LOCUS       sequence-341445-        7246 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid designed for the genomic tagging of saccharomyces cerevisiae
            Bem1 with mCherry-AID.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7246)
  AUTHORS   Daalman WK, Sweep E, Laan L
  TITLE     A tractable physical model for the yeast polarity predicts epistasis
            and fitness.
  JOURNAL   Philos Trans R Soc Lond B Biol Sci. 2023 May 22;378(1877):20220044. 
            doi: 10.1098/rstb.2022.0044. Epub 2023 Apr 3.
  PUBMED    37004720
REFERENCE   2  (bases 1 to 7246)
  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: "Philos 
            Trans R Soc Lond B Biol Sci."; date: "2023-05-22"; volume: 
            "378(1877):20220044. doi"; pages: " 10.1098/rstb.2022.0044. Epub 
            2023 Apr 3"
COMMENT     Sequence Label: Hygro-Bem1-mCherry-AID
FEATURES             Location/Qualifiers
     source          1..7246
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    88..109
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     CDS             887..1594
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(1032..1050)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1300..1319
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             1637..1768
                     /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"
     terminator      1816..2005
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     gene            complement(2153..3728)
                     /label=hphMX6
                     /note="yeast selectable marker conferring hygromycin 
                     resistance"
     terminator      complement(2153..2350)
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     CDS             complement(2356..3384)
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     CDS             complement(2356..3384)
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     promoter        complement(3385..3728)
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     promoter        complement(3841..3859)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(3842..3859)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     4882..4903
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5057..5076)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5176..5198
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5236..5254)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5322..5426
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5427..5495,5496..6287)
                     /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     complement(5645..5664)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6458..7046
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6947..6966
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7200..7217
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 gcctctcccc gcgcgttggc cgattcatta atgcagctgg cacgacaggt ttcccgactg
       61 gaaagcgggc agtgagcgca acgcaattaa tgtgagttag ctcactcatt aggcacccca
      121 ggctttacac tttatgcttc cgcggctcgt atgttgtgtg gaattaagag atttaaccat
      181 gacgccgcca tccaacggtc atggggcgct tagtgccaga aattcgagcc aaccttctac
      241 ggcgaataat ggaagtaatt cattcaagag ctcagccgtc ccacagactt cgcctggtca
      301 gccgagaaag ggccaaaacg gttctaatac tatagaagat atcaacaacc tagtgaatct
      361 tcccgattat atatctcgct cagaaatggt gcattcctta ttcgttgttt tgaataatgg
      421 tttcgacaga gaatttgaaa gagacgaaaa tcaaaacaat attaaaacat tacaagaaaa
      481 cgatacagca acctttgcaa cagcatctca aacttctaat tttgcctcaa ctaaccaaga
      541 caacacttta actggcgaag acttgaagtt aaacaagaaa ctctcagatt tatctttatc
      601 gggctccaag caggctccag cccaatcaac ttctggattg aaaactacta aaattaaatt
      661 ttattacaaa gacgatattt ttgctttaat gctgaagggt gacacaacct ataaagaact
      721 tcgtagtaaa atcgctccaa ggattgatac agataatttt aaattacaaa ccaaattatt
      781 tgatggtagc ggggaggaaa ttaagactga ttcacaagtc agtaacataa tccaagccaa
