LOCUS       sequence-337356-        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"
     CDS             578..1285
                     /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(723..741)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     991..1010
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             1328..1459
                     /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      1507..1696
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     gene            complement(1844..3419)
                     /label=hphMX6
                     /note="yeast selectable marker conferring hygromycin 
                     resistance"
     terminator      complement(1844..2041)
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     CDS             complement(2047..3075)
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     CDS             complement(2047..3075)
                     /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(3076..3419)
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     promoter        complement(3532..3550)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(3533..3550)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     4573..4594
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4748..4767)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4867..4889
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4927..4945)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5013..5117
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5118..5186,5187..5978)
                     /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(5336..5355)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6149..6737
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6638..6657
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6891..6908
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7025..7046
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
ORIGIN
        1 gggccaaaac ggttctaata ctatagaaga tatcaacaac ctagtgaatc ttcccgatta
       61 tatatctcgc tcagaaatgg tgcattcctt attcgttgtt ttgaataatg gtttcgacag
      121 agaatttgaa agagacgaaa atcaaaacaa tattaaaaca ttacaagaaa acgatacagc
      181 aacctttgca acagcatctc aaacttctaa ttttgcctca actaaccaag acaacacttt
      241 aactggcgaa gacttgaagt taaacaagaa actctcagat ttatctttat cgggctccaa
      301 gcaggctcca gcccaatcaa cttctggatt gaaaactact aaaattaaat tttattacaa
      361 agacgatatt tttgctttaa tgctgaaggg tgacacaacc tataaagaac ttcgtagtaa
      421 aatcgctcca aggattgata cagataattt taaattacaa accaaattat ttgatggtag
      481 cggggaggaa attaagactg attcacaagt cagtaacata atccaagcca aactgaaaat
      541 ttccgttcac gatattagac tcggtggcgg aggaagcatg gtgagcaagg gcgaggagga
      601 taacatggcc atcatcaagg agttcatgcg cttcaaggtg cacatggagg gctccgtgaa
      661 cggccacgag ttcgagatcg agggcgaggg cgagggccgc ccctacgagg gcacccagac
      721 cgccaagctg aaggtgacca agggtggccc cctgcccttc gcctgggaca tcctgtcccc
      781 tcagttcatg tacggctcca aggcctacgt gaagcacccc gccgacatcc ccgactactt
      841 gaagctgtcc ttccccgagg gcttcaagtg ggagcgcgtg atgaacttcg aggacggcgg
      901 cgtggtgacc gtgacccagg actcctccct gcaggacggc gagttcatct acaaggtgaa
      961 gctgcgcggc accaacttcc cctccgacgg ccccgtaatg cagaagaaga ccatgggctg
     1021 ggaggcctcc tccgagcgga tgtaccccga ggacggcgcc ctgaagggcg agatcaagca
     1081 gaggctgaag ctgaaggacg gcggccacta cgacgctgag gtcaagacca cctacaaggc
     1141 caagaagccc gtgcagctgc ccggcgccta caacgtcaac atcaagttgg acatcacctc
     1201 ccacaacgag gactacacca tcgtggaaca gtacgaacgc gccgagggcc gccactccac
     1261 cggcggcatg gacgagctgt acaaggtcga cggtgcaggc gccggggctg gtgctggggc
