LOCUS       sequence-451858-        5089 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  To insert biotin ligase, miniTurboID, at the C-terminus of genes; 
            yeast BioID, KanMX6 marker.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5089)
  AUTHORS   Pfannenstein J, Tyryshkin M, Gulden ME, Doud EH, Mosley AL, Reese JC
  TITLE     Characterization of BioID tagging systems in budding yeast and 
            exploring the interactome of the Ccr4-Not complex.
  JOURNAL   G3 (Bethesda). 2024 Sep 13:jkae221. doi: 10.1093/g3journal/jkae221.
  PUBMED    39271111
REFERENCE   2  (bases 1 to 5089)
  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: "G3 
            (Bethesda). 2024 Sep 13:jkae221. doi: 10.1093/g3journal/jkae221."
COMMENT     Sequence Label: pFA6a-mTID-HA3-KanMX6
FEATURES             Location/Qualifiers
     source          1..5089
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             880..969
                     /codon_start=1
                     /product="three tandem HA epitope tags"
                     /label=3xHA
                     /translation="YPYDVPDYAGYPYDVPDYAGSYPYDVPDYA"
     terminator      1002..1189
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(1057..1081)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     gene            1264..2620
                     /label=kanMX
                     /note="yeast selectable marker conferring kanamycin 
                     resistance (Wach et al., 1994)"
     promoter        1264..1607
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             1608..2417
                     /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(1675..1694)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      2423..2620
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(2724..2743)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2725..2743)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2830..2847)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3001..3589)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3081..3100)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3760..4551,4552..4620))
                     /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     4383..4402
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4621..4725)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4793..4811
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(4849..4871)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4971..4990
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        5071..5089
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     5071..5088
                     /label=SP6
                     /note="SP6 promoter, forward primer"
ORIGIN
        1 acgcggccgc cagctgaagc ttcgtacgct gcaggtcgac ggatccccgg gttaattaac
       61 gcaagagacc cgcccgtagc aactatgatt cctttactga atgcgaaaca gatattgggt
      121 caattagatg ggggttccgt agctgtcctt ccggtggtgg atagcacgaa ccaatatcta
      181 cttgatagga tcggggaatt gaagtcaggt gatgcttgca tcgcagagta ccaacaggct
      241 ggtagaggga gcagaggcag gaagtggttt tcaccatttg gagcaaatct gtacttgagc
      301 atgttctgga gacttaagag aggtccagcg gcgatagggc tgggacccgt tatcggaatt
      361 gtgatggcag aagccctgcg taaactaggt gctgataaag tacgtgtcaa atggccaaac
      421 gatttatatc tacaagacag aaagttggcg gggattttgg tggaacttgc cggcataacc
      481 ggggacgccg cgcagatagt cattggtgct ggaatcaatg tggcgatgag acgtgttgag
      541 gagtcagttg taaatcaggg ttggataact cttcaggagg ccgggataaa cttagacagg
      601 aatactctag ctgccatgct tatacgtgag ctacgtgccg ccctggagct tttcgagcaa
      661 gaaggtctag ctccttacct tagccgttgg gagaagttgg ataacttcat caatagacca
      721 gttaagttaa tcatagggga caaagagata ttcgggatca gtagaggtat agataagcag
      781 ggagcgctac tactggagca ggatggggtt atcaaaccgt ggatgggcgg ggagatatcc
      841 ctgaggtctg ccgagaaggc tagcttaatt aacatctttt acccatacga tgttcctgac
      901 tatgcgggct atccctatga cgtcccggac tatgcaggat cctatccata tgacgttcca
      961 gattacgctg ctcagtgctg aggcgcgcca cttctaaata agcgaatttc ttatgattta
     1021 tgatttttat tattaaataa gttataaaaa aaataagtgt atacaaattt taaagtgact
     1081 cttaggtttt aaaacgaaaa ttcttattct tgagtaactc tttcctgtag gtcaggttgc
     1141 tttctcaggt atagtatgag gtcgctctta ttgaccacac ctctaccggc agatccgcta
     1201 gggataacag ggtaatatag atctgtttag cttgcctcgt ccccgccggg tcacccggcc
     1261 agcgacatgg aggcccagaa taccctcctt gacagtcttg acgtgcgcag ctcaggggca
