LOCUS       sequence-382500-        7690 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Marker for yeast early Golgi. Integration of 2xGFP tag at SYS1 C 
            terminus. Uses auxotrophic marker URA3(Candida albicans). .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7690)
  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 7690)
  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: SYS1-2EGFP-ter-URA3
FEATURES             Location/Qualifiers
     source          1..7690
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1042..1755
                     /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
                     FVTAAGITHGIDELYK"
     CDS             1792..2508
                     /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"
     CDS             1792..2505
                     /codon_start=1
                     /product="green fluorescent protein"
                     /label=GFP
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     terminator      2529..2716
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(2584..2608)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     protein_bind    2999..3032
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     terminator      3987..4184
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     protein_bind    4238..4271
                     /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(5334..5353)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(5335..5353)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5440..5457)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5611..6199)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5691..5710)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6370..7161,7162..7230))
                     /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     6993..7012
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7231..7335)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7403..7421
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7459..7481)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7581..7600
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        join(7681..7690,1..9)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     join(7681..7690,1..8)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
ORIGIN
        1 cactatagaa cgcggccgcc agctgaagct tccagatata aggaagtgta tttgctttct
       61 aataacttgt gtacggccat aacccggctt ttaatcagtt tatttaacaa ggtatacaaa
      121 tcgccaaacg taacaagaga taggcgctat tattcattta atattgtcgt attcaaaata
      181 ctaacggtga acaatttgct ccattgtaac actataggac aaggctaatc atgaataaat
      241 atctccccac ccaaaatatt cgtagctcgc gatattcacg gtaatattct gtaacttcat
      301 gttaagagat gatctttagg aaggctaagg aagggtacac atagaagtga gataaaaaat
      361 aggcatttga attactgtaa catggtttcg ataagaaggt atttgcgagt gccgaatgag
      421 ttgaaacctt cccagatctt caaacaagat tctctctctc caagtaagat tggacttcaa
      481 attgttcttt tacagatttt ttattacacc acagcgatcg ttctttttta ttgttgggcg
      541 aagctagcgg gatatgatct caatatcaag gaatggctgt tttcttggga gaatatcgac
      601 tttacgaacg catacggttt atcaatctca ctgttatggc ttctggattc attaatatgt
      661 gtttttttct taactgttat cgttggacgg agtaaactcg catgggattt tgctatcact
      721 atacatgcca tcaattttat tgtggtgttc ctctatactc ggaaatttcc ttctttttct
      781 tggttttttc ttcaaatttt atcatcttta atactaatat ttttggggac atggacaaca
      841 agatggagag agcttagaga cacctttttt gagggcttgg ttgatcctaa tgaaggagaa
      901 gttggattgg ttacgcctag tcagcagcat agtaatcatt cagaattgga gcaatcacca
      961 atacaactaa aagacttaga aagccaaata gcttcgtacg ctgcaggtcg acggatcggt
     1021 gacggtgctg gtttaattaa catgtctaaa ggtgaagaat tattcactgg tgttgtccca
     1081 attttggttg aattagatgg tgatgttaat ggtcacaaat tttctgtctc cggtgaaggt
     1141 gaaggtgatg ctacttacgg taaattgacc ttaaaattta tttgtactac tggtaaattg
     1201 ccagttccat ggccaacctt agtcactact ttaacttatg gtgttcaatg tttttctaga