     1321 aggtacccct aaagatccag ccaaacctcc ggccaaggca caagttgtgg gatggccacc
     1381 ggtgagatca taccggaaga acgtgatggt ttcctgccaa aaatcaagcg gtggcccgga
     1441 ggcggcggcg ttcgtgaagg tatcacccgg gtaactcgag taagcttggt accgcggcta
     1501 gctaagatcc gctctaaccg aaaaggaagg agttagacaa cctgaagtct aggtccctat
     1561 ttattttttt atagttatgt tagtattaag aacgttattt atatttcaaa tttttctttt
     1621 ttttctgtac agacgcgtgt acgcatgtaa cattatactg aaaaccttgc ttgagaaggt
     1681 tttgggacgc tcgaagatcc agctcagcgt acgtcgacaa tacgactcac tatagggaga
     1741 ccggcagatc cgcggccgca taggccacta gtggatctga tatcatcgat gaattcgagc
     1801 tcgttttcga cactggatgg cggcgttagt atcgaatcga cagcagtata gcgaccagca
     1861 ttcacatacg attgacgcat gatattactt tctgcgcact taacttcgca tctgggcaga
     1921 tgatgtcgag gcgaaaaaaa atataaatca cgctaacatt tgattaaaat agaacaacta
     1981 caatataaaa aaactataca aatgacaagt tcttgaaaac aagaatcttt ttattgtcag
     2041 tactgattat tcctttgccc tcggacgagt gctggggcgt cggtttccac tatcggcgag
     2101 tacttctaca cagccatcgg tccagacggc cgcgcttctg cgggcgattt gtgtacgccc
     2161 gacagtcccg gctccggatc ggacgattgc gtcgcatcga ccctgcgccc aagctgcatc
     2221 atcgaaattg ccgtcaacca agctctgata gagttggtca agaccaatgc ggagcatata
     2281 cgcccggagc cgcggcgatc ctgcaagctc cggatgcctc cgctcgaagt agcgcgtctg
     2341 ctgctccata caagccaacc acggcctcca gaagaagatg ttggcgacct cgtattggga
     2401 atccccgaac atcgcctcgc tccagtcaat gaccgctgtt atgcggccat tgtccgtcag
     2461 gacattgttg gagccgaaat ccgcgtgcac gaggtgccgg acttcggggc agtcctcggc
     2521 ccaaagcatc agctcatcga gagcctgcgc gacggacgca ctgacggtgt cgtccatcac
     2581 agtttgccag tgatacacat ggggatcagc aatcgcgcat atgaaatcac gccatgtagt
     2641 gtattgaccg attccttgcg gtccgaatgg gccgaacccg ctcgtctggc taagatcggc
     2701 cgcagcgatc gcatccatgg cctccgcgac cggctgcaga acagcgggca gttcggtttc
     2761 aggcaggtct tgcaacgtga caccctgtgc acggcgggag atgcaatagg tcaggctctc
     2821 gctgaattcc ccaatgtcaa gcacttccgg aatcgggagc gcggccgatg caaagtgccg
     2881 ataaacataa cgatctttgt agaaaccatc ggcgcagcta tttacccgca ggacatatcc
     2941 acgccctcct acatcgaagc tgaaagcacg agattcttcg ccctccgaga gctgcatcag
     3001 gtcggagacg ctgtcgaact tttcgatcag aaacttctcg acagacgtcg cggtgagttc
     3061 aggcttttta cccatggttg tttatgttcg gatgtgatgt gagaactgta tcctagcaag
     3121 attttaaaag gaagtatatg aaagaagaac ctcagtggca aatcctaacc ttttatattt
     3181 ctctacaggg gcgcggcgtg gggacaattc aacgcgtctg tgaggggagc gtttccctgc
     3241 tcgcaggtct gcagcgagga gccgtaattt ttgcttcgcg ccgtgcggcc atcaaaatgt
     3301 atggatgcaa atgattatac atggggatgt atgggctaaa tgtacgggcg acagtcacat
     3361 catgcccctg agctgcgcac gtcaagactg tcaaggaggg tattctgggc ctccatgtcg
     3421 ctggccgggt gacccggcgg ggacgaggca agctaaacag atctggcgcg ccttaattaa
     3481 cccggggatc cgtcgacctg cagcgtacga agcttcagct ggcggccgcg ttctatagtg
     3541 tcacctaaat cgtatgtgat ctagtgttag aagacgaagc atttttcttc tttacttggc
     3601 agatgcacat aggttgaaca ttttaatcaa ctttatgatg atttgcgaga gtaataagat
     3661 tctcccgaac agtaataata ttaaggtatc caattcttcc gaaacagcgt gatttgtaat
     3721 aaaaataaat aaggacatat atgtatatag aaagttgctt aaacttttta gaccacttgg
     3781 aaatattttt tatttatatt tttcttcaaa tgtaaagttg ggtacctaag cttaaattca
     3841 cctgaaaatg aattatatga aagattttac accacatgta aaaaaaaacg tacaaaaagc
     3901 caaatatttc cactcaatca taatgcatga attataggta ttttaagtta tattgctgac
     3961 gtaaaattca gaagagacta caagaaaaat tctgaacata tttattctta aatatgcttt
     4021 tgtgcttcca tatgtgtcaa aggataatgt ggctccctgg ggacaaggca agctcagtgt
     4081 cccttctcgc tgtcttttcg aaagagtcat atgtataccg gcaatagtaa cttgcctgat
     4141 ttatgcataa tgccgttgta tttgctagcc agttttccac cgctgtctgt gggttgcgac
