LOCUS       sequence-415809-        4933 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid with KanMX-tetO7-CYCpr cassette from the yTHC collection. 
            Used for amplifying cassette with homology to regions of interest 
            for genomic integration..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4933)
  AUTHORS   Evans-Yamamoto D, Dube AK, Saha G, Plante S, Bradley D, 
            Gagnon-Arsenault I, Landry CR
  TITLE     Parallel nonfunctionalization of CK1delta/epsilon kinase ohnologs 
            following a whole-genome duplication event.
  JOURNAL   Mol Biol Evol. 2023 Nov 18:msad246. doi: 10.1093/molbev/msad246.
  PUBMED    37979156
REFERENCE   2  (bases 1 to 4933)
  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: "Mol Biol 
            Evol. 2023 Nov 18:msad246. doi: 10.1093/molbev/msad246."
COMMENT     Sequence Label: pKB33 (pUC19-KANMX-tetO7-CYCpr)
FEATURES             Location/Qualifiers
     source          1..4933
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     gene            complement(14..1370)
                     /label=kanMX
                     /note="yeast selectable marker conferring kanamycin 
                     resistance (Wach et al., 1994)"
     terminator      complement(14..211)
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     CDS             complement(217..1026)
                     /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     940..959
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(1027..1370)
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     terminator      1473..1660
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(1528..1552)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     protein_bind    join(1718..1736,1737..1759,1760..1778,1779..1801,1802..1820,
                     1821..1843,1844..1862,1863..1885,1886..1904,1905..1927,
                     1928..1946,1947..1969,1970..1988)
                     /label=tetracycline response element
                     /bound_moiety="tetracycline repressor TetR"
                     /note="contains seven copies of the tetracycline operator 
                     tetO"
     primer_bind     complement(2194..2211)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2194..2210)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2203..2225)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(2419..2438)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2538..2560
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2598..2616)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2684..2788
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2789..2857,2858..3649)
                     /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(3007..3026)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3820..4408
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4309..4328
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4562..4579
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    4696..4717
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4732..4737,4738..4755,4756..4762)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4770..4786
                     /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)."
     primer_bind     4794..4810
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4872..4888
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 atcgaatcga cagcagtata gcgaccagca ttcacatacg attgacgcat gatattactt
       61 tctgcgcact taacttcgca tctgggcaga tgatgtcgag gcgaaaaaaa atataaatca
      121 cgctaacatt tgattaaaat agaacaacta caatataaaa aaactataca aatgacaagt
      181 tcttgaaaac aagaatcttt ttattgtcag tactgattag aaaaactcat cgagcatcaa
      241 atgaaactgc aatttattca tatcaggatt atcaatacca tatttttgaa aaagccgttt
      301 ctgtaatgaa ggagaaaact caccgaggca gttccatagg atggcaagat cctggtatcg
      361 gtctgcgatt ccgactcgtc caacatcaat acaacctatt aatttcccct cgtcaaaaat
      421 aaggttatca agtgagaaat caccatgagt gacgactgaa tccggtgaga atggcaaaag
      481 cttatgcatt tctttccaga cttgttcaac aggccagcca ttacgctcgt catcaaaatc
      541 actcgcatca accaaaccgt tattcattcg tgattgcgcc tgagcgagac gaaatacgcg
      601 atcgctgtta aaaggacaat tacaaacagg aatcgaatgc aaccggcgca ggaacactgc
      661 cagcgcatca acaatatttt cacctgaatc aggatattct tctaatacct ggaatgctgt
      721 tttgccgggg atcgcagtgg tgagtaacca tgcatcatca ggagtacgga taaaatgctt
      781 gatggtcgga agaggcataa attccgtcag ccagtttagt ctgaccatct catctgtaac
      841 atcattggca acgctacctt tgccatgttt cagaaacaac tctggcgcat cgggcttccc
      901 atacaatcga tagattgtcg cacctgattg cccgacatta tcgcgagccc atttataccc
      961 atataaatca gcatccatgt tggaatttaa tcgcggcctc gaaacgtgag tcttttcctt
