagcacga cttcttcaag tccgccatgc
     2161 ccgaaggcta cgtccaggag cgcaccatct ccttcaagga cgacggcacc tacaagaccc
     2221 gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg catcgagctg aagggcatcg
     2281 acttcaagga ggacggcaac atcctggggc acaagctgga gtacaacttc aacagccaca
     2341 acgtctatat cacggccgac aagcagaaga acggcatcaa ggcgaacttc aagatccgcc
     2401 acaacgtcga ggacggcagc gtgcagctcg ccgaccacta ccagcagaac acccccatcg
     2461 gcgacggccc cgtgctgctg cccgacaacc actacctgag cacccagtcc aagctgagca
     2521 aagaccccaa cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc gccgccggga
     2581 tcactctcgg catggacgag ctgtacaagg ggatcgagga aaacctgtac ttccaatcca
     2641 atgcagtttt ggaataccaa caaagggaag gtaaaggatc ctcatctaaa agcatgccac
     2701 ctgattcctc gtcaacgact attcacacat gctctgaagc acaaactggg gaagacaaag
     2761 gcttacttga ccctcatcta tcagtattgg aattgttgag taagacaggg cattcgcctt
     2821 cgccaatggg acagaacttg gttacgtcca ttgacatatc cggcaaccac aatgtaaacg
     2881 atagtatatc aggatcgtgg caagccattc agcctttgga cttgggcgct tctttcatcc
     2941 cagaaaggtg ctcttctcaa acgactaatg gaagtattct atcatctagt gatacttctg
     3001 aagaagaaca agagctgctt caagcgcctg ctgctgatat aataaatatt attaaacaag
     3061 gacaagaagg agcaaatgtc gtttcaccgt ctcatccctt taaacagtta cagaagataa
     3121 tatcgctgcc cttgcctggt aaggaaaaaa ctccttttaa cgagcaagat gatgatggtg
     3181 atgaagatga ggcttttgag gaagatagtg tgacgataac aaaaagcttg acttcttcgg
     3241 tgaatgagga agataatgat gacgaagcta tacatacaaa gtctcggtcc tattgtcgta
     3301 acaaaaaagc cgaaaactca aaaaagaaat cccctaaatc gaataaaaaa cccaagagaa
     3361 agaagcagaa gttcttcacg agctggttca cttggggcat ttcaattact ataggaatta
     3421 gttttggatg ttgtgtgaca tactttgtca ctgcagcata cgaacaccaa acagttaagt
     3481 ctttaagttt gcggccctcc atattggcct ctttactttc cttagattcc tctagcgaca
     3541 caatcaatac gcccgccact gcatctcctt cttcaactga gcagttttta tggtttgata
     3601 agggtacgct tcagataaac ttccactccg atggattcat tatgaaaagt cttaccataa
     3661 tcaaggagac ttggggcaaa atgaatactt ttgtcttgca tgctctttct aaacctttaa
     3721 aatttttgga gaaccttaat aaaagctctg aattttccat tgatgaatct aatcggattt
     3781 tggcactggg ttatattcta ttgtgactcg agtcatgtaa ttagttatgt cacgcttaca
     3841 ttcacgccct ccccccacat ccgctctaac cgaaaaggaa ggagttagac aacctgaagt
     3901 ctaggtccct atttattttt ttatagttat gttagtatta agaacgttat ttatatttca
     3961 aatttttctt ttttttctgt acagacgcgt gtacgcatgt aacattatac tgaaaacctt
     4021 gcttgagaag gttttgggac gctcgaaggc tttaatttgc ggccaagctt gcatgcctgc
     4081 aggtcgactc tagaggatcc ccgggataac ttcgtataat gtatgctata cgaagttatg
     4141 gattttctta acttcttcgg cgacagcatc accgacttcg gtggtactgt tggaaccacc
     4201 taaatcacca gttctgatac ctgcatccaa aaccttttta actgcatctt caatggcctt
     4261 accttcttca ggcaagttca atgacaattt caacatcatt gcagcagaca agatagtggc
     4321 gatagggttg accttattct ttggcaaatc tggagcagaa ccgtggcatg gttcgtacaa
     4381 accaaatgcg gtgttcttgt ctggcaaaga ggccaaggac gcagatggca acaaacccaa
     4441 ggaacctggg ataacggagg cttcatcgga gatgatatca ccaaacatgt tgctggtgat
     4501 tataatacca tttaggtggg ttgggttctt aactaggatc atggcggcag aatcaatcaa
