LOCUS       sequence-382504-        5391 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Marker for DAG. Yeast expression of human PKC beta under LSY2 
            promoter. Uses auxotrophic marker TRP1(Kluyveromyces lactis). .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5391)
  AUTHORS   Li D, Yang SG, He CW, Zhang ZT, Liang Y, Li H, Zhu J, Su X, Gong Q, 
            Xie Z
  TITLE     Excess diacylglycerol at the endoplasmic reticulum disrupts 
            endomembrane homeostasis and autophagy.
  JOURNAL   BMC Biol. 2020 Aug 28;18(1):107. doi: 10.1186/s12915-020-00837-w.
  PUBMED    32859196
REFERENCE   2  (bases 1 to 5391)
  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: "2020-08-28"; volume: "18(1):107. doi"; pages: " 
            10.1186/s12915-020-00837-w"
COMMENT     Sequence Label: UG76-pLYS2-GFP-PKCbeta
FEATURES             Location/Qualifiers
     source          1..5391
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        271..510
                     /gene="S. cerevisiae LYS2"
                     /label=LYS2 promoter
     CDS             910..1623
                     /codon_start=1
                     /product="green fluorescent protein"
                     /label=GFP
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     CDS             910..1623
                     /codon_start=1
                     /product="yeast-enhanced green fluorescent protein"
                     /label=yeGFP
                     /note="codons optimized for translation in S. cerevisiae 
                     and C. albicans"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     terminator      1653..1840
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(1708..1732)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     protein_bind    1892..1925
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    2928..2961
                     /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(3014..3033)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3015..3033)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3120..3137)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3291..3879)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3371..3390)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4050..4841,4842..4910))
                     /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     4673..4692
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4911..5015)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5083..5101
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5139..5161)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     5261..5280
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        5361..5379
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     5361..5378
                     /label=SP6
                     /note="SP6 promoter, forward primer"
ORIGIN
        1 gctgaagctt agcagttgct ttctcctatg ggaagagctt tctaagtctg aagaagtaaa
       61 cagttctttg ctatttcaca cttcctggtt gatggtcact tgctgcctga aatatatata
      121 tatgtatgac atatgtactt gttttctttt ttgtgccttt gttacgtcta tattcattga
      181 aactgattat tcgattttct tcttgctgac cgcttctaga ggcatcgcac agttttagcg
      241 aggaaaactc ttcaatagtt ttgccagcgg aattccactt gcaattacat aaaaaattcc
      301 ggcggttttt cgcgtgtgac tcaatgtcga aatacctgcc taatgaacat gaacatcgcc
      361 caaatgtatt tgaagacccg ctgggagaag ttcaagatat ataagtaaca agcagccaat
      421 agtataaaaa aaaatctgag tttattacct ttcctggaat ttcagtgaaa aactgctaat
      481 tatagagaga tatcacagag ttactcacta gctagcatgg tgcatgaggt caagaaccac
      541 aaattcaccg cccgcttctt caagcagccc accttctgca gccactgcac cgacttcatc
      601 tggggcttcg ggaagcaggg attccagtgc caagtttgct gctttgtggt gcacaagcgg
      661 tgccatgaat ttgtcacatt ctcctgccct ggcgctgaca agggtccagc ctccgatgac
      721 ccccgcagca aacacaagtt taagatccac acgtactcca gccccacgtt ttgtgaccac
      781 tgtgggtcac tgctgtatgg actcatccac caggggatga aatgtgacac ctgcatgatg
      841 aatgtgcaca agcgctgcgt gatgaatgtt cccagcctgt gtggcacgga ccacacggag
      901 cgcggatcca tgtctaaagg tgaagaatta ttcactggtg ttgtcccaat tttggttgaa
      961 ttagatggtg atgttaatgg tcacaaattt tctgtctccg gtgaaggtga aggtgatgct
     1021 acttacggta aattgacctt aaaatttatt tgtactactg gtaaattgcc agttccatgg
     1081 ccaaccttag tcactacttt aacttatggt gttcaatgtt tttctagata cccagatcat
     1141 atgaaacaac atgacttttt caagtctgcc atgccagaag gttatgttca agaaagaact
     1201 atttttttca aagatgacgg taactacaag accagagctg aagtcaagtt tgaaggtgat
     1261 accttagtta atagaatcga attaaaaggt attgatttta aagaagatgg taacatttta
     1321 ggtcacaaat tggaatacaa ctataactct cacaatgttt acatcatggc tgacaaacaa
