LOCUS       sequence-422260-        4843 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid backbone for yeast expression, use auxotrophic marker MET15 
            (Saccharomyces cerevisiae).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4843)
  AUTHORS   He CW, Cui XF, Ma SJ, Xu Q, Ran YP, Chen WZ, Mu JX, Li H, Zhu J, 
            Gong Q, Xie Z
  TITLE     Membrane recruitment of Atg8 by Hfl1 facilitates turnover of 
            vacuolar membrane proteins in yeast cells approaching stationary 
            phase.
  JOURNAL   BMC Biol. 2021 Jun 4;19(1):117. doi: 10.1186/s12915-021-01048-7.
  PUBMED    34088313
REFERENCE   2  (bases 1 to 4843)
  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: "BMC Biol.";
            date: "2021-06-4"; volume: "19(1):117. doi"; pages: " 
            10.1186/s12915-021-01048-7"
COMMENT     Sequence Label: ClhN-MET15
FEATURES             Location/Qualifiers
     source          1..4843
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      101..288
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(156..180)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     protein_bind    340..373
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(556..1890)
                     /codon_start=1
                     /gene="S. cerevisiae MET17"
                     /gene_synonym="MET15; MET25"
                     /product="O-acetylserine and O-acetylhomoserine 
                     sulfhydrylase, required for sulfur amino acid synthesis"
                     /label=MET17
                     /translation="MPSHFDTVQLHAGQENPGDNAHRSRAVPIYATTSYVFENSKHGSQ
                     LFGLEVPGYVYSRFQNPTSNVLEERIAALEGGAAALAVSSGQAAQTLAIQGLAHTGDNI
                     VSTSYLYGGTYNQFKISFKRFGIEARFVEGDNPEEFEKVFDERTKAVYLETIGNPKYNV
                     PDFEKIVAIAHKHGIPVVVDNTFGAGGYFCQPIKYGADIVTHSATKWIGGHGTTIGGII
                     VDSGKFPWKDYPEKFPQFSQPAEGYHGTIYNEAYGNLAYIVHVRTELLRDLGPLMNPFA
                     SFLLLQGVETLSLRAERHGENALKLAKWLEQSPYVSWVSYPGLASHSHHENAKKYLSNG
                     FGGVLSFGVKDLPNADKETDPFKLSGAQVVDNLKLASNLANVGDAKTLVIAPYFTTHKQ
                     LNDKEKLASGVTKDLIRVSVGIEFIDDIIADFQQSFETVFAGQKP"
     promoter        complement(1891..2241)
                     /gene="S. cerevisiae MET17"
                     /gene_synonym="MET15; MET25"
                     /label=MET17 promoter
                     /note="expression is repressed by methionine"
     protein_bind    2394..2427
                     /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(2480..2499)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2481..2499)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2586..2603)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2757..3345)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2837..2856)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3516..4307,4308..4376))
                     /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     4139..4158
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4377..4481)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4549..4567
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(4605..4627)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4727..4746
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        join(4827..4843,1..2)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     join(4827..4843,1)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
ORIGIN
        1 gaacgcggcc gccagctgaa gcttcgtacg ctgcaggtcg tgggattcct gggttaatta
       61 acatcgctag ccctaggtga ggcgcgccac ttctaaataa gcgaatttct tatgatttat
      121 gatttttatt attaaataag ttataaaaaa aataagtgta tacaaatttt aaagtgactc
      181 ttaggtttta aaacgaaaat tcttattctt gagtaactct ttcctgtagg tcaggttgct
      241 ttctcaggta tagtatgagg tcgctcttat tgaccacacc tctaccggca gatccgctag
      301 ggataacagg gtaatatctg caggtcgaca acccttaata taacttcgta taatgtatgc
      361 tatacgaagt tattaggtct agatgcgaat taacactgtt ctttgatgtt agaacaattt
      421 aggttcaaag tacgagtcac gacatgtaac aagggaaaaa aaggatattc atttcaataa
      481 agttcgtttt ataaaagtat agtacttgtg agagaaagta ggtttataca taattttaca
      541 actcattacg cacactcatg gtttttggcc agcgaaaaca gtttcaaaag attgctggaa
      601 gtctgcaata atgtcatcaa taaattcgat accaacagag acacgaatta agtccttggt
      661 aacaccagat gccaactttt ctttgtcatt taattgtttg tgggtagtga agtatggagc
      721 aatgactaag gtcttggcat caccaacatt ggccaagtta gaggcaagct ttaaattgtc
      781 aacaacttga gcaccagaaa gtttgaatgg gtcagtttcc ttgtcggcat ttggtaagtc
      841 ttttacaccg aaagataaga caccaccgaa accgttagat agatacttct tagcattttc
      901 atgatgagaa tgagatgcta aaccagggta tgaaacccaa gatacgtatg gggattgttc