     1321 tgatgtgact gtcgcccgta catttagccc atacatcccc atgtataatc atttgcatcc
     1381 atacattttg atggccgcac ggcgcgaagc aaaaattacg gctcctcgct gcagacctgc
     1441 gagcagggaa acgctcccct cacagacgcg ttgaattgtc cccacgccgc gcccctgtag
     1501 agaaatataa aaggttagga tttgccactg aggttcttct ttcatatact tccttttaaa
     1561 atcttgctag gatacagttc tcacatcaca tccgaacata aacaaccatg ggtaaggaaa
     1621 agactcacgt ttcgaggccg cgattaaatt ccaacatgga tgctgattta tatgggtata
     1681 aatgggctcg cgataatgtc gggcaatcag gtgcgacaat ctatcgattg tatgggaagc
     1741 ccgatgcgcc agagttgttt ctgaaacatg gcaaaggtag cgttgccaat gatgttacag
     1801 atgagatggt cagactaaac tggctgacgg aatttatgcc tcttccgacc atcaagcatt
     1861 ttatccgtac tcctgatgat gcatggttac tcaccactgc gatccccggc aaaacagcat
     1921 tccaggtatt agaagaatat cctgattcag gtgaaaatat tgttgatgcg ctggcagtgt
     1981 tcctgcgccg gttgcattcg attcctgttt gtaattgtcc ttttaacagc gatcgcgtat
     2041 ttcgtctcgc tcaggcgcaa tcacgaatga ataacggttt ggttgatgcg agtgattttg
     2101 atgacgagcg taatggctgg cctgttgaac aagtctggaa agaaatgcat aagcttttgc
     2161 cattctcacc ggattcagtc gtcactcatg gtgatttctc acttgataac cttatttttg
     2221 acgaggggaa attaataggt tgtattgatg ttggacgagt cggaatcgca gaccgatacc
     2281 aggatcttgc catcctatgg aactgcctcg gtgagttttc tccttcatta cagaaacggc
     2341 tttttcaaaa atatggtatt gataatcctg atatgaataa attgcagttt catttgatgc
     2401 tcgatgagtt tttctaatca gtactgacaa taaaaagatt cttgttttca agaacttgtc
     2461 atttgtatag tttttttata ttgtagttgt tctattttaa tcaaatgtta gcgtgattta
     2521 tatttttttt cgcctcgaca tcatctgccc agatgcgaag ttaagtgcgc agaaagtaat
     2581 atcatgcgtc aatcgtatgt gaatgctggt cgctatactg ctgtcgattc gatactaacg
     2641 ccgccatcca gtttaaacga gctcgaattc atcgatgata tcagatccac tagtggccta
     2701 tgcggccgcg gatctgccgg tctccctata gtgagtcgta ttaatttcga taagccaggt
     2761 taacctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc
     2821 ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc
     2881 agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa
     2941 catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt
     3001 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
     3061 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg
     3121 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
     3181 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
     3241 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
     3301 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
     3361 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc
     3421 taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac
     3481 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg
     3541 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt
     3601 gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt
     3661 catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa
     3721 atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga
     3781 ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt
     3841 gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg
     3901 agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga
     3961 gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga
     4021 agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg
     4081 catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc
     4141 aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc
     4201 gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca
     4261 taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac
     4321 caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg
     4381 ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc
     4441 ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg
     4501 tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac
     4561 aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat
     4621 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
     4681 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
     4741 agtgccacct gacgtctaag aaaccattat tatcatgaca ttaacctata aaaataggcg
     4801 tatcacgagg ccctttcgtc tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat
     4861 gcagctcccg gagacggtca cagcttgtct gtaagcggat gccgggagca gacaagcccg
     4921 tcagggcgcg tcagcgggtg ttggcgggtg tcggggctgg cttaactatg cggcatcaga
     4981 gcagattgta ctgagagtgc accatatgga catattgtcg ttagaacgcg gctacaatta
     5041 atacataacc ttatgtatca tacacatacg atttaggtga cactataga
//
