LOCUS       sequence-402961-        6826 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  C-terminal tagging of target proteins with AID*/3xHA degron.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6826)
  AUTHORS   Milholland KL, Gregor JB, Hoda S, Piriz-Antunez S, Duenas-Santero E,
            Vu BG, Patel KP, Moye-Rowley WS, Vazquez de Aldana CR, Correa-Bordes
            J, Briggs SD, Hall MC
  TITLE     Rapid, efficient auxin-inducible protein degradation in Candida 
            pathogens.
  JOURNAL   mSphere. 2023 Aug 18:e0028323. doi: 10.1128/msphere.00283-23.
  PUBMED    37594261
REFERENCE   2  (bases 1 to 6826)
  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: "mSphere. 
            2023 Aug 18:e0028323. doi: 10.1128/msphere.00283-23."
COMMENT     Sequence Label: pHLP700
FEATURES             Location/Qualifiers
     source          1..6826
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(33..55)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     155..174
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        198..413
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     CDS             414..1217
                     /codon_start=1
                     /gene="S. cerevisiae URA3"
                     /product="orotidine-5'-phosphate decarboxylase, required 
                     for uracil biosynthesis"
                     /label=URA3
                     /note="yeast auxotrophic marker, counterselectable with 
                     5-fluoroorotic acid (5-FOA)"
                     /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
                     ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     rep_origin      complement(1348..1803)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1485..1506)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1697..1716
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1929..1951
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1943..1960
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1944..1960
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1967..1986
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1967..1985
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     2015..2031
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2015..2031
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     CDS             2069..2200
                     /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             2216..2305
                     /codon_start=1
                     /product="three tandem HA epitope tags"
                     /label=3xHA
                     /translation="YPYDVPDYAGYPYDVPDYAGSYPYDVPDYA"
     terminator      2319..2506
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(2374..2398)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     primer_bind     complement(4501..4517)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4525..4541)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        complement(4570..4588)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4609..4625)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4609..4625)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4622..4644)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4633..4649
                     /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(4657..4663,4664..4681,4682..4687))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4702..4723
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4840..4857)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5011..5599)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5091..5110)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5770..6561,6562..6630))
                     /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     6393..6412
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6631..6735)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6803..6821
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 cgtctcgcgc gtttcggtga tgacggtgaa aacctctgac acatgcagct cccggagacg
       61 gtcacagctt gtctgtaagc ggatgccggg agcagacaag cccgtcaggg cgcgtcagcg