      961 taaccatttg gctaacttca atgcattttc accgtgtctt tcagctctca aagataatgt
     1021 ttcaacacct tgtagtagca agaaagaggc aaatgggttc atcaatggac ccaaatctct
     1081 taatagttca gttctaacat gaacgatgta tgccaagtta ccgtaggctt cattgtagat
     1141 agtaccgtga tatccttcgg caggttgaga gaattgaggg aacttttctg ggtagtcctt
     1201 ccatgggaac ttaccagagt caacaataat accaccgata gtagtaccat gaccaccaat
     1261 ccatttggta gcagaatgtg ttacaatatc agcaccgtat ttaattggct gacagaagta
     1321 accaccggca ccaaatgtgt tgtcaacgac aactggaata ccgtgtttgt gagcaattgc
     1381 aacaattttt tcaaaatccg gaacattgta ctttggatta ccaatggttt ccaaataaac
     1441 agccttggtt ctttcatcaa agaccttttc gaattcttct ggattgtcac cttcaacaaa
     1501 tctagcctcg ataccaaatc ttttgaacga gattttgaac tggttataag taccaccgta
     1561 taagtaagaa gtggaaacga tgttgtcacc agtgtgtgcc aaaccttgga tggcaagggt
     1621 ttgagcggct tgaccggagg aaacagccaa agcagcagca ccaccttcta aagcagcaat
     1681 tctttcttcc aaaacattac tggttgggtt ttggaaacgg gaatagacgt aacctggaac
     1741 ttctagacca aacaattgcg aaccatgctt agagttttcg aaaacataag aagtggtggc
     1801 gtaaattggt acagctctgg atctgtgagc attgtcacca gggttctctt ggccggcgtg
     1861 tagttgaaca gtatcgaaat gagatggcat tgtatggatg ggggtaatag aattgtatct
     1921 atgtatctga cgaccctgta ttacacgaag gaaatagtag atgaaaagac aagagagcaa
     1981 gaaaaaggaa aaacttcctt tctaacagac gctttactta accttatata tatcttattt
     2041 ttttcatcga gcgtgttcaa ttggacaagg tgccaccttt tcgacacttc ggttattatg
     2101 ttacacagtt ttcatgagga tggcgccttg actaacttaa ttagtcattt gccaaccacc
     2161 acagttcccc aatatcgaca gcttcacgtg ccatttgcca ttttgcgatc atgtgacctc
     2221 aagagaaaaa gcaaaaaata atgagagcta agggattcga acccttgcat ccgaagatat
     2281 cagagatttt agaggttaaa tccacctaaa tctgacgcct taaaccactc ggccaaactc
     2341 ccattgtatg ttggaagaat tggatttaaa tggagctctc gagaaccctt aatataactt
     2401 cgtataatgt atgctatacg aagttattag gtgatatcag atccactagt ggcctatgcg
     2461 gccgcggatc tgccggtctc cctatagtga gtcgtattaa tttcgataag ccaggttaac
     2521 ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc
     2581 gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct
     2641 cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg
     2701 tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc
     2761 cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga
     2821 aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct
     2881 cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg
     2941 gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag
     3001 ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat
     3061 cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac
     3121 aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac
     3181 tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc
     3241 ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt
     3301 tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc
     3361 ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg
     3421 agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca
     3481 atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca
     3541 cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag
     3601 ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac
     3661 ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc
     3721 agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct
     3781 agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc
     3841 gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg
     3901 cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc
     3961 gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat
     4021 tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag
     4081 tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat
     4141 aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg
     4201 cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca
     4261 cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga
     4321 aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc
     4381 ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag cggatacata
     4441 tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg
     4501 ccacctgacg tctaagaaac cattattatc atgacattaa cctataaaaa taggcgtatc
     4561 acgaggccct ttcgtctcgc gcgtttcggt gatgacggtg aaaacctctg acacatgcag
     4621 ctcccggaga cggtcacagc ttgtctgtaa gcggatgccg ggagcagaca agcccgtcag
     4681 ggcgcgtcag cgggtgttgg cgggtgtcgg ggctggctta actatgcggc atcagagcag
     4741 attgtactga gagtgcacca tatggacata ttgtcgttag aacgcggcta caattaatac
     4801 ataaccttat gtatcataca catacgattt aggtgacact ata
//
