LOCUS       sequence-453627-        6807 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses TurboID-FRB fusion from HIS3 promoter; yeast BioID.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6807)
  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 6807)
  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: pRS416-HIS3p-TID-FRB-HA3
FEATURES             Location/Qualifiers
     source          1..6807
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     551..570
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(789..893)
                     /gene="bla"
                     /label=AmpR promoter
     misc_feature    930..1433
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     primer_bind     1474..1492
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(1530..1552)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     1652..1671
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        1695..1910
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     CDS             1911..2714
                     /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(2845..3300)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2982..3003)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3194..3213
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3426..3448
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3440..3457
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3441..3457
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3467..3486
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3467..3485
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        3904..4091
                     /gene="S. cerevisiae HIS3"
                     /label=HIS3 promoter
     CDS             5112..5390
                     /codon_start=1
                     /product="T2098L mutant of FKBP-rapamycin binding domain of
                     human FRAP"
                     /label=FRB*
                     /note="binds synthetic ligands such as AP21967"
                     /translation="ILWHEMWHEGLEEASRLYFGERNVKGMFEVLEPLHAMMERGPQTL
                     KETSFNQAYGRDLMEAQEWCRKYMKSGNVKDLLQAWDLYYHVFRRISK"
     CDS             5397..5486
                     /codon_start=1
                     /product="three tandem HA epitope tags"
                     /label=3xHA
                     /translation="YPYDVPDYAGYPYDVPDYAGSYPYDVPDYA"
     primer_bind     complement(5535..5555)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(5535..5553)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5574..5590)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5574..5590)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5587..5609)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5598..5614
                     /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(5622..5628,5629..5646,5647..5652))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5667..5688
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5805..5822)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5976..6564)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6056..6075)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6735..6807,1..719,720..788))
                     /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"
ORIGIN
        1 cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg
       61 ataccgcgag acccacgctc accggctcca gatttatcag caataaacca gccagccgga
      121 agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt
      181 tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt
      241 gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc
      301 caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc
      361 ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca
      421 gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag
      481 tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg
      541 tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa
      601 cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa
      661 cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga
      721 gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga
      781 atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg
      841 agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt
      901 ccccgaaaag tgccacctgg gtccttttca tcacgtgcta taaaaataat tataatttaa
      961 attttttaat ataaatatat aaattaaaaa tagaaagtaa aaaaagaaat taaagaaaaa
     1021 atagtttttg ttttccgaag atgtaaaaga ctctaggggg atcgccaaca aatactacct
     1081 tttatcttgc tcttcctgct ctcaggtatt aatgccgaat tgtttcatct tgtctgtgta
     1141 gaagaccaca cacgaaaatc ctgtgatttt acattttact tatcgttaat cgaatgtata
     1201 tctatttaat ctgcttttct tgtctaataa atatatatgt aaagtacgct ttttgttgaa
     1261 attttttaaa cctttgttta tttttttttc ttcattccgt aactcttcta ccttctttat
     1321 ttactttcta aaatccaaat acaaaacata aaaataaata aacacagagt aaattcccaa
     1381 attattccat cattaaaaga tacgaggcgc gtgtaagtta caggcaagcg atccgtctaa
     1441 gaaaccatta ttatcatgac attaacctat aaaaataggc gtatcacgag gccctttcgt
     1501 ctcgcgcgtt tcggtgatga cggtgaaaac ctctgacaca tgcagctccc ggagacggtc
     1561 acagcttgtc tgtaagcgga tgccgggagc agacaagccc gtcagggcgc gtcagcgggt
     1621 gttggcgggt gtcggggctg gcttaactat gcggcatcag agcagattgt actgagagtg
     1681 caccatacca ccttttcaat tcatcatttt ttttttattc ttttttttga tttcggtttc
     1741 cttgaaattt ttttgattcg gtaatctccg aacagaagga agaacgaagg aaggagcaca
     1801 gacttagatt ggtatatata cgcatatgta gtgttgaaga aacatgaaat tgcccagtat
     1861 tcttaaccca actgcacaga acaaaaacct gcaggaaacg aagataaatc atgtcgaaag
     1921 ctacatataa ggaacgtgct gctactcatc ctagtcctgt tgctgccaag ctatttaata
     1981 tcatgcacga aaagcaaaca aacttgtgtg cttcattgga tgttcgtacc accaaggaat