      121 ggtgttggcg ggtgtcgggg ctggcttaac tatgcggcat cagagcagat tgtactgaga
      181 gtgcaccata ccaccttttc aattcatcat ttttttttta ttcttttttt tgatttcggt
      241 ttccttgaaa tttttttgat tcggtaatct ccgaacagaa ggaagaacga aggaaggagc
      301 acagacttag attggtatat atacgcatat gtagtgttga agaaacatga aattgcccag
      361 tattcttaac ccaactgcac agaacaaaaa cctgcaggaa acgaagataa atcatgtcga
      421 aagctacata taaggaacgt gctgctactc atcctagtcc tgttgctgcc aagctattta
      481 atatcatgca cgaaaagcaa acaaacttgt gtgcttcatt ggatgttcgt accaccaagg
      541 aattactgga gttagttgaa gcattaggtc ccaaaatttg tttactaaaa acacatgtgg
      601 atatcttgac tgatttttcc atggagggca cagttaagcc gctaaaggca ttatccgcca
      661 agtacaattt tttactcttc gaagacagaa aatttgctga cattggtaat acagtcaaat
      721 tgcagtactc tgcgggtgta tacagaatag cagaatgggc agacattacg aatgcacacg
      781 gtgtggtggg cccaggtatt gttagcggtt tgaagcaggc ggcagaagaa gtaacaaagg
      841 aacctagagg ccttttgatg ttagcagaat tgtcatgcaa gggctcccta tctactggag
      901 aatatactaa gggtactgtt gacattgcga agagcgacaa agattttgtt atcggcttta
      961 ttgctcaaag agacatgggt ggaagagatg aaggttacga ttggttgatt atgacacccg
     1021 gtgtgggttt agatgacaag ggagacgcat tgggtcaaca gtatagaacc gtggatgatg
     1081 tggtctctac aggatctgac attattattg ttggaagagg actatttgca aagggaaggg
     1141 atgctaaggt agagggtgaa cgttacagaa aagcaggctg ggaagcatat ttgagaagat
     1201 gcggccagca aaactaaaaa actgtattat aagtaaatgc atgtatacta aactcacaaa
     1261 ttagagcttc aatttaatta tatcagttat taccctatgc ggtgtgaaat accgcacaga
     1321 tgcgtaagga gaaaataccg catcaggaaa ttgtaagcgt taatattttg ttaaaattcg
     1381 cgttaaattt ttgttaaatc agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc
     1441 cttataaatc aaaagaatag accgagatag ggttgagtgt tgttccagtt tggaacaaga
     1501 gtccactatt aaagaacgtg gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg
     1561 atggcccact acgtgaacca tcaccctaat caagtttttt ggggtcgagg tgccgtaaag
     1621 cactaaatcg gaaccctaaa gggagccccc gatttagagc ttgacgggga aagccggcga
     1681 acgtggcgag aaaggaaggg aagaaagcga aaggagcggg cgctagggcg ctggcaagtg
     1741 tagcggtcac gctgcgcgta accaccacac ccgccgcgct taatgcgccg ctacagggcg
     1801 cgtccattcg ccattcaggc tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc
     1861 gctattacgc cagctggcga aagggggatg tgctgcaagg cgattaagtt gggtaacgcc
     1921 agggttttcc cagtcacgac gttgtaaaac gacggccagt gaattgtaat acgactcact
     1981 atagggcgaa ttggagctcc accgcggtgg ccgctctaga actagtggat ccggtgcagg
     2041 cgccggggct ggtgctgggg caggtacccc taaagatcca gccaaacctc cggccaaggc
     2101 acaagttgtg ggatggccac cggtgagatc ataccggaag aacgtgatgg tttcctgcca
     2161 aaaatcaagc ggtggcccgg aggcggcggc gttcgtgaag gtatcacccg ggaaataccc
     2221 atacgatgtt cctgactatg cgggctatcc ctatgacgtc ccggactatg caggatccta
     2281 tccatatgac gttccagatt acgcttctag ataacgaggc gaatttctta tgatttatga
     2341 tttttattat taaataagtt ataaaaaaaa taagtgtata caaattttaa agtgactctt
     2401 aggttttaaa acgaaaattc ttattcttga gtaactcttt cctgtaggtc aggttgcttt
     2461 ctcaggtata gcatgaggtc gctcttattg accacacctc taccggcatg ccgagcaaat
     2521 gcctgcaaat cgctccccat ttcacccaat tgtagatatg ctaactccag caatgagttg
     2581 atgaatctcg gtgtgtattt tatgtcctca gaggacaaca cctgttgtaa tcgtgtcgag
     2641 cgtcaaaact agagaataat aaagaaaacg atcttttcaa aaagaaaaaa ccttttagtt
     2701 ttcctttgtt gttgttgtgg gtgtgtgcta tttatattat atagtttact cataatacca
     2761 taaaatattc ggtttgatta ggttatttta ataagctaat ttgtttctaa tcgtgtaatt
     2821 tatgctgtgt atattaagta gtgtgtgcac tgcccaaaaa tgtttgttgt ttatagtcgg
     2881 ttaaagagaa aaaagaaaaa aagatccata cacacacgtt aattagttgt tcaacgtaat
     2941 acactcatat tttgttctta tttgctttcg gtcgctgttc tcaccaagat ttattgccaa
     3001 cgaaacaatt ttttttatat attttcagat ttttcttttt ttcctttcct ttccttttct
     3061 aattttcact cctggttttc tttctttctt agaaacatta tctcgatatt aatattaaaa
     3121 aaatataatc attcaaaatg gacggtggta tgttttagtt tagcttcaat tctaattgat
     3181 tgattaatca gttgattggt ttcaatatga caaatgggta gggtgggaaa acttcatttt
     3241 caattcagat caaacttttt tgttgtcgac ataatatttc tcgtttggga tgttactgtc