      841 actgaaaatt tccgttcacg atattagact cggtggcgga ggaagcatgg tgagcaaggg
      901 cgaggaggat aacatggcca tcatcaagga gttcatgcgc ttcaaggtgc acatggaggg
      961 ctccgtgaac ggccacgagt tcgagatcga gggcgagggc gagggccgcc cctacgaggg
     1021 cacccagacc gccaagctga aggtgaccaa gggtggcccc ctgcccttcg cctgggacat
     1081 cctgtcccct cagttcatgt acggctccaa ggcctacgtg aagcaccccg ccgacatccc
     1141 cgactacttg aagctgtcct tccccgaggg cttcaagtgg gagcgcgtga tgaacttcga
     1201 ggacggcggc gtggtgaccg tgacccagga ctcctccctg caggacggcg agttcatcta
     1261 caaggtgaag ctgcgcggca ccaacttccc ctccgacggc cccgtaatgc agaagaagac
     1321 catgggctgg gaggcctcct ccgagcggat gtaccccgag gacggcgccc tgaagggcga
     1381 gatcaagcag aggctgaagc tgaaggacgg cggccactac gacgctgagg tcaagaccac
     1441 ctacaaggcc aagaagcccg tgcagctgcc cggcgcctac aacgtcaaca tcaagttgga
     1501 catcacctcc cacaacgagg actacaccat cgtggaacag tacgaacgcg ccgagggccg
     1561 ccactccacc ggcggcatgg acgagctgta caaggtcgac ggtgcaggcg ccggggctgg
     1621 tgctggggca ggtaccccta aagatccagc caaacctccg gccaaggcac aagttgtggg
     1681 atggccaccg gtgagatcat accggaagaa cgtgatggtt tcctgccaaa aatcaagcgg
     1741 tggcccggag gcggcggcgt tcgtgaaggt atcacccggg taactcgagt aagcttggta
     1801 ccgcggctag ctaagatccg ctctaaccga aaaggaagga gttagacaac ctgaagtcta
     1861 ggtccctatt tattttttta tagttatgtt agtattaaga acgttattta tatttcaaat
     1921 ttttcttttt tttctgtaca gacgcgtgta cgcatgtaac attatactga aaaccttgct
     1981 tgagaaggtt ttgggacgct cgaagatcca gctcagcgta cgtcgacaat acgactcact
     2041 atagggagac cggcagatcc gcggccgcat aggccactag tggatctgat atcatcgatg
     2101 aattcgagct cgttttcgac actggatggc ggcgttagta tcgaatcgac agcagtatag
     2161 cgaccagcat tcacatacga ttgacgcatg atattacttt ctgcgcactt aacttcgcat
     2221 ctgggcagat gatgtcgagg cgaaaaaaaa tataaatcac gctaacattt gattaaaata
     2281 gaacaactac aatataaaaa aactatacaa atgacaagtt cttgaaaaca agaatctttt
     2341 tattgtcagt actgattatt cctttgccct cggacgagtg ctggggcgtc ggtttccact
     2401 atcggcgagt acttctacac agccatcggt ccagacggcc gcgcttctgc gggcgatttg
     2461 tgtacgcccg acagtcccgg ctccggatcg gacgattgcg tcgcatcgac cctgcgccca
     2521 agctgcatca tcgaaattgc cgtcaaccaa gctctgatag agttggtcaa gaccaatgcg
     2581 gagcatatac gcccggagcc gcggcgatcc tgcaagctcc ggatgcctcc gctcgaagta
     2641 gcgcgtctgc tgctccatac aagccaacca cggcctccag aagaagatgt tggcgacctc
     2701 gtattgggaa tccccgaaca tcgcctcgct ccagtcaatg accgctgtta tgcggccatt
     2761 gtccgtcagg acattgttgg agccgaaatc cgcgtgcacg aggtgccgga cttcggggca
     2821 gtcctcggcc caaagcatca gctcatcgag agcctgcgcg acggacgcac tgacggtgtc
     2881 gtccatcaca gtttgccagt gatacacatg gggatcagca atcgcgcata tgaaatcacg
     2941 ccatgtagtg tattgaccga ttccttgcgg tccgaatggg ccgaacccgc tcgtctggct
     3001 aagatcggcc gcagcgatcg catccatggc ctccgcgacc ggctgcagaa cagcgggcag
     3061 ttcggtttca ggcaggtctt gcaacgtgac accctgtgca cggcgggaga tgcaataggt
     3121 caggctctcg ctgaattccc caatgtcaag cacttccgga atcgggagcg cggccgatgc
     3181 aaagtgccga taaacataac gatctttgta gaaaccatcg gcgcagctat ttacccgcag
     3241 gacatatcca cgccctccta catcgaagct gaaagcacga gattcttcgc cctccgagag
     3301 ctgcatcagg tcggagacgc tgtcgaactt ttcgatcaga aacttctcga cagacgtcgc
     3361 ggtgagttca ggctttttac ccatggttgt ttatgttcgg atgtgatgtg agaactgtat
     3421 cctagcaaga ttttaaaagg aagtatatga aagaagaacc tcagtggcaa atcctaacct
     3481 tttatatttc tctacagggg cgcggcgtgg ggacaattca acgcgtctgt gaggggagcg
     3541 tttccctgct cgcaggtccg cagcgaggag ccgtaatttt tgcttcgcgc cgtgcggcca
     3601 tcaaaatgta tggatgcaaa tgattataca tggggatgta tgggctaaat gtacgggcga
     3661 cagtcacatc atgcccctga gctgcgcacg tcaagactgt caaggagggt attctgggcc
     3721 tccatgtcgc tggccgggtg acccggcggg gacgaggcaa gctaaacaga tctggcgcgc
     3781 cttaattaac ccggggatcc gtcgacctgc agcgtacgaa gcttcagctg gcggccgcgt
     3841 tctatagtgt cacctaaatc gtatgtgatc tagtgttaga agacgaagca tttttcttct
     3901 ttacttggca gatgcacata ggttgaacat tttaatcaac tttatgatga tttgcgagag
