LOCUS       sequence-199314-        5849 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid used for the C-terminal tagging of Cdc14 with mCherry and 
            the auxin-inducible degron in yeast..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5849)
  AUTHORS   Papagiannakis A, Niebel B, Wit EC, Heinemann M
  TITLE     Autonomous Metabolic Oscillations Robustly Gate the Early and Late 
            Cell Cycle.
  JOURNAL   Mol Cell. 2017 Jan 19;65(2):285-295. doi: 
            10.1016/j.molcel.2016.11.018. Epub 2016 Dec 15.
  PUBMED    27989441
REFERENCE   2  (bases 1 to 5849)
  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"; doi: 
            "10.1016/j.molcel.2016.11.018"; journalName: "Mol Cell"; date: 
            "2017-01-19- 19"; volume: "65"; issue: "2"; pages: "285-295"
COMMENT     Sequence Label: pG23A
FEATURES             Location/Qualifiers
     source          1..5849
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    1..22
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     CDS             572..1279
                     /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(717..735)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     985..1004
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             1322..1453
                     /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      1501..1690
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     gene            complement(1745..2864)
                     /label=natMX6
                     /note="yeast selectable marker conferring nourseothricin 
                     resistance"
     terminator      complement(1745..1942)
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     CDS             complement(1948..2520)
                     /codon_start=1
                     /gene="Streptomyces noursei nat1"
                     /product="nourseothricin acetyltransferase"
                     /label=NrsR
                     /note="confers resistance to nourseothricin"
                     /translation="MGTTLDDTAYRYRTSVPGDAEAIEALDGSFTTDTVFRVTATGDGF
                     TLREVPVDPPLTKVFPDDESDDESDDGEDGDPDSRTFVAYGDDGDLAGFVVVSYSGWNR
                     RLTVEDIEVAPEHRGHGVGRALMGLATEFARERGAGHLWLEVTNVNAPAIHAYRRMGFT
                     LCGLDTALYDGTASDGEQALYMSMPCP"
     promoter        complement(2521..2864)
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     primer_bind     3398..3419
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3573..3592)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3692..3714
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3752..3770)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3838..3942
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3943..4011,4012..4803)
                     /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(4161..4180)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4974..5562
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5463..5482
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5716..5733
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 taatgtgagt tagctcactc attaggcacc ccaggcttta cactttatgc ttccgcggct
       61 cgtatgttgt gtggaattaa gagatttaac catgacgccg ccatccaacg gtcatggggc
      121 gcttagtgcc agaaattcga gccaaccttc tacggcgaat aatggaagta attcattcaa
      181 gagctcagcc gtcccacaga cttcgcctgg tcagccgaga aagggccaaa acggttctaa
      241 tactatagaa gatatcaaca ataatcgtaa tccaacttcc catgcgaata ggaaggtagt
      301 aatagaaagc aacaactcag atgacgaatc catgcaggat accaacggca caagtaatca
      361 ttatcccaaa gtttcgcgca agaaaaatga tatctcctct gcttcatcgt cccgaatgga
      421 agataacgaa cccagcgcaa cgaacatcaa taatgccgca gatgatacca tactaagaca
      481 actattgccc aagaatagga gagtaacgtc agggagaagg actacaagcg ccgccggtgg
      541 tataagaaaa ataagtggct ccatcaagaa aatggtgagc aagggcgagg aggataacat
      601 ggccatcatc aaggagttca tgcgcttcaa ggtgcacatg gagggctccg tgaacggcca
      661 cgagttcgag atcgagggcg agggcgaggg ccgcccctac gagggcaccc agaccgccaa
      721 gctgaaggtg accaagggtg gccccctgcc cttcgcctgg gacatcctgt cccctcagtt
      781 catgtacggc tccaaggcct acgtgaagca ccccgccgac atccccgact acttgaagct
      841 gtccttcccc gagggcttca agtgggagcg cgtgatgaac ttcgaggacg gcggcgtggt
      901 gaccgtgacc caggactcct ccctgcagga cggcgagttc atctacaagg tgaagctgcg
      961 cggcaccaac ttcccctccg acggccccgt aatgcagaag aagaccatgg gctgggaggc
     1021 ctcctccgag cggatgtacc ccgaggacgg cgccctgaag ggcgagatca agcagaggct
     1081 gaagctgaag gacggcggcc actacgacgc tgaggtcaag accacctaca aggccaagaa
     1141 gcccgtgcag ctgcccggcg cctacaacgt caacatcaag ttggacatca cctcccacaa
     1201 cgaggactac accatcgtgg aacagtacga acgcgccgag ggccgccact ccaccggcgg
     1261 catggacgag ctgtacaagg tcgacggtgc aggcgccggg gctggtgctg gggcaggtac
     1321 ccctaaagat ccagccaaac ctccggccaa ggcacaagtt gtgggatggc caccggtgag
     1381 atcataccgg aagaacgtga tggtttcctg ccaaaaatca agcggtggcc cggaggcggc
     1441 ggcgttcgtg aaggtatcac ccgggtaact cgagtaagct tggtaccgcg gctagctaag
     1501 atccgctcta accgaaaagg aaggagttag acaacctgaa gtctaggtcc ctatttattt
     1561 ttttatagtt atgttagtat taagaacgtt atttatattt caaatttttc ttttttttct