     4201 catgctacct ggtacctagg aataacttca ccgtaaacaa tattttctct tgcaacgtca
     4261 atcaatttgg ttgcgatacc caacttcctt gctgtcctgc acacccaaat tctcgatatg
     4321 ccaatcttaa aatcggggta cacattctgt accaatcttc tagaatcata aaccatccaa
     4381 cgtccacgac tagatgtttt accattgccc ccataaaggt tctctataat tattattccg
     4441 accgccctgt catttcttat gtatacaaac gctttgcctt tttcttctgt ggtactgttc
     4501 caaatgacat tctcatcgtg tggcgcattc agctcgttat tcactagtgt cattatctcc
     4561 gtcatggtaa tagtggactc ttgttccaaa ctggaacaac actcaactct atctcgggct
     4621 attcttttga tttataaggg attttgccga tttcggtcta ttggttaaaa aatgagctga
     4681 tttaacaaaa atttaacgcg aattttaaca aaatattaac gtttacaatt ttatggtgca
     4741 ctctcagtac aatctgctct gatgccgcat agttaagcca gccccgacac ccgccaacac
     4801 ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga caagctgtga
     4861 ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgagac
     4921 gaaagggcct cgtgatacgc ctatttttat aggttaatgt catgataata atggtttctt
     4981 agacgtcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct
     5041 aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat
     5101 attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt cccttttttg
     5161 cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg
     5221 aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc
     5281 ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa gttctgctat
     5341 gtggcgcggt attatcccgt attgacgccg ggcaagagca actcggtcgc cgcatacact
     5401 attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt acggatggca
     5461 tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact
     5521 tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg
     5581 atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg
     5641 agcgtgacac cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg
     5701 aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg gataaagttg
     5761 caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag
     5821 ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc
     5881 gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga
     5941 tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa gtttactcat
     6001 atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc
     6061 tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag
     6121 accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct
     6181 gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac
     6241 caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc
     6301 tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg
     6361 ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt
     6421 tggactcaag acgatagtta ccggataagg cacagcggtc gggctgaacg gggggttcgt
     6481 gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc
     6541 tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca
     6601 gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata
     6661 gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg
     6721 ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct
     6781 ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta
     6841 ccgcctttga gtgagctgat accgctcgcc gcagccgaac gaccgagcgc agcgagtcag
     6901 tgagcgagga agcggaagag cgcccaatac gcaaaccgcc tctccccgcg cgttggccga
     6961 ttcattaatg cagctggcac gacaggtttc ccgactggaa agcgggcagt gagcgcaacg
     7021 caattaatgt gagttagctc actcattagg caccccaggc tttacacttt atgcttccgc
     7081 ggctcgtatg ttgtgtggaa ttaagagatt taaccatgac gccgccatcc aacggtcatg
     7141 gggcgcttag tgccagaaat tcgagccaac cttctacggc gaataatgga agtaattcat
     7201 tcaagagctc agccgtccca cagacttcgc ctggtcagcc gagaaa
//
