LOCUS       sequence-327175-        5900 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Integrative expression of nucleus-targeting yEGFP-AID*(mini auxin 
            inducible degron)-CUP1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5900)
  AUTHORS   Lu Z, Peng B, Ebert BE, Dumsday G, Vickers CE
  TITLE     Auxin-mediated protein depletion for metabolic engineering in 
            terpene-producing yeast.
  JOURNAL   Nat Commun. 2021 Feb 16;12(1):1051. doi: 10.1038/s41467-021-21313-1.
  PUBMED    33594068
REFERENCE   2  (bases 1 to 5900)
  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: "Nat 
            Commun."; date: "2021-02-16"; volume: "12(1):1051. doi"; pages: " 
            10.1038/s41467-021-21313-1"
COMMENT     Sequence Label: pILGFP105
FEATURES             Location/Qualifiers
     source          1..5900
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             431..1141
                     /codon_start=1
                     /product="yeast-enhanced green fluorescent protein"
                     /label=yeGFP
                     /note="codons optimized for translation in S. cerevisiae 
                     and C. albicans"
                     /translation="SKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKF
                     ICTTGKLPVPWPTLVTTFGYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDGN
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKVN
                     FKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLEF
                     VTAAGITHGMDELYK"
     CDS             1148..1279
                     /codon_start=1
                     /product="minimal degron from the auxin-responsive IAA17 
                     protein of Arabidopsis thaliana (Morawska and Ulrich, 
                     2013)"
                     /label=AID*
                     /note="acts as an auxin-inducible degron (AID) in the 
                     presence of TIR1"
                     /translation="PKDPAKPPAKAQVVGWPPVRSYRKNVMVSCQKSSGGPEAAAFVK"
     CDS             1472..1492
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     terminator      1502..1772
                     /gene="S. cerevisae PGK1"
                     /label=PGK1 terminator
                     /note="transcription terminator for 3-phosphoglycerate 
                     kinase gene"
     primer_bind     complement(1921..1937)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1921..1937)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1934..1956)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1945..1961
                     /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(1969..1975,1976..1993,1994..1999))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2014..2035
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2152..2169)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2323..2911)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2403..2422)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3082..3873,3874..3942))
                     /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     3705..3724
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3943..4047)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4115..4133
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(4171..4193)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4293..4312
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4506..4528
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4520..4537
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4521..4537
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(5651..5684)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        join(5895..5900,1..408)
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
ORIGIN
        1 ccacacacca tagcttcaaa atgtttctac tcctttttta ctcttccaga ttttctcgga
       61 ctccgcgcat cgccgtacca cttcaaaaca cccaagcaca gcatactaaa tttcccctct
      121 ttcttcctct agggtgtcgt taattacccg tactaaaggt ttggaaaaga aaaaagagac
      181 cgcctcgttt ctttttcttc gtcgaaaaag gcaataaaaa tttttatcac gtttcttttt
      241 cttgaaaatt tttttttttg atttttttct ctttcgatga cctcccattg atatttaagt
      301 taataaacgg tcttcaattt ctcaagtttc agtttcattt ttcttgttct attacaactt
      361 tttttacttc ttgctcatta gaaagaaagc atagcaatct aatctaagtt ttaattacaa
      421 aatgggatcc tctaaaggtg aagaattatt cactggtgtt gtcccaattt tggttgaatt
      481 agatggtgat gttaatggtc acaaattttc tgtctccggt gaaggtgaag gtgatgctac
      541 ttacggtaaa ttgaccttaa aatttatttg tactactggt aaattgccag ttccatggcc
      601 aaccttagtc actactttcg gttatggtgt tcaatgtttt gcgagatacc cagatcatat
      661 gaaacaacat gactttttca agtctgccat gccagaaggt tatgttcaag aaagaactat
      721 ttttttcaaa gatgacggta actacaagac cagagctgaa gtcaagtttg aaggtgatac
      781 cttagttaat agaatcgaat taaaaggtat tgattttaaa gaagatggta acattttagg
      841 tcacaaattg gaatacaact ataactctca caatgtttac atcatggctg acaaacaaaa
      901 gaatggtatc aaagttaact tcaaaattag acacaacatt gaagatggtt ctgttcaatt
      961 agctgaccat tatcaacaaa atactccaat tggtgatggt ccagtcttgt taccagacaa
     1021 ccattactta tccactcaat ctgccttatc caaagatcca aacgaaaaga gagaccacat
     1081 ggtcttgtta gaatttgtta ctgctgctgg tattacccat ggtatggatg aattgtacaa
     1141 aagatctcca aaagacccag ctaaacctcc agccaaggcc caagtcgtcg gctggccacc
     1201 agttaggtct tacagaaaga acgtaatggt tagttgtcaa aagtcctccg gtggtccaga
     1261 agctgctgcc tttgttaaga tgttcagcga attaattaac ttccaaaatg aaggtcatga
