LOCUS       sequence-256852-        7735 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Integration of 2xGFP tag at SEC7 C terminus, use auxotrophic marker 
            URA3(Candida albicans). Marker for yeast late golgi/early endosome..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7735)
  AUTHORS   Zhu J, Zhang ZT, Tang SW, Zhao BS, Li H, Song JZ, Li D, Xie Z
  TITLE     A Validated Set of Fluorescent-Protein-Based Markers for Major 
            Organelles in Yeast (Saccharomyces cerevisiae).
  JOURNAL   MBio. 2019 Sep 3;10(5). pii: mBio.01691-19. doi: 
            10.1128/mBio.01691-19.
  PUBMED    31481383
REFERENCE   2  (bases 1 to 7735)
  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: "MBio."; 
            date: "2019-09-3"; volume: "10(5). pii: mBio.01691-19. doi"; pages: 
            " 10.1128/mBio.01691-19"
COMMENT     Sequence Label: ZJOM156
FEATURES             Location/Qualifiers
     source          1..7735
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1110..1823
                     /codon_start=1
                     /product="green fluorescent protein"
                     /label=GFP
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     CDS             1110..1823
                     /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             1860..2576
                     /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             1860..2573
                     /codon_start=1
                     /product="green fluorescent protein"
                     /label=GFP
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     terminator      2597..2784
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(2652..2676)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     protein_bind    3067..3100
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     terminator      4055..4252
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     protein_bind    4306..4339
                     /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(5379..5398)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(5380..5398)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5485..5502)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5656..6244)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5736..5755)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6415..7206,7207..7275))
                     /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     7038..7057
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7276..7380)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7448..7466
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7504..7526)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7626..7645
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        join(7726..7735,1..9)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     join(7726..7735,1..8)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
ORIGIN
        1 cactatagaa cgcggccgcc agctgaagct tgatattgca aaacgtatct aaattcaatg
       61 agtaccattg gaatcaaata ggtgacgtat tcgataaatt gtttgattta accactgcta
      121 atgaattgtt cgattatgat ccgttacagc aaggaagaaa atcatctgta tctcatcacc
      181 aaacaactaa tgataccagc caacactcag atgatgatag caacgacaga agggagaacg
      241 attctaatat cagtgagacg gtcgaaagag cgcatcaaga agaatccagc gaagacgttg
      301 gtggtgacat ggttgaaaca ctcaatgggc aaactaagct taataatggc aattccgttc
      361 cgacggtaaa ggacgaattg aatccaaagc ccgctagtct aagtatcccc aagaaaacta
      421 agcacatgaa acgaaacgaa agtaacgaag atatacgtag gagaataaac atcaagaatt
      481 ctattgttgt caaatgtgtt ttacagcttt tgatgataga gctgctcaac gaattattcg
      541 agaacgaaga ttttgcccac tgtatccctt acaaggaagc cattagaatt acaagattgt
      601 tggagaaatc gtacgaattt tctcgtgact ttaatgaaga ttatgggtta agaacaagac
      661 tagtagaggc tcgtgtagtt gataaaatac caaacctgtt gaaacaagaa acgagtgctg
      721 ctgcagttct ccttgacatt atgtttcagt tgtatctgaa cgatgatgag aagaaggctg
      781 atttaataac tcgtctgatc accatttgca tccaggtcgt agagggttat gtttctttgg
      841 atgatagaac catggaacgt agtatcaatg cctggcgttc agtgatagtt gaaatattac
      901 aaggttacta cgaatttgat gatgaagact ttagactata ctgtcctgct atgtatgctc