     1261 tacccagatc atatgaaaca acatgacttt ttcaagtctg ccatgccaga aggttatgtt
     1321 caagaaagaa ctattttttt caaagatgac ggtaactaca agaccagagc tgaagtcaag
     1381 tttgaaggtg ataccttagt taatagaatc gaattaaaag gtattgattt taaagaagat
     1441 ggtaacattt taggtcacaa attggaatac aactataact ctcacaatgt ttacatcatg
     1501 gctgacaaac aaaagaatgg tatcaaagtt aacttcaaaa ttagacacaa cattgaagat
     1561 ggttctgttc aattagctga ccattatcaa caaaatactc caattggtga tggtccagtc
     1621 ttgttaccag acaaccatta cttatccact caatctgcct tatccaaaga tccaaacgaa
     1681 aagagagacc acatggtctt gttagaattt gttactgctg ctggtattac ccatggtatt
     1741 gatgaattgt acaaaattcc agttccaacg cgtgatctag agaagccatt catgtctaaa
     1801 ggtgaagaat tattcactgg tgttgtccca attttggttg aattagatgg tgatgttaat
     1861 ggtcacaaat tttctgtctc cggtgaaggt gaaggtgatg ctacttacgg taaattgacc
     1921 ttaaaattta tttgtactac tggtaaattg ccagttccat ggccaacctt agtcactact
     1981 ttaacttatg gtgttcaatg tttttctaga tacccagatc atatgaaaca acatgacttt
     2041 ttcaagtctg ccatgccaga aggttatgtt caagaaagaa ctattttttt caaagatgac
     2101 ggtaactaca agaccagagc tgaagtcaag tttgaaggtg ataccttagt taatagaatc
     2161 gaattaaaag gtattgattt taaagaagat ggtaacattt taggtcacaa attggaatac
     2221 aactataact ctcacaatgt ttacatcatg gctgacaaac aaaagaatgg tatcaaagtt
     2281 aacttcaaaa ttagacacaa cattgaagat ggttctgttc aattagctga ccattatcaa
     2341 caaaatactc caattggtga tggtccagtc ttgttaccag acaaccatta cttatccact
     2401 caatctgcct tatccaaaga tccaaacgaa aagagagacc acatggtctt gttagaattt
     2461 gttactgctg ctggtattac ccatggtatg gatgaattat ataaataagg cgcgccactt
     2521 ctaaataagc gaatttctta tgatttatga tttttattat taaataagtt ataaaaaaaa
     2581 taagtgtata caaattttaa agtgactctt aggttttaaa acgaaaattc ttattcttga
     2641 gtaactcttt cctgtaggtc aggttgcttt ctcaggtata gtatgaggtc gctcttattg
     2701 accacacctc taccggcaga tccgctaggg ataacagggt aatatagatc tgtttagctt
     2761 gcctcgtccc cgccgggtca cccggccagc gacatggagg cccagaatac cctccttgac
     2821 agtcttgacg tgcgcagctc aggggcatga tgtgactgtc gcccgtacat ttagcccata
     2881 catccccatg tataatcatt tgcatccata cattttgatg gccgcacggc gcgaagcaaa
     2941 aattacggct cctcgctgcg gacctgcgag cagggaaacg ctcccctcac agacgcgtat
     3001 aacttcgtat aatgtatgct atacgaagtt attgaattgt ccccacgccg cgcccctgta
     3061 gagaaatata aaaggttagg atttgccact gaggttcttc tttcatatac ttccttttaa
     3121 aatcttgcta ggatacagtt ctcacatcac atccgaacat aaacaaccat gacagtcaac
     3181 actaagacct atagtgagag agcagaaact catgcctcac cagtagcaca acgattattt
     3241 cgattaatgg aactgaagaa aaccaattta tgtgcatcaa ttgatgttga taccactaag
     3301 gaattccttg aattaattga taaattgggt ccttatgtat gcttaatcaa gactcatatt
     3361 gatataatca atgatttttc ctatgaatcc actattgaac cattattaga actttcacgt
     3421 aaacatcaat ttatgatttt tgaagataga aaatttgctg atattggtaa taccgtgaag
     3481 aaacaatata ttggtggagt ttataaaatt agtagttggg cagatattac taatgctcat
     3541 ggtgtcactg ggaatggagt agttgaagga ttaaaacagg gagctaaaga aaccaccacc
     3601 aaccaagagc caagagggtt attgatgtta gctgaattat catcagtggg atcattagca
     3661 tatggagaat attctcaaaa aactgttgaa attgctaaat ccgataagga atttgttatt
     3721 ggatttattg cccaacgtga tatgggtgga caagaagaag gatttgattg gcttattatg
     3781 acacctggag ttggattaga tgataaaggt gatggattag gacaacaata tagaactgtt
     3841 gatgaagttg ttagcactgg aactgatatt atcattgttg gtagaggatt gtttggtaaa
     3901 ggaagagatc cagatattga aggtaaaagg tatagagatg ctggttggaa tgcttatttg
     3961 aaaaagactg gccaattata atcagtactg acaataaaaa gattcttgtt ttcaagaact
     4021 tgtcatttgt atagtttttt tatattgtag ttgttctatt ttaatcaaat gttagcgtga
     4081 tttatatttt ttttcgcctc gacatcatct gcccagatgc gaagttaagt gcgcagaaag
     4141 taatatcatg cgtcaatcgt atgtgaatgc tggtcgctat actgctgtcg attcgatact
     4201 aacgccgcca tccagtgtcg aaaacgagct cgaattcata acttcgtata atgtatgcta
     4261 tacgaagtta tatcgatgat atcagatcca ctagtggcct atgcggccgc ggtgagagga
     4321 gatgaaacaa catattcgag gcaattttgg caatattata gtcattcaag agaaaacagt