     2041 tactggagtt agttgaagca ttaggtccca aaatttgttt actaaaaaca catgtggata
     2101 tcttgactga tttttccatg gagggcacag ttaagccgct aaaggcatta tccgccaagt
     2161 acaatttttt actcttcgaa gacagaaaat ttgctgacat tggtaataca gtcaaattgc
     2221 agtactctgc gggtgtatac agaatagcag aatgggcaga cattacgaat gcacacggtg
     2281 tggtgggccc aggtattgtt agcggtttga agcaggcggc agaagaagta acaaaggaac
     2341 ctagaggcct tttgatgtta gcagaattgt catgcaaggg ctccctatct actggagaat
     2401 atactaaggg tactgttgac attgcgaaga gcgacaaaga ttttgttatc ggctttattg
     2461 ctcaaagaga catgggtgga agagatgaag gttacgattg gttgattatg acacccggtg
     2521 tgggtttaga tgacaaggga gacgcattgg gtcaacagta tagaaccgtg gatgatgtgg
     2581 tctctacagg atctgacatt attattgttg gaagaggact atttgcaaag ggaagggatg
     2641 ctaaggtaga gggtgaacgt tacagaaaag caggctggga agcatatttg agaagatgcg
     2701 gccagcaaaa ctaaaaaact gtattataag taaatgcatg tatactaaac tcacaaatta
     2761 gagcttcaat ttaattatat cagttattac cctatgcggt gtgaaatacc gcacagatgc
     2821 gtaaggagaa aataccgcat caggaaattg taagcgttaa tattttgtta aaattcgcgt
     2881 taaatttttg ttaaatcagc tcatttttta accaataggc cgaaatcggc aaaatccctt
     2941 ataaatcaaa agaatagacc gagatagggt tgagtgttgt tccagtttgg aacaagagtc
     3001 cactattaaa gaacgtggac tccaacgtca aagggcgaaa aaccgtctat cagggcgatg
     3061 gcccactacg tgaaccatca ccctaatcaa gttttttggg gtcgaggtgc cgtaaagcac
     3121 taaatcggaa ccctaaaggg agcccccgat ttagagcttg acggggaaag ccggcgaacg
     3181 tggcgagaaa ggaagggaag aaagcgaaag gagcgggcgc tagggcgctg gcaagtgtag
     3241 cggtcacgct gcgcgtaacc accacacccg ccgcgcttaa tgcgccgcta cagggcgcgt
     3301 ccattcgcca ttcaggctgc gcaactgttg ggaagggcga tcggtgcggg cctcttcgct
     3361 attacgccag ctggcgaaag ggggatgtgc tgcaaggcga ttaagttggg taacgccagg
     3421 gttttcccag tcacgacgtt gtaaaacgac ggccagtgag cgcgcgtaat acgactcact
     3481 atagggcgaa ttgggtaccc attcataagt ttcagagcaa tgtccttatt ctggaacttg
     3541 gatttatggc tcttttggtt taatttcgcc tgattcttga tctcctttag cttctcgacg
     3601 tgggcctttt tcttgccata tggatccgct gcacggtcct gttccctagc atgtacgtga
     3661 gcgtatttcc ttttaaacca cgacgctttg tcttcattca acgtttccca ttgttttttt
     3721 ctactattgc tttgctgtgg gaaaaactta tcgaaagatg acgacttttt cttaattctc
     3781 gttttaagag cttggtgagc gctaggagtc actgccaggt atcgtttgaa cacggcatta
     3841 gtcagggaag tcataacaca gtcctttccc gcaattttct ttttctatta ctcttggcct
     3901 cctctagtac actctatatt tttttatgcc tcggtaatga ttttcatttt tttttttcca
     3961 cctagcggat gactcttttt ttttcttagc gattggcatt atcacataat gaattataca
     4021 ttatataaag taatgtgatt tcttcgaaga atatactaaa aaatgagcag gcaagataaa
     4081 cgaaggcaaa gaagcttaaa atggcaagag atccgcctgt ggccacgatg aaagataaca
     4141 cggttccctt aaaattgata gcccttcttg ccaatggtga gtttcattcc ggggaacaac
     4201 ttggcgaaac tctgggaatg agcagggcag cgattaacaa gcatatccaa accttgagag
     4261 actggggagt agatgtgttc accgtgcctg gtaaaggtta ttctttgccg gaacctatcc
     4321 ctctattaaa tgcgaaacag atactaggac aactggacgg tggtagtgtt gccgtattgc
     4381 cggttgttga ctcaaccaat caatacttgc tggacagaat tggcgaactt aaaagcggtg
     4441 atgcgtgtat tgcagaatat cagcaagcag ggagaggctc aagaggtagg aaatggtttt
     4501 caccttttgg cgccaatctt tacttaagta tgttctggcg tttgaagaga gggcccgcgg
     4561 ctattggttt aggtcccgtt atcgggatag ttatggctga agcgctacgt aaacttgggg
     4621 ccgacaaggt cagagtgaag tggcctaatg atctatactt gcaagataga aagctagcgg
     4681 gtatcctggt ggagctagcg ggaatcacag gggacgctgc ccaaatagta atcggtgctg
     4741 gtataaatgt cgcaatgcgt agagtcgagg aatcagtggt gaaccagggc tggatcacgt
     4801 tgcaagaggc gggcataaac ctggaccgta acacccttgc agcgacccta ataagagaac
     4861 tacgtgctgc attggagctt tttgagcagg agggcctggc tccgtaccta cccaggtggg
     4921 agaaattaga caatttcata aaccgtccag tcaaactaat aattggagat aaggagattt
     4981 tcggtatttc tcgtgggata gacaagcagg gagcgttgtt attagaacag gatggtgtca
     5041 ttaaaccttg gatgggcggt gaaattagtc ttcgtagtgc cgaaaaggct agcggttctg
     5101 gtggtgctag catcctctgg catgagatgt ggcatgaagg cctggaagag gcatctcgtt
     5161 tgtactttgg ggaaaggaac gtgaaaggca tgtttgaggt gctggagccc ttgcatgcta
     5221 tgatggaacg gggcccccag actctgaagg aaacatcctt taatcaggcc tatggtcgag
     5281 atttaatgga ggcccaagag tggtgcagga agtacatgaa atcagggaat gtcaaggacc
     5341 tcctccaagc ctgggacctc tattatcatg tgttccgacg aatctcaaag ctcgagtacc
     5401 catacgatgt tcctgactat gcgggctatc cgtatgacgt cccggactat gcaggatctt
     5461 atccatatga cgttccagat tacgctgctt gatctagagc ggccgccacc gcggtggagc
     5521 tccagctttt gttcccttta gtgagggtta attgcgcgct tggcgtaatc atggtcatag
     5581 ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc
     5641 ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc
     5701 tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa
     5761 cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg
     5821 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
     5881 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
     5941 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     6001 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     6061 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     6121 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     6181 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     6241 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     6301 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     6361 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
     6421 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     6481 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     6541 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     6601 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     6661 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     6721 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     6781 tttcgttcat ccatagttgc ctgactc
//