     1621 gtacagacgc gtgtacgcat gtaacattat actgaaaacc ttgcttgaga aggttttggg
     1681 acgctcgaag atccagctca gcgtacgtcg acactggatg gcggcgttag tatcgaatcg
     1741 acagcagtat agcgaccagc attcacatac gattgacgca tgatattact ttctgcgcac
     1801 ttaacttcgc atctgggcag atgatgtcga ggcgaaaaaa aatataaatc acgctaacat
     1861 ttgattaaaa tagaacaact acaatataaa aaaactatac aaatgacaag ttcttgaaaa
     1921 caagaatctt tttattgtca gtactgatta ggggcagggc atgctcatgt agagcgcctg
     1981 ctcgccgtcc gaggcggtgc cgtcgtacag ggcggtgtcc aggccgcaga gggtgaaccc
     2041 catccgccgg tacgcgtgga tcgccggtgc gttgacgttg gtgacctcca gccagaggtg
     2101 cccggcgccc cgctcgcggg cgaactccgt cgcgagcccc atcaacgcgc gcccgacccc
     2161 gtgcccccgg tgctccgggg cgacctcgat gtcctcgacg gtcagccggc ggttccagcc
     2221 ggagtacgag acgaccacga agcccgccag gtcgccgtcg tccccgtacg cgacgaacgt
     2281 ccgggagtcc gggtcgccgt cctccccgtc gtccgattcg tcgtccgatt cgtcgtcggg
     2341 gaacaccttg gtcaggggcg ggtccaccgg cacctcccgc agggtgaagc cgtccccggt
     2401 ggcggtgacg cggaagacgg tgtcggtggt gaaggaccca tccagtgcct cgatggcctc
     2461 ggcgtccccc gggacactgg tgcggtaccg gtaagccgtg tcgtcaagag tggtacccat
     2521 ggttgtttat gttcggatgt gatgtgagaa ctgtatccta gcaagatttt aaaaggaagt
     2581 atatgaaaga agaacctcag tggcaaatcc taacctttta tatttctcta caggggcgcg
     2641 gcgtggggac aattcaacgc gtctgtgagg ggagcgtttc cctgctcgca ggtctgcagc
     2701 gaggagccgt aatttttgct tcgcgccgtg cggccatcaa aatgtatgga tgcaaatgat
     2761 tatacatggg gatgtatggg ctaaatgtac gggcgacagt cacatcatgc ccctgagctg
     2821 cgcacgtcaa gactgtcaag gagggtattc tgggcctcca tgtcgctggc cgggtgaccc
     2881 ggcggggaca aggcaagctc agtgtccctt ctcgctgtct tttcgaaaga gtcatatgta
     2941 taccggcaat agtaacttgc ctgatttatg cataatgccg ttgtatttgc tagccagttt
     3001 tccaccgctg tctgtgggtt gcgaccatgc tacctggtac ctaggaataa cttcaccgta
     3061 aacaatattt tctcttgcaa cgtcaatcaa tttggttgcg atacccaact tccttgctgt
     3121 cctgcacacc caaattctcg atatgccaat cttaaaatcg gggtacacat tctgtaccaa
     3181 tcttctagaa tcataaacca tccaacgtcc acgactagat gttttaccat tgcccccata
     3241 aaggttctct ataattatta ttccgaccgc cctgtcattt cttatgtata caaacgcttt
     3301 gcctttttct tctgtggtac tgttccaaat gacattctca tcgtgtggcg cattcagctc
     3361 gttattcact agtgtcatta tctccgtcat ggtaatagtg gactcttgtt ccaaactgga
     3421 acaacactca actctatctc gggctattct tttgatttat aagggatttt gccgatttcg
     3481 gtctattggt taaaaaatga gctgatttaa caaaaattta acgcgaattt taacaaaata
     3541 ttaacgttta caattttatg gtgcactctc agtacaatct gctctgatgc cgcatagtta
     3601 agccagcccc gacacccgcc aacacccgct gacgcgccct gacgggcttg tctgctcccg
     3661 gcatccgctt acagacaagc tgtgaccgtc tccgggagct gcatgtgtca gaggttttca
     3721 ccgtcatcac cgaaacgcgc gagacgaaag ggcctcgtga tacgcctatt tttataggtt
     3781 aatgtcatga taataatggt ttcttagacg tcaggtggca cttttcgggg aaatgtgcgc
     3841 ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa
     3901 taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc
     3961 cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa
     4021 acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa
     4081 ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg
     4141 atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa
     4201 gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc
     4261 acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc
     4321 atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta
     4381 accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag
     4441 ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca
     4501 acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca acaattaata
     4561 gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc
     4621 tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca
     4681 ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca
     4741 actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg
     4801 taactgtcag accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa
     4861 tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt
     4921 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat
     4981 cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg
     5041 gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga
     5101 gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac
     5161 tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt
     5221 ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcacag
     5281 cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc
     5341 gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag
     5401 gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca
     5461 gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt
     5521 cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc
     5581 tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc
     5641 cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc
     5701 cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg aagagcgccc aatacgcaaa
     5761 ccgcctctcc ccgcgcgttg gccgattcat taatgcagct ggcacgacag gtttcccgac
     5821 tggaaagcgg gcagtgagcg caacgcaat
//