     4561 ttgatgttga accttcaatg taggaaattc gttcttgatg gtttcctcca cagtttttct
     4621 ccataatctt gaagaggcca aaacattagc tttatccaag gaccaaatag gcaatggtgg
     4681 ctcatgttgt agggccatga aagcggccat tcttgtgatt ctttgcactt ctggaacggt
     4741 gtattgttca ctatcccaag cgacaccatc accatcatct tcctttctct taccaaagta
     4801 aatacctccc actaattctc tgacaacaac gaagtcagta cctttagcaa attgtggctt
     4861 gattggagat aagtctaaaa gagagtcgga tgcaaagtta catggtctta agttggcgta
     4921 caattgaagt tctttacgga tttttagtaa accttgttca ggtctaacac tacctgtacc
     4981 ccatttagga ccacccacag cacctaacaa aacggcatca gccttcttgg aggcttccag
     5041 cgcctcatct ggaagtggga cacctgtagc ttcgatagca gcaccaccaa ttaaatgatt
     5101 ttcgaaatcg aacttgacat tggaacgaac atcagaaata gctttaagaa ccttaatggc
     5161 ttcggctgtg atttcttgac caacgtggtc acctggcaaa acgacgatct tcttaggggc
     5221 agacattaga atggtatatc cttgaaatat atatatatat tgctgaaatg taaaaggtaa
     5281 gaaaagttag aaagtaagac gattgctaac cacctattgg aaaaacaata ggtccttaaa
     5341 taatattgtc aacttcaagt attgtgatgc aagcatttag tcatgaacgc ttctctattc
     5401 tatatgaaaa gccggttccg gcgctctcac ctttcctttt tctcccaatt tttcagttga
     5461 aaaaggtata tgcgtcaggc gacctctgaa attaacaaaa aatttccagt catcgaattt
     5521 gattctgtgc gatagcgccc ctgtgtgttc tcgttatgtt gaggaaaaaa ataatggtta
     5581 ataacttcgt ataatgtatg ctatacgaag ttatcccggg taccgagctc gaattcgtaa
     5641 cttacacgcg cctcgtatct tttaatgatg gaataatttg ggaatttact ctgtgtttat
     5701 ttatttttat gttttgtatt tggattttag aaagtaaata aagaaggtag aagagttacg
     5761 gaatgaagaa aaaaaaataa acaaaggttt aaaaaatttc aacaaaaagc gtactttaca
     5821 tatatattta ttagacaaga aaagcagatt aaatagatat acattcgatt aacgataagt
     5881 aaaatgtaaa atcacaggat tttcgtgtgt ggtcttctac acagacaaga tgaaacaatt
     5941 cggcattaat acctgagagc aggaagagca agataaaagg tagtatttgt tggcgatccc
     6001 cctagagtct tttacatctt cggaaaacaa aaactatttt ttctttaatt tcttttttta
     6061 ctttctattt ttaatttata tatttatatt aaaaaattta aattataatt atttttatag
     6121 cacgtgatga attcgtaatc atgtcatagc tgtttcctgt gtgaaattgt tatccgctca
     6181 caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag
     6241 tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt
     6301 cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc
     6361 gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg
     6421 tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa
     6481 agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg
     6541 cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga
     6601 ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg
     6661 tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg
     6721 gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc
     6781 gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg
     6841 gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca
     6901 ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt
     6961 ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag
     7021 ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg
     7081 gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caa
//