     1021 acccatggtt gtttatgttc ggatgtgatg tgagaactgt atcctagcaa gattttaaaa
     1081 ggaagtatat gaaagaagaa cctcagtggc aaatcctaac cttttatatt tctctacagg
     1141 ggcgcggcgt ggggacaatt caacgcgtct gtgaggggag cgtttccctg ctcgcaggtc
     1201 tgcagcgagg agccgtaatt tttgcttcgc gccgtgcggc catcaaaatg tatggatgca
     1261 aatgattata catggggatg tatgggctaa atgtacgggc gacagtcaca tcatgcccct
     1321 gagctgcgca cgtcaagact gtcaaggagg gtattctggg cctccatgtc gctggccggg
     1381 tgacccggcg gggacgaggc aagctaaaca gatctggcgc gccttaatta acccggggat
     1441 caactcgagt aattcgcgcc acttctaaat aagcgaattt cttatgattt atgattttta
     1501 ttattaaata agttataaaa aaaataagtg tatacaaatt ttaaagtgac tcttgggttt
     1561 taaaacgaaa attcttattc ttgagtaact ctttcctgta ggtcaggttg ctttctcagg
     1621 tatagtatga ggtcgctctt attgaccaca cctctaccgg cagatccgct agggataaca
     1681 gggtgatata gatcaattcc tcgatcgagt ttaccactcc ctatcagtga tagagaaaag
     1741 tgaaagtcga gtttaccact ccctatcagt gatagagaaa agtggaagtc gagtttacca
     1801 ctccctatca gtgatagaga aaagtgaaag tcgagtttac cactccctat cagtgataga
     1861 gaaaagtgaa agtcgagttt accactccct atcagtgata gagaaaagtg aaagtcgagt
     1921 ttaccactcc ctatcagtga tagagaaaag tgaaagtcga gtttaccact ccctatcagt
     1981 gatagagaaa agtgaaagtc gagctcggta ccctatggca tgcatgtgct ctgtatgtat
     2041 ataaaactct tgttttcttc ttttctctaa atattctttc cttatacatt aggtcctttg
     2101 tagcataaat tactatactt ctatagacac gcaaacacaa atacacacac taaattaccg
     2161 gatcaattcg ggggatccat ggagctcgaa ttcactggcc gtcgttttac aacgtcgtga
     2221 ctgggaaaac cctggcgtta cccaacttaa tcgccttgca gcacatcccc ctttcgccag
     2281 ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc caacagttgc gcagcctgaa
     2341 tggcgaatgg cgcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg
     2401 catatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc agccccgaca
     2461 cccgccaaca cccgctgacg cgccctgacg ggcttgtctg ctcccggcat ccgcttacag
     2521 acaagctgtg accgtctccg ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa
     2581 acgcgcgaga cgaaagggcc tcgtgatacg cctattttta taggttaatg tcatgataat
     2641 aatggtttct tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg
     2701 tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat
     2761 gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat
     2821 tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt
     2881 aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag
     2941 cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa
     3001 agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg
     3061 ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct
     3121 tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac
     3181 tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca
     3241 caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat
     3301 accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact
     3361 attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc
     3421 ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga
     3481 taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg
     3541 taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg
     3601 aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca
     3661 agtttactca tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta
     3721 ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca
     3781 ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg
     3841 cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga
     3901 tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa
     3961 tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc
     4021 tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg
     4081 tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac
     4141 ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct
     4201 acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc
     4261 ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg
     4321 gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg
     4381 ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct
     4441 ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga
     4501 taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg
     4561 cagcgagtca gtgagcgagg aagcggaaga gcgcccaata cgcaaaccgc ctctccccgc
     4621 gcgttggccg attcattaat gcagctggca cgacaggttt cccgactgga aagcgggcag
     4681 tgagcgcaac gcaattaatg tgagttagct cactcattag gcaccccagg ctttacactt
     4741 tatgcttccg gctcgtatgt tgtgtggaat tgtgagcgga taacaatttc acacaggaaa
     4801 cagctatgac catgattacg ccaagcttgc atgcctgcag gtcgactcta gaagcggata
     4861 acaatttcac acaggaaaca gctatgacat gattacgaat tcgagctcgt ttaaactgga
     4921 tggcggcgtt agt
//
