LOCUS       sequence-166726-        4525 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  N-ICE plasmid pK3F-GAL1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4525)
  AUTHORS   Zhang Y, Serratore ND, Briggs SD
  TITLE     N-ICE plasmids for generating N-terminal 3 x FLAG tagged genes that 
            allow Inducible, Constitutive, or Endogenous expression in 
            Saccharomyces cerevisiae.
  JOURNAL   Yeast. 2016 Dec 12. doi: 10.1002/yea.3226.
  PUBMED    27943405
REFERENCE   2  (bases 1 to 4525)
  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: "Yeast. 2016
            Dec 12. doi: 10.1002/yea.3226."
COMMENT     Sequence Label: pK3F-GAL1
FEATURES             Location/Qualifiers
     source          1..4525
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    complement(67..100)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     gene            142..1498
                     /label=kanMX
                     /note="yeast selectable marker conferring kanamycin 
                     resistance (Wach et al., 1994)"
     promoter        142..485
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             486..1295
                     /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     complement(553..572)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      1301..1498
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     promoter        1550..1991
                     /gene="S. cerevisiae GAL1"
                     /label=GAL1 promoter
                     /note="inducible promoter, regulated by Gal4"
     protein_bind    1550..1667
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     primer_bind     1962..1985
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     protein_bind    complement(2009..2042)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             2065..2136
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags"
                     /label=3xFLAG
                     /translation="DYKDDDDKDYKDDDDKDYKDDDDK"
     promoter        complement(2174..2192)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2175..2192)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2273..2292)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2392..2414
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2452..2470)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2538..2642
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2643..2711,2712..3503)
                     /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(2861..2880)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3674..4262
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4163..4182
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4416..4433
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     join(4520..4525,1..14)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        join(4520..4525,1..13)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 gactcactat agggagaccg gcagatccgc ggccgcatag gccactagtg gatctgatat
       61 cacctaataa cttcgtatag catacattat acgaagttat attaagggtt ctcgacgtcc
      121 ccgccgggtc acccggccag cgacatggag gcccagaata ccctccttga cagtcttgac
      181 gtgcgcagct caggggcatg atgtgactgt cgcccgtaca tttagcccat acatccccat
      241 gtataatcat ttgcatccat acattttgat ggccgcacgg cgcgaagcaa aaattacggc
      301 tcctcgctgc agacctgcga gcagggaaac gctcccctca cagacgcgtt gaattgtccc
      361 cacgccgcgc ccctgtagag aaatataaaa ggttaggatt tgccactgag gttcttcttt
      421 catatacttc cttttaaaat cttgctagga tacagttctc acatcacatc cgaacataaa
      481 caaccatggg taaggaaaag actcacgttt cgaggccgcg attaaattcc aacatggatg
      541 ctgatttata tgggtataaa tgggctcgcg ataatgtcgg gcaatcaggt gcgacaatct
      601 atcgattgta tgggaagccc gatgcgccag agttgtttct gaaacatggc aaaggtagcg
      661 ttgccaatga tgttacagat gagatggtca gactaaactg gctgacggaa tttatgcctc
      721 ttccgaccat caagcatttt atccgtactc ctgatgatgc atggttactc accactgcga
      781 tccccggcaa aacagcattc caggtattag aagaatatcc tgattcaggt gaaaatattg
      841 ttgatgcgct ggcagtgttc ctgcgccggt tgcattcgat tcctgtttgt aattgtcctt
      901 ttaacagcga tcgcgtattt cgtctcgctc aggcgcaatc acgaatgaat aacggtttgg
      961 ttgatgcgag tgattttgat gacgagcgta atggctggcc tgttgaacaa gtctggaaag
     1021 aaatgcataa gcttttgcca ttctcaccgg attcagtcgt cactcatggt gatttctcac
     1081 ttgataacct tatttttgac gaggggaaat taataggttg tattgatgtt ggacgagtcg
     1141 gaatcgcaga ccgataccag gatcttgcca tcctatggaa ctgcctcggt gagttttctc
     1201 cttcattaca gaaacggctt tttcaaaaat atggtattga taatcctgat atgaataaat
     1261 tgcagtttca tttgatgctc gatgagtttt tctaatcagt actgacaata aaaagattct
     1321 tgttttcaag aacttgtcat ttgtatagtt tttttatatt gtagttgttc tattttaatc
     1381 aaatgttagc gtgatttata ttttttttcg cctcgacatc atctgcccag atgcgaagtt
     1441 aagtgcgcag aaagtaatat catgcgtcaa tcgtatgtga atgctggtcg ctatactgct
     1501 gtcgattcga tactaacgcc gccatccagt gtcgagatct ctagtagtac ggattagaag
     1561 ccgccgagcg ggcgacagcc ctccgacgga agactctcct ccgtgcgtcc tcgtcttcac
     1621 cggtcgcgtt cctgaaacgc agatgtgcct cgcgccgcac tgctccgaac aataaagatt
     1681 ctacaatact agcttttatg gttatgaaga ggaaaaattg gcagtaacct ggccccacaa
     1741 accttcaaat taacgaatca aattaacaac cataggatga taatgcgatt agttttttag
     1801 ccttatttct ggggtaatta atcagcgaag cgatgatttt tgatctatta acagatatat
     1861 aaatggaaaa gctgcataac cactttaact aatactttca acattttcag tttgtattac
     1921 ttcttattca aatgtcataa aagtatcaac aaaaaattgt taatatacct ctatacttta
     1981 acgtcaagga gaaaaaactc tagacctaat aacttcgtat agcatacatt atacgaagtt
     2041 atattaaggg ttgtcgacac catggactat aaagacgacg acgacaaaga ttataaggac
     2101 gacgacgata aggattacaa ggacgacgac gataaaggat ccatgggcgg ccgcgagatc
     2161 ctggcggccg cgttctatag tgtcacctaa atcgtatgtg tatgatacat aaggttatgt
     2221 attaattgta gccgcgttct aacgacaata tgtccatatg gtgcactctc agtacaatct
     2281 gctctgatgc cgcatagtta agccagcccc gacacccgcc aacacccgct gacgcgccct
     2341 gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct
     2401 gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gagacgaaag ggcctcgtga
     2461 tacgcctatt tttataggtt aatgtcatga taataatggt ttcttagacg tcaggtggca
     2521 cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata
     2581 tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga
     2641 gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc
     2701 ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg
     2761 cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc
     2821 ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat
     2881 cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact
     2941 tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat
     3001 tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga
     3061 tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc
     3121 ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga
     3181 tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag
     3241 cttcccggca acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc
     3301 gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt
     3361 ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct
     3421 acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg
     3481 cctcactgat taagcattgg taactgtcag accaagttta ctcatatata ctttagattg
     3541 atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca
     3601 tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga
     3661 tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa
     3721 aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga
     3781 aggtaactgg cttcagcaga gcgcagatac caaatactgt ccttctagtg tagccgtagt
     3841 taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt
     3901 taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat
     3961 agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct
     4021 tggagcgaac gacctacacc gaactgagat acctacagcg tgagcattga gaaagcgcca
     4081 cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag
     4141 agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc
     4201 gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga
     4261 aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca
     4321 tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc tttgagtgag
     4381 ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg
     4441 aagagcgccc aatacgcaaa ccgcctctcc ccgcgcgttg gccgattcat taatgcaggt
     4501 taacctggct tatcgaaatt aatac
//