     4381 aaatgtatga aaaatggcaa gaaccttatc tctataggct taataaatct gcctttttgg
     4441 ccaatttttc aaaacagatc ctatttctta tttatacaca atctatattt aacataatta
     4501 agtacgtgat gacattgaag ataataataa gtaaagagtt tacgtactta aattttatat
     4561 agttttagta tacttgcaat gtgtgaatgg atctagtaag ctgaggtact tctctagcgt
     4621 gaatatgaaa cgagtaacag ggtggtacat aatttacgaa cagaactcaa gttgataaat
     4681 cctttgcgtt cttaaccatt tgtaaaagtt cgtcaacaac attgaaaata ttgcttcgag
     4741 cagatgtttt atcgtttctg caaatttcca gtcctgaaca cctctgttgt cgtaaatgta
     4801 ataatgccac actggattcg attctagtaa ttacatgatc aaaaaataac aacgcatcct
     4861 tctcagaaat cgattgcata ataccatttt caactttagg gttaaattta atataacttc
     4921 cctccgtccg agagtctagt tcaataaatc cgccgcttaa agctgacatt atagattcca
     4981 gtctgtttga aatgactctt aaacgaaaca acatcgtttc attgattaat tttgactgct
     5041 catcttcaga tgcaaattcg tgtctggaag caagaatctt aggatacgca atagtaacga
     5101 gttctttggt ttcgagtccc tttagcttat gctcgccaaa gtcaaaaaat gcccaaccaa
     5161 tactttccag catggcaatt tctctttcaa gaacctctcc gataatttct tcaccataaa
     5221 cttccttgag agattccttg cccttcacta ctcgctcatg atacttcatt atcttgttga
     5281 attcagagta aaaatcacta ctcattgcaa tctgaccgcg gatctgccgg tctccctata
     5341 gtgagtcgta ttaatttcga taagccaggt taacctgcat taatgaatcg gccaacgcgc
     5401 ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg
     5461 ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc
     5521 cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag
     5581 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca
     5641 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca
     5701 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg
     5761 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag
     5821 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt
     5881 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca
     5941 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg
     6001 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt
     6061 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc
     6121 cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg
     6181 cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg
     6241 gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta
     6301 gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg
     6361 gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg
     6421 ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc
     6481 atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc
     6541 agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc
     6601 ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag
     6661 tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat
     6721 ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg
     6781 caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt
     6841 gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag
     6901 atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg
     6961 accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt
     7021 aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct
     7081 gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac
     7141 tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat
     7201 aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat
     7261 ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca
     7321 aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtctaag aaaccattat
     7381 tatcatgaca ttaacctata aaaataggcg tatcacgagg ccctttcgtc tcgcgcgttt
     7441 cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca cagcttgtct
     7501 gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg ttggcgggtg
     7561 tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc accatatgga
     7621 catattgtcg ttagaacgcg gctacaatta atacataacc ttatgtatca tacacatacg
     7681 atttaggtga
//