     1381 aagaatggta tcaaagttaa cttcaaaatt agacacaaca ttgaagatgg ttctgttcaa
     1441 ttagctgacc attatcaaca aaatactcca attggtgatg gtccagtctt gttaccagac
     1501 aaccattact tatccactca atctgcctta tccaaagatc caaacgaaaa gagagaccac
     1561 atggtcttgt tagaatttgt tactgctgct ggtattaccc atggtatgga tgaattgtac
     1621 aaacctaggt gaggcgcgcc acttctaaat aagcgaattt cttatgattt atgattttta
     1681 ttattaaata agttataaaa aaaataagtg tatacaaatt ttaaagtgac tcttaggttt
     1741 taaaacgaaa attcttattc ttgagtaact ctttcctgta ggtcaggttg ctttctcagg
     1801 tatagtatga ggtcgctctt attgaccaca cctctaccgg cagatccgct agggataaca
     1861 gggtaatatc tgcaggtcga caacccttaa tataacttcg tataatgtat gctatacgaa
     1921 gttattaggt ctagagcggc cgcggaacga tcattcacta tatatatatc aatttatata
     1981 tacgtatgtg taattgaaga aagatacgtt ttttcctcta ttgagaggcc tgctggatga
     2041 atagctttac cttttctaaa tccttgatac catcagtctc tactcctcca cttacatcga
     2101 caccgatcgc atttggtaac atattaatgg caacagaaac gttatcagga ttcaatccac
     2161 cagcgataat gaattttatc tcgggatgac ttgcagacca actggaaatt gcactccaat
     2221 tcaatttctc accagtgcca ccttcaccag aatcgaacaa cgtcagcaca ttgtctacgt
     2281 gttcatacag gtccagtagt aattcacaat cctgtgggaa ctggaacctc ttaatgattg
     2341 gaattgaaga tgggatcaaa gatctgtatt ctttaatatc ttcatctcca tgtaattgta
     2401 tcacatctag attatattcg tggtacagtt gaaggacatc atcaacggac tgatttctaa
     2461 acaccccgac tagtttagta cctttcacgt tctcttgttg gtgaactgca gttgaaatac
     2521 ctttcgcaac agatgagtca atggttcttt tcctaccggg aacacaaatg atacctaagt
     2581 aatcagcacc atcatccaca gcagtctttg cagcttcaac ggtttgcaaa ccacacactt
     2641 taacgagcat cacagtagat ttcgcaagag ggtagctcgc tcagagtacc caagtaaatg
     2701 attagtaaac tgatgtttga tagttcaatt tttcaatgaa ataaccttat attaaaattg
     2761 atattactat tatacaaaaa taaagaataa aggatttgag tttatacata aaataccatt
     2821 attatttgtt cagtgagaga tactggggta tatgggatgt gtgtagtgat accatgcaat
     2881 catgtatcaa acatgggccc ggtaccgagc tctcgagaac ccttaatata acttcgtata
     2941 atgtatgcta tacgaagtta ttaggtgata tcagatccac tagtggccta tgcggccgcg
     3001 gatctgccgg tctccctata gtgagtcgta ttaatttcga taagccaggt taacctgcat
     3061 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc
     3121 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
     3181 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     3241 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     3301 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     3361 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     3421 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     3481 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     3541 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     3601 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     3661 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     3721 tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     3781 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     3841 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     3901 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     3961 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     4021 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     4081 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     4141 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     4201 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     4261 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     4321 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     4381 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     4441 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     4501 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     4561 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     4621 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     4681 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     4741 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     4801 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     4861 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     4921 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     4981 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     5041 gacgtctaag aaaccattat tatcatgaca ttaacctata aaaataggcg tatcacgagg
     5101 ccctttcgtc tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg
     5161 gagacggtca cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg
     5221 tcagcgggtg ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta
     5281 ctgagagtgc accatatgga catattgtcg ttagaacgcg gctacaatta atacataacc
     5341 ttatgtatca tacacatacg atttaggtga cactatagaa cgcggccgcc a
//