      961 tggtgattca aatattagat aagagcgttc caacggagtt gagacatgct ataaaacaat
     1021 ttctaagcag agttggtgaa ttataccttt ctactgatgc ttcgtacgct gcaggtcgac
     1081 ggatcggtga cggtgctggt ttaattaaca tgtctaaagg tgaagaatta ttcactggtg
     1141 ttgtcccaat tttggttgaa ttagatggtg atgttaatgg tcacaaattt tctgtctccg
     1201 gtgaaggtga aggtgatgct acttacggta aattgacctt aaaatttatt tgtactactg
     1261 gtaaattgcc agttccatgg ccaaccttag tcactacttt aacttatggt gttcaatgtt
     1321 tttctagata cccagatcat atgaaacaac atgacttttt caagtctgcc atgccagaag
     1381 gttatgttca agaaagaact atttttttca aagatgacgg taactacaag accagagctg
     1441 aagtcaagtt tgaaggtgat accttagtta atagaatcga attaaaaggt attgatttta
     1501 aagaagatgg taacatttta ggtcacaaat tggaatacaa ctataactct cacaatgttt
     1561 acatcatggc tgacaaacaa aagaatggta tcaaagttaa cttcaaaatt agacacaaca
     1621 ttgaagatgg ttctgttcaa ttagctgacc attatcaaca aaatactcca attggtgatg
     1681 gtccagtctt gttaccagac aaccattact tatccactca atctgcctta tccaaagatc
     1741 caaacgaaaa gagagaccac atggtcttgt tagaatttgt tactgctgct ggtattaccc
     1801 atggtatgga tgaattgtac aaaattccag ttccaacgcg tgatctagag aagccattca
     1861 tgtctaaagg tgaagaatta ttcactggtg ttgtcccaat tttggttgaa ttagatggtg
     1921 atgttaatgg tcacaaattt tctgtctccg gtgaaggtga aggtgatgct acttacggta
     1981 aattgacctt aaaatttatt tgtactactg gtaaattgcc agttccatgg ccaaccttag
     2041 tcactacttt aacttatggt gttcaatgtt tttctagata cccagatcat atgaaacaac
     2101 atgacttttt caagtctgcc atgccagaag gttatgttca agaaagaact atttttttca
     2161 aagatgacgg taactacaag accagagctg aagtcaagtt tgaaggtgat accttagtta
     2221 atagaatcga attaaaaggt attgatttta aagaagatgg taacatttta ggtcacaaat
     2281 tggaatacaa ctataactct cacaatgttt acatcatggc tgacaaacaa aagaatggta
     2341 tcaaagttaa cttcaaaatt agacacaaca ttgaagatgg ttctgttcaa ttagctgacc
     2401 attatcaaca aaatactcca attggtgatg gtccagtctt gttaccagac aaccattact
     2461 tatccactca atctgcctta tccaaagatc caaacgaaaa gagagaccac atggtcttgt
     2521 tagaatttgt tactgctgct ggtattaccc atggtatgga tgaattatat aaataaggcg
     2581 cgccacttct aaataagcga atttcttatg atttatgatt tttattatta aataagttat
     2641 aaaaaaaata agtgtataca aattttaaag tgactcttag gttttaaaac gaaaattctt
     2701 attcttgagt aactctttcc tgtaggtcag gttgctttct caggtatagt atgaggtcgc
     2761 tcttattgac cacacctcta ccggcagatc cgctagggat aacagggtaa tatagatctg
     2821 tttagcttgc ctcgtccccg ccgggtcacc cggccagcga catggaggcc cagaataccc
     2881 tccttgacag tcttgacgtg cgcagctcag gggcatgatg tgactgtcgc ccgtacattt
     2941 agcccataca tccccatgta taatcatttg catccataca ttttgatggc cgcacggcgc
     3001 gaagcaaaaa ttacggctcc tcgctgcaga cctgcgagca gggaaacgct cccctcacag
     3061 acgcgtataa cttcgtataa tgtatgctat acgaagttat tgaattgtcc ccacgccgcg
     3121 cccctgtaga gaaatataaa aggttaggat ttgccactga ggttcttctt tcatatactt
     3181 ccttttaaaa tcttgctagg atacagttct cacatcacat ccgaacataa acaaccatga
     3241 cagtcaacac taagacctat agtgagagag cagaaactca tgcctcacca gtagcacagc
     3301 gattatttcg attaatggaa ctgaagaaaa ccaatttatg tgcatcaatt gacgttgata
     3361 ccactaagga attccttgaa ttaattgata aattaggtcc ttatgtatgc ttaatcaaga
     3421 ctcatattga tataatcaat gatttttcct atgaatccac tattgaacca ttattagaac
     3481 tttcacgtaa acatcaattt atgatttttg aagatagaaa atttgctgat attggtaata
     3541 ccgtaaagaa acaatatatt ggtggagttt ataaaattag tagttgggca gatattacca
     3601 atgctcatgg tgtcactggg aatggagtgg ttgaaggatt aaaacaggga gctaaagaaa
     3661 ccaccaccaa ccaagagcca agagggttat tgatgttagc tgaattatca tcagtgggat
     3721 cattagcata tggagaatat tctcaaaaaa ctgttgaaat tgctaaatcc gataaggaat
     3781 ttgttattgg atttattgcc caacgtgata tgggtggcca agaagaagga tttgattggc
     3841 ttattatgac acctggagtt ggattagatg ataaaggtga tggattagga caacaatata
     3901 gaactgttga tgaagttgtt agcactggaa ctgatattat cattgttggt agaggattgt
     3961 ttggtaaagg aagagatcca gatattgaag gtaaaaggta tagaaatgct ggttggaatg
     4021 cttatttgaa aaagactggc caattataat cagtactgac aataaaaaga ttcttgtttt
     4081 caagaacttg tcatttgtat agttttttta tattgtagtt gttctatttt aatcaaatgt
     4141 tagcgtgatt tatatttttt ttcgcctcga catcatctgc ccagatgcga agttaagtgc
     4201 gcagaaagta atatcatgcg tcaatcgtat gtgaatgctg gtcgctatac tgctgtcgat
