LOCUS       sequence-495242-        4568 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Yeast gene tagging with mIAA7.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4568)
  AUTHORS   Hubbe P, Sharma C, Pajonk O, Peters N, Guschtschin-Schmidt N, 
            Friemel N, Schuck S
  TITLE     An optimised system for rapid auxin-inducible protein degradation in
            budding yeast
  JOURNAL   bioRxiv 2025.09.24.678194
REFERENCE   2  (bases 1 to 4568)
  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: "bioRxiv 
            2025.09.24.678194"
COMMENT     Sequence Label: pFA6a-mIAA7-ALFA-kanNP1
FEATURES             Location/Qualifiers
     source          1..4568
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             265..303
                     /codon_start=1
                     /product="rationally designed epitope tag that forms a 
                     stable alpha-helix (Gotzke et al., 2019)"
                     /label=ALFA-tag
                     /note="It may be beneficial to flank the ALFA-tag sequence 
                     with proline residues to preserve the helical structure."
                     /translation="SRLEEELRRRLTE"
     terminator      326..513
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(381..405)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     promoter        567..1086
                     /gene="S. cerevisiae TPI1"
                     /label=TPI1 promoter
                     /note="strong constitutive promoter for yeast triose 
                     phosphate isomerase"
     CDS             1087..1896
                     /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(1154..1173)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      1902..2099
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(2203..2222)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2204..2222)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2309..2326)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2480..3068)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2560..2579)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3239..4030,4031..4099))
                     /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     3862..3881
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4100..4204)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4272..4290
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(4328..4350)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4450..4469
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        4550..4568
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     4550..4567
                     /label=SP6
                     /note="SP6 promoter, forward primer"
ORIGIN
        1 acgcggccgc cagctgaagc ttcgtacgct gcaggtcgac ggatccccgg gttaattaac
       61 ggcttctctg agaccgtgga cctgatgctg aacctgcagt ccaataagga gggctctgtg
      121 gatctgaaga acgtgagcgc cgtgcctaag gagaagacca cactgaagga cccatccaag
      181 ccccctgcca aggcacaggt ggtgggatgg ccacccgtgc ggaactacag aaagaatatg
      241 atgacccagc agaagacaag ctcctcaaga ttagaagagg agctgaggag aagattgacc
      301 gagtaagcgc gccacttcta aataagcgaa tttcttatga tttatgattt ttattattaa
      361 ataagttata aaaaaaataa gtgtatacaa attttaaagt gactcttagg ttttaaaacg
      421 aaaattctta ttcttgagta actctttcct gtaggtcagg ttgctttctc aggtatagta
      481 tgaggtcgct cttattgacc acacctctac cggcagatcc gctagggata acagggtaat
      541 atagatctgt ttagcttgcc tcgtccccta cgttagtgtg agcgggattt aaactgtgag
      601 gaccttaata cattcagaca cttctgcggt atcaccctac ttattccctt cgagattata
      661 tctaggaacc catcaggttg gtggaagatt acccgttcta agacttttca gcttcctcta
      721 ttgatgttac acctggacac cccttttctg gcatccagtt tttaatcttc agtggcatgt
      781 gagattctcc gaaattaatt aaagcaatca cacaattctc tcggatacca cctcggttga
      841 aactgacagg tggtttgtta cgcatgctaa tgcaaaggag cctatatacc tttggctcgg
      901 ctgctgtaac agggaatata aagggcagca taatttagga gtttagtgaa cttgcaacat
      961 ttactatttt cccttcttac gtaaatattt ttctttttaa ttctaaatca atctttttca
     1021 attttttgtt tgtattcttt tcttgcttaa atctataact acaaaaaaca catacataaa
     1081 ctaaaaatgg gtaaggaaaa gactcacgtt tcgaggccgc gattaaattc caacatggat
     1141 gctgatttat atgggtataa atgggctcgc gataatgtcg ggcaatcagg tgcgacaatc
     1201 tatcgattgt atgggaagcc cgatgcgcca gagttgtttc tgaaacatgg caaaggtagc
     1261 gttgccaatg atgttacaga tgagatggtc agactaaact ggctgacgga atttatgcct
     1321 cttccgacca tcaagcattt tatccgtact cctgatgatg catggttact caccactgcg
     1381 atccccggca aaacagcatt ccaggtatta gaagaatatc ctgattcagg tgaaaatatt
     1441 gttgatgcgc tggcagtgtt cctgcgccgg ttgcattcga ttcctgtttg taattgtcct
     1501 tttaacagcg atcgcgtatt tcgtctcgct caggcgcaat cacgaatgaa taacggtttg
     1561 gttgatgcga gtgattttga tgacgagcgt aatggctggc ctgttgaaca agtctggaaa
     1621 gaaatgcata agcttttgcc attctcaccg gattcagtcg tcactcatgg tgatttctca
     1681 cttgataacc ttatttttga cgaggggaaa ttaataggtt gtattgatgt tggacgagtc
     1741 ggaatcgcag accgatacca ggatcttgcc atcctatgga actgcctcgg tgagttttct
     1801 ccttcattac agaaacggct ttttcaaaaa tatggtattg ataatcctga tatgaataaa
     1861 ttgcagtttc atttgatgct cgatgagttt ttctaatcag tactgacaat aaaaagattc
     1921 ttgttttcaa gaacttgtca tttgtatagt ttttttatat tgtagttgtt ctattttaat
     1981 caaatgttag cgtgatttat attttttttc gcctcgacat catctgccca gatgcgaagt
     2041 taagtgcgca gaaagtaata tcatgcgtca atcgtatgtg aatgctggtc gctatactgc
     2101 tgtcgattcg atactaacgc cgccatccag tttaaacgag ctcgaattca tcgatgatat
     2161 cagatccact agtggcctat gcggccgcgg atctgccggt ctccctatag tgagtcgtat
     2221 taatttcgat aagccaggtt aacctgcatt aatgaatcgg ccaacgcgcg gggagaggcg
     2281 gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc
     2341 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag
     2401 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
     2461 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
     2521 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
     2581 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     2641 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
     2701 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     2761 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     2821 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
     2881 agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg
     2941 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
     3001 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
     3061 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     3121 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa
     3181 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
     3241 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag
     3301 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca
     3361 gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc
     3421 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt
     3481 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg
     3541 ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca
     3601 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg
     3661 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca
     3721 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg
     3781 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct
     3841 cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca
     3901 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca
     3961 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg
     4021 tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac
     4081 ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt
     4141 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc
     4201 cgcgcacatt tccccgaaaa gtgccacctg acgtctaaga aaccattatt atcatgacat
     4261 taacctataa aaataggcgt atcacgaggc cctttcgtct cgcgcgtttc ggtgatgacg
     4321 gtgaaaacct ctgacacatg cagctcccgg agacggtcac agcttgtctg taagcggatg
     4381 ccgggagcag acaagcccgt cagggcgcgt cagcgggtgt tggcgggtgt cggggctggc
     4441 ttaactatgc ggcatcagag cagattgtac tgagagtgca ccatatggac atattgtcgt
     4501 tagaacgcgg ctacaattaa tacataacct tatgtatcat acacatacga tttaggtgac
     4561 actataga
//
