LOCUS       sequence-327697-        5422 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Knocking out GAL80 using KanMX4 marker.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5422)
  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 5422)
  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: pIR3DH8K
FEATURES             Location/Qualifiers
     source          1..5422
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(25..41)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(25..41)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(38..60)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    49..65
                     /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(73..79,80..97,98..103))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    118..139
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(256..273)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(427..1015)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(507..526)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1186..1977,1978..2046))
                     /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     1809..1828
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2047..2151)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2219..2237
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(2275..2297)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     2397..2416
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2610..2632
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2624..2641
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2625..2641
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        3481..3911
                     /gene="S. cerevisiae TPI1"
                     /label=TPI1 promoter
                     /note="strong constitutive promoter for yeast triose 
                     phosphate isomerase"
     CDS             3912..4721
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin"
                     /translation="MGKEKTHVSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(3979..3998)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
ORIGIN
        1 gcatgcaagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct
       61 cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg
      121 agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct
      181 gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg
      241 gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc
      301 ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg
      361 aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct
      421 ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca
      481 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct
      541 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc
      601 gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt
      661 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc
      721 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc
      781 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg
      841 gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc
      901 agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag
      961 cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga
     1021 tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat
     1081 tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag
     1141 ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat
     1201 cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc
     1261 cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat
     1321 accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag
     1381 ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg
     1441 ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc
     1501 tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca
     1561 acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg
     1621 tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc
     1681 actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta
     1741 ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc
     1801 aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg
     1861 ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc
     1921 cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc
     1981 aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat
     2041 actcatactc ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag
     2101 cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc
     2161 ccgaaaagtg ccacctgacg tctaagaaac cattattatc atgacattaa cctataaaaa
     2221 taggcgtatc acgaggccct ttcgtctcgc gcgtttcggt gatgacggtg aaaacctctg
     2281 acacatgcag ctcccggaga cggtcacagc ttgtctgtaa gcggatgccg ggagcagaca
     2341 agcccgtcag ggcgcgtcag cgggtgttgg cgggtgtcgg ggctggctta actatgcggc
     2401 atcagagcag attgtactga gagtgcacca tatgcggtgt gaaataccgc acagatgcgt
     2461 aaggagaaaa taccgcatca ggcgccattc gccattcagg ctgcgcaact gttgggaagg
     2521 gcgatcggtg cgggcctctt cgctattacg ccagctggcg aaagggggat gtgctgcaag
     2581 gcgattaagt tgggtaacgc cagggttttc ccagtcacga cgttgtaaaa cgacggccag
     2641 tgaattcgtt taaacggccg cctctgccat ggcaaagaat gctttccatg acgatcatcg
     2701 tagtgcccaa ttgggtgcct ctatgatggg tatggcttgg gcaagtgtct ttttatgtat
     2761 cgtggaattt atcctgctgg tcttctggtc tgttagggca aggttggcct ctacttactc
     2821 catcgacaat tcaagataca gaacctcctc cagatggaat cccttccata gagagaagga
     2881 gcaagcaact gacccaatat tgactgccac tggacctgaa gacatgcaac aaagtgcaag
     2941 catagtgggg ccttcttcca atgctaatcc ggtcactgcc actgctgcta cggaaaacca
     3001 acctaaaggt attaacttct tcactataag aaaatcacac gagcgcccgg acgatgtctc
     3061 tgtttaaaga tgctaagaga tagtgatgat atttcataaa taatgtaatt ctatatatgt
     3121 taattacctt ttttgcgagg catatttatg gtgaaggata agttttgacc atcaaagaag
     3181 gttaatgtgg ctgtggtttc agggtccata aagcttttca attcatcatt ttttttttat
     3241 tctttttttt gatttcggtt tccttgaaat ttttttgatt cggtaatctc cgaacagaag
     3301 gaagaacgaa ggaaggagca cagacttaga ttggtatata tacgcatatg tagtgttgaa
     3361 gaaacatgaa attgcccagt attcttaacc caactgcaca gaacaaaaac catttaaact
     3421 gtgaggacct taatacattc agacacttct gcggtatcac cctacttatt cccttcgaga
     3481 ttatatctag gaacccatca ggttggtgga agattacccg ttctaagact tttcagcttc
     3541 ctctattgat gttacacctg gacacccctt ttctggcatc cagtttttaa tcttcagtgg
     3601 catgtgagat tctccgaaat taattaaagc aatcacacaa ttctctcgga taccacctcg
     3661 gttgaaactg acaggtggtt tgttacgcat gctaatgcaa aggagcctat atacctttgg
     3721 ctcggctgct gtaacaggga atataaaggg cagcataatt taggagttta gtgaacttgc
     3781 aacatttact attttccctt cttacgtaaa tatttttctt tttaattcta aatcaatctt
     3841 tttcaatttt ttgtttgtat tcttttcttg cttaaatcta taactacaaa aaacacatac
     3901 ataaactaaa aatgggtaag gaaaagactc acgtttcgag gccgcgatta aattccaaca
     3961 tggatgctga tttatatggg tataaatggg ctcgcgataa tgtcgggcaa tcaggtgcga
     4021 caatctatcg attgtatggg aagcccgatg cgccagagtt gtttctgaaa catggcaaag
     4081 gtagcgttgc caatgatgtt acagatgaga tggtcagact aaactggctg acggaattta
     4141 tgcctcttcc gaccatcaag cattttatcc gtactcctga tgatgcatgg ttactcacca
     4201 ctgcgatccc cggcaaaaca gcattccagg tattagaaga atatcctgat tcaggtgaaa
     4261 atattgttga tgcgctggca gtgttcctgc gccggttgca ttcgattcct gtttgtaatt
     4321 gtccttttaa cagcgatcgc gtatttcgtc tcgctcaggc gcaatcacga atgaataacg
     4381 gtttggttga tgcgagtgat tttgatgacg agcgtaatgg ctggcctgtt gaacaagtct
     4441 ggaaagaaat gcataagctt ttgccattct caccggattc agtcgtcact catggtgatt
     4501 tctcacttga taaccttatt tttgacgagg ggaaattaat aggttgtatt gatgttggac
     4561 gagtcggaat cgcagaccga taccaggatc ttgccatcct atggaactgc ctcggtgagt
     4621 tttctccttc attacagaaa cggctttttc aaaaatatgg tattgataat cctgatatga
     4681 ataaattgca gtttcatttg atgctcgatg agtttttcta agactacaac aagagatctt
     4741 cggtctcaac cgtgcctaat gcagctccca taagagtcgg attcgtcggt ctcaacgcag
     4801 ccaaaggatg ggcaatcaag acacattacc ccgccatact gcaactatcg tcacaatttc
     4861 aaatcactgc cttatacagt ccaaaaattg agacttctat tgccaccatc cagcgtctaa
     4921 aattgagtaa tgccactgct tttcccactt tagagtcatt tgcatcatct tccactatag
     4981 atatgatagt gatagctatc caagtggcca gtcattatga cgttgttatg cctctcttgg
     5041 aattctccaa aaataatccg aacctcaagt atcttttcgt agaatgggcc cttgcatgtt
     5101 cactagatca agccgaatcc atttataagg ctgctgctga acgtggggtt caaaccatca
     5161 tctctttaca aggtcgtaaa tcaccatata ttttgagagc aaaagaatta atatctcaag
     5221 gctatatcgg cgacattaat tctatcgaga ttgctggaaa tggcggttgg tacggctacg
     5281 aaaggcctgt taaatcacca aaatacatct atgaaatcgg gaacggtgta gatctggtaa
     5341 ccacaacatt tggtcacaca atcgatattt tacaatacat gacaagttcg tacttttcca
     5401 ggataaatgc aatggtttaa ac
//