     4261 tcgatactaa cgccgccatc cagtgtcgaa aacgagctcg aattcataac ttcgtataat
     4321 gtatgctata cgaagttata tcgatgatat cagatccact agtggcctat gcggccgcgg
     4381 cagattaaaa tatgcttagt tgtagaacac agaaaaaatg agaaggttga tatactttta
     4441 ctcatattaa tcctgattta gtggcggtga tcctttttag attcgtttaa ttcataaaca
     4501 aggtacatca tatagcaata acaaatatgt aaactatgta gaaagactta tcaagtttag
     4561 ccattttggt cctcattcat gggcatttta tactaacttc atttttgccg aaataagaaa
     4621 ggatgtaatc ttaacctaaa atatgtaagc aattaagggc gcacgtagtc acttgcgaac
     4681 aaataggaaa agaaagaaac aagcacggat actgaatgac tgagtgtatt actacacgat
     4741 ttatctcaac gaatttaagt gtggaggacg atcggctgca cgtaggagtc aacgtttttt
     4801 tactgtgttc atcacaaaac cattaagctg agtcatccat attgcaaaca ataaggtagc
     4861 accgcttcca tcaaaacact agaacatacc gatgttttca cggatttatt ggaaacaaat
     4921 agcagatgag ggcgttttta cttgtaatga aggcgaatga gaaggcaagg cttacaggct
     4981 tgttgtttat gccaagcatc aatactcgct atggaggagc gccatcagcg ataacaacaa
     5041 agtttgcaag tgcttgcttg gatataaaag gcttgacgct taagataaat agggccatac
     5101 ttgcttgctt ttactatttt tgttcctcat aagcctgatt gactacgcta ttataaaccc
     5161 ctacattatc ctataatcaa cttcacttgt gtaacgttga atgagttcaa ctgagtaata
     5221 ctcggtcgga aatatacgct ccatctggca ggaaacatag gctgcattta tatcttgcac
     5281 aattatgccc ttcgtaggtg tatgttttca tgttttccat tatgtcaatg tagatgtatc
     5341 tttggacagg caacttttgg ccgcggatct gccggtctcc ctatagtgag tcgtattaat
     5401 ttcgataagc caggttaacc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt
     5461 gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct
     5521 gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga
     5581 taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc
     5641 cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg
     5701 ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg
     5761 aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt
     5821 tctcccttcg ggaagcgtgg cgctttctca atgctcacgc tgtaggtatc tcagttcggt
     5881 gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg
     5941 cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact
     6001 ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt
     6061 cttgaagtgg tggcctaact acggctacac tagaaggaca gtatttggta tctgcgctct
     6121 gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac
     6181 cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc
     6241 tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg
     6301 ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta
     6361 aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca
     6421 atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc
     6481 ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc
     6541 tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc
     6601 agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat
     6661 taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt
     6721 tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc
     6781 cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag
     6841 ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt
     6901 tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac
     6961 tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg
     7021 cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat
     7081 tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc
     7141 gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc
     7201 tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa
     7261 atgttgaata ctcatactct tcctttttca atattattga agcatttatc agggttattg
     7321 tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg
     7381 cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc attattatca tgacattaac
     7441 ctataaaaat aggcgtatca cgaggccctt tcgtctcgcg cgtttcggtg atgacggtga
     7501 aaacctctga cacatgcagc tcccggagac ggtcacagct tgtctgtaag cggatgccgg
     7561 gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc gggtgtcggg gctggcttaa
     7621 ctatgcggca tcagagcaga ttgtactgag agtgcaccat atggacatat tgtcgttaga
     7681 acgcggctac aattaataca taaccttatg tatcatacac atacgattta ggtga
//