     3961 taataagatt ctcccgaaca gtaataatat taagatatcc aattcttcca aaacagcgtg
     4021 atttgtaata aaaataaata aggacatata tgtatataga aagttgctta aactttttag
     4081 accacttgga aatatttttt atttatattt ttcttcaaat gtaaagtttg gtacctaagc
     4141 ttaaattcac ctgaaaatga attatatgaa agattttaca ccacatgtta aaaaaaacgt
     4201 acaaaaagcc aaatatttcc actcaataat aatgcatgaa ttataggtat tttaagttat
     4261 attgctgacg taaaattcag aagagactac aagaaaaatt ctgaacatat ctattcttaa
     4321 ctatgctttt gtgcttccat atgtgtcaaa ggataatgtg gctccctggg gacaaggcaa
     4381 gctcagtgtc ccttctcgct gtcttttcga aagagtcata tgtataccgg caatagtaac
     4441 ttgcctgatt tatgcataat gccgttgtat ttgctagcca gttttccacc gctgtctgtg
     4501 ggttgcgacc atgctacctg gtacctagga ataacttcac cgtaaacaat attttctctt
     4561 gcaacgtcaa tcaatttggt tgcgataccc aacttccttg ctgtcctgca cacccaaatt
     4621 ctcgatatgc caatcttaaa atcggggtac acattctgta ccaatcttct agaatcataa
     4681 accatccaac gtccacgact agatgtttta ccattgcccc cataaaggtt ctctataatt
     4741 attattccga ccgccctgtc atttcttatg tatacaaacg ctttgccttt ttcttctgtg
     4801 gtactgttcc aaatgacatt ctcatcgtgt ggcgcattca gctcgttatt cactagtgtc
     4861 attatctccg tcatggtaat agtggactct tgttccaaac tggaacaaca ctcaactcta
     4921 tctcgggcta ttcttttgat ttataaggga ttttgccgat ttcggtctat tggttaaaaa
     4981 atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg tttacaattt
     5041 tatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag ccccgacacc
     5101 cgccaacacc cgctgacgcg ccctgacggg cttgtctgct cccggcatcc gcttacagac
     5161 aagctgtgac cgtctccggg agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac
     5221 gcgcgagacg aaagggcctc gtgatacgcc tatttttata ggttaatgtc atgataataa
     5281 tggtttctta gacgtcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt
     5341 tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc
     5401 ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc
     5461 ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa
     5521 aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg
     5581 gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag
     5641 ttctgctatg tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc
     5701 gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta
     5761 cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg
     5821 cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca
     5881 acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac
     5941 caaacgacga gcgtgacacc acgatgcctg tagcaatggc aacaacgttg cgcaaactat
     6001 taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg
     6061 ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata
     6121 aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta
     6181 agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa
     6241 atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag
     6301 tttactcata tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg
     6361 tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact
     6421 gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg
     6481 taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc
     6541 aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata
     6601 ctgttcttct agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta
     6661 catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc
     6721 ttaccgggtt ggactcaaga cgatagttac cggataaggc acagcggtcg ggctgaacgg
     6781 ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac
     6841 agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg
     6901 taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt
     6961 atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct
     7021 cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg
     7081 ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata
     7141 accgtattac cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca
     7201 gcgagtcagt gagcgaggaa gcggaagagc gcccaatacg caaacc
//