     3301 acattaataa tacacacaca tcagcttata attttgaaag taatttatca gatatgttgt
     3361 gacgatcagt ggaaatggct aacttcaatg tatctgttct tccccttttt caaagttcac
     3421 gttttttgat tgattgattg atctgtcggc agtggtttca aaaccattcg gtgagtaatc
     3481 ctatcaatca atgttacgac aaaaggctca atattcaaaa ttgcaatgtt ttatgttttc
     3541 ctacgtgtac ttgtgcaagg caattgattc aacattgctt ttggtgtttg acgagtttct
     3601 agtttggact tgtgttgtta tctgggctat acagatttcc cggctcacta tgaatttttt
     3661 ttttcgacgc tcagtgcaca caactataaa caacacaaac acaaacacag caagaaaaaa
     3721 aaaaaacgaa cattgaattg aaaccaagcc aactgaaaaa ttccttattt aaatgactgt
     3781 catactaacc catttttata gaagaagttg ctgctttagt tatcgataac ggttctcata
     3841 tgaaaatttc ggtgatccct gagcaggtgg cggaaacatt ggatgctgag aaccatttca
     3901 ttgttcgtga agtgttcgat gtgcacctat ccgaccaagg ctttgaacta tctaccagaa
     3961 gtgtgagccc ctaccggaag gattacatct cggatgatga ctctgatgaa gactctgctt
     4021 gctatggcgc attcatcgac caagagcttg tcgggaagat tgaactcaac tcaacatgga
     4081 acgatctagc ctctatcgaa cacattgttg tgtcgcacac gcaccgaggc aaaggagtcg
     4141 cgcacagtct catcgaattt gcgaaaaagt gggcactaag cagacagctc cttggcatac
     4201 gattagagac acaaacgaac aatgtacctg cctgcaattt gtacgcaaaa tgtggcttta
     4261 ctctcggcgg cattgacctc ttcacgtata aaactagacc tcaagtctcg aacgaaacag
     4321 cgatgtactg gtactggttc tcgggagcac aggatgacgc ctaacatatg tgaagtgtga
     4381 agggggagat tttcacttta ttagatttgt atatatgtat aataaataaa taaataagtt
     4441 aaataaataa ttagataagg gtggtaatta ttactattta caatcaaagg tggtcctgca
     4501 cgataccgtc gacctcgagg gtatcgatac cgtcgacctc gagggggggc ccggtaccag
     4561 cttttgttcc ctttagtgag ggttaatttc gagcttggcg taatcatggt catagctgtt
     4621 tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa
     4681 gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact
     4741 gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc
     4801 ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg
     4861 ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc
     4921 cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag
     4981 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca
     5041 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca
     5101 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg
     5161 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag
     5221 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt
     5281 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca
     5341 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg
     5401 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt
     5461 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc
     5521 cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg
     5581 cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg
     5641 gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta
     5701 gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg
     5761 gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg
     5821 ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc
     5881 atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc
     5941 agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc
     6001 ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag
     6061 tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat
     6121 ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg
     6181 caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt
     6241 gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag
     6301 atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg
     6361 accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt
     6421 aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct
     6481 gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac
     6541 tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat
     6601 aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat
     6661 ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca
     6721 aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtctaag aaaccattat
     6781 tatcatgaca ttaacctata aaaataggcg tatcacgagg cccttt
//