     1321 gtgccaatgc caatgtggta gctgcaaaaa taatgaacaa tgccaaaaat catgtagctg
     1381 cccaacgggg tgtaacagcg acgacaaatg cccctgcggt aacaagtctg aagaaaccaa
     1441 gaagtcatgc tgctctggga aatgtacccc acctaagaag aaaagaaagg tttaaactag
     1501 tattgaattg aattgaaatc gatagatcaa tttttttctt ttctctttcc ccatccttta
     1561 cgctaaaata atagtttatt ttattttttg aatatttttt atttatatac gtatatatag
     1621 actattattt atcttttaat gattattaag atttttatta aaaaaaaatt cgctcctctt
     1681 ttaatgcctt tatgcagttt ttttttccca ttcgatattt ctatgttcgg gttcagcgta
     1741 ttttaagttt aataactcga aaattctgcg ttcgttaaag ctttcgagaa ggatattatt
     1801 tgctaagtaa atgcatgtat actaaactca caaattagag cttcaattta attatatcag
     1861 ttattacccg ggaatctcgg tcgtaatgat ttaaatgcat gcaagcttgg cgtaatcatg
     1921 gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca acatacgagc
     1981 cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc
     2041 gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat
     2101 cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac
     2161 tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt
     2221 aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca
     2281 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc
     2341 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
     2401 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
     2461 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag
     2521 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca
     2581 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa
     2641 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc
     2701 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag
     2761 aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg
     2821 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca
     2881 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc
     2941 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag
     3001 gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata
     3061 tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat
     3121 ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg
     3181 ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc
     3241 tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc
     3301 aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc
     3361 gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc
     3421 gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc
     3481 ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa
     3541 gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat
     3601 gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata
     3661 gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca
     3721 tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag
     3781 gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc
     3841 agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc
     3901 aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata
     3961 ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta
     4021 gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac ctgacgtcta
     4081 agaaaccatt attatcatga cattaaccta taaaaatagg cgtatcacga ggccctttcg
     4141 tctcgcgcgt ttcggtgatg acggtgaaaa cctctgacac atgcagctcc cggagacggt
     4201 cacagcttgt ctgtaagcgg atgccgggag cagacaagcc cgtcagggcg cgtcagcggg
     4261 tgttggcggg tgtcggggct ggcttaacta tgcggcatca gagcagattg tactgagagt
     4321 gcaccatatg cggtgtgaaa taccgcacag atgcgtaagg agaaaatacc gcatcaggcg
     4381 ccattcgcca ttcaggctgc gcaactgttg ggaagggcga tcggtgcggg cctcttcgct
     4441 attacgccag ctggcgaaag ggggatgtgc tgcaaggcga ttaagttggg taacgccagg
     4501 gttttcccag tcacgacgtt gtaaaacgac ggccagtgaa ttcatttaaa tttttttgat
     4561 tcggtaatct ccgaacagaa ggaagaacga aggaaggagc acagacttag attggtatat
     4621 atacgcatat gtagtgttga agaaacatga aattgcccag tattcttaac ccaactgcac
     4681 agaacaaaaa cctgcaggaa acgaagataa aacatgtcca caaaatcata taccagtaga
     4741 gctgagactc atgcaagtcc ggttgcatcg aaacttttac gtttaatgga tgaaaagaag
     4801 accaatttgt gtgcttctct tgacgttcgt tcgactgatg agctattgaa acttgttgaa
     4861 acgttgggtc catacatttg ccttttgaaa acacacgttg atatcttgga tgatttcagt
     4921 tatgagggta ctgtcgttcc attgaaagca ttggcagaga aatacaagtt cttgatattt
     4981 gaggacagaa aattcgccga tatcggtaac acagtcaaat tacaatatac atcgggcgtt
     5041 taccgtatcg cagaatggtc tgatatcacc aacgcccacg gggttactgg tgctggtatt
     5101 gttgctggct tgaaacaagg tgcgcaagag gtcaccaaag aaccaagggg attattgatg
     5161 cttgctgaat tgtcttccaa gggttctcta gcacacggtg aatatactaa gggtaccgtt
     5221 gatattgcaa agagtgataa agatttcgtt attgggttca ttgctcagaa cgatatggga
     5281 ggaagagaag aagggtttga ttggctaatc atgaccccag gtgtaggttt agacgacaaa
     5341 ggcgatgcat tgggtcagca gtacagaacc gtcgacgaag ttgtaagtgg tggatcagat
     5401 atcatcattg ttggcagagg acttttcgcc aagggtagag atcctaaggt tgaaggtgaa
     5461 agatacagaa atgctggatg ggaagcgtac caaaagagaa tcagcgctcc ccattaatta
     5521 tacaggaaac ttaatagaac aaatcacata tttaatctaa tagccacctg cattggcacg
     5581 gtgcaacact acttcaactt catcttacaa aaagatcacg tgatctgttg tattgggatc
     5641 tctagaccta ataacttcgt atagcataca ttatacgaag ttatattaag ggttgctcga
     5701 ccagaaagcg accacccaac tttggctgat aatagcgtat aaacaatgca tactttgtac
     5761 gttcaaaata caatgcagta gatatattta tgcatattac atataataca tatcacatag
     5821 gaagcaacag gcgcgttgga cttttaattt tcgaggaccg cgaatcctta catcacaccc
     5881 aatcccccac aagtgatccc
//
