LOCUS       sequence-171079-        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"
     primer_bind     89..108
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     342..359
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     446..465
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        446..464
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    complement(518..551)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     gene            593..1949
                     /label=kanMX
                     /note="yeast selectable marker conferring kanamycin 
                     resistance (Wach et al., 1994)"
     promoter        593..936
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             937..1746
                     /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(1004..1023)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      1752..1949
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     promoter        2001..2442
                     /gene="S. cerevisiae GAL1"
                     /label=GAL1 promoter
                     /note="inducible promoter, regulated by Gal4"
     protein_bind    2001..2118
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     primer_bind     2413..2436
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     protein_bind    complement(2460..2493)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             2516..2587
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags"
                     /label=3xFLAG
                     /translation="DYKDDDDKDYKDDDDKDYKDDDDK"
     promoter        complement(2625..2643)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2626..2643)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2724..2743)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2843..2865
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2903..2921)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2989..3093
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3094..3162,3163..3954)
                     /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(3312..3331)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(4125..4525,1..188)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa
       61 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg
      121 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc
      181 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg
      241 ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg
      301 agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg
      361 aagcggaaga gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat
      421 gcaggttaac ctggcttatc gaaattaata cgactcacta tagggagacc ggcagatccg
      481 cggccgcata ggccactagt ggatctgata tcacctaata acttcgtata gcatacatta
      541 tacgaagtta tattaagggt tctcgacgtc cccgccgggt cacccggcca gcgacatgga
      601 ggcccagaat accctccttg acagtcttga cgtgcgcagc tcaggggcat gatgtgactg
      661 tcgcccgtac atttagccca tacatcccca tgtataatca tttgcatcca tacattttga
      721 tggccgcacg gcgcgaagca aaaattacgg ctcctcgctg cagacctgcg agcagggaaa
      781 cgctcccctc acagacgcgt tgaattgtcc ccacgccgcg cccctgtaga gaaatataaa
      841 aggttaggat ttgccactga ggttcttctt tcatatactt ccttttaaaa tcttgctagg
      901 atacagttct cacatcacat ccgaacataa acaaccatgg gtaaggaaaa gactcacgtt
      961 tcgaggccgc gattaaattc caacatggat gctgatttat atgggtataa atgggctcgc
     1021 gataatgtcg ggcaatcagg tgcgacaatc tatcgattgt atgggaagcc cgatgcgcca
     1081 gagttgtttc tgaaacatgg caaaggtagc gttgccaatg atgttacaga tgagatggtc
     1141 agactaaact ggctgacgga atttatgcct cttccgacca tcaagcattt tatccgtact
     1201 cctgatgatg catggttact caccactgcg atccccggca aaacagcatt ccaggtatta
     1261 gaagaatatc ctgattcagg tgaaaatatt gttgatgcgc tggcagtgtt cctgcgccgg
     1321 ttgcattcga ttcctgtttg taattgtcct tttaacagcg atcgcgtatt tcgtctcgct
     1381 caggcgcaat cacgaatgaa taacggtttg gttgatgcga gtgattttga tgacgagcgt
     1441 aatggctggc ctgttgaaca agtctggaaa gaaatgcata agcttttgcc attctcaccg
     1501 gattcagtcg tcactcatgg tgatttctca cttgataacc ttatttttga cgaggggaaa
     1561 ttaataggtt gtattgatgt tggacgagtc ggaatcgcag accgatacca ggatcttgcc
     1621 atcctatgga actgcctcgg tgagttttct ccttcattac agaaacggct ttttcaaaaa
     1681 tatggtattg ataatcctga tatgaataaa ttgcagtttc atttgatgct cgatgagttt
     1741 ttctaatcag tactgacaat aaaaagattc ttgttttcaa gaacttgtca tttgtatagt
     1801 ttttttatat tgtagttgtt ctattttaat caaatgttag cgtgatttat attttttttc
     1861 gcctcgacat catctgccca gatgcgaagt taagtgcgca gaaagtaata tcatgcgtca
     1921 atcgtatgtg aatgctggtc gctatactgc tgtcgattcg atactaacgc cgccatccag
     1981 tgtcgagatc tctagtagta cggattagaa gccgccgagc gggcgacagc cctccgacgg
     2041 aagactctcc tccgtgcgtc ctcgtcttca ccggtcgcgt tcctgaaacg cagatgtgcc
     2101 tcgcgccgca ctgctccgaa caataaagat tctacaatac tagcttttat ggttatgaag
     2161 aggaaaaatt ggcagtaacc tggccccaca aaccttcaaa ttaacgaatc aaattaacaa
     2221 ccataggatg ataatgcgat tagtttttta gccttatttc tggggtaatt aatcagcgaa
     2281 gcgatgattt ttgatctatt aacagatata taaatggaaa agctgcataa ccactttaac
     2341 taatactttc aacattttca gtttgtatta cttcttattc aaatgtcata aaagtatcaa
     2401 caaaaaattg ttaatatacc tctatacttt aacgtcaagg agaaaaaact ctagacctaa
     2461 taacttcgta tagcatacat tatacgaagt tatattaagg gttgtcgaca ccatggacta
     2521 taaagacgac gacgacaaag attataagga cgacgacgat aaggattaca aggacgacga
     2581 cgataaagga tccatgggcg gccgcgagat cctggcggcc gcgttctata gtgtcaccta
     2641 aatcgtatgt gtatgataca taaggttatg tattaattgt agccgcgttc taacgacaat
     2701 atgtccatat ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagccc
     2761 cgacacccgc caacacccgc tgacgcgccc tgacgggctt gtctgctccc ggcatccgct
     2821 tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc accgtcatca
     2881 ccgaaacgcg cgagacgaaa gggcctcgtg atacgcctat ttttataggt taatgtcatg
     2941 ataataatgg tttcttagac gtcaggtggc acttttcggg gaaatgtgcg cggaacccct
     3001 atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga
     3061 taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc
     3121 cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg
     3181 aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga actggatctc
     3241 aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat gatgagcact
     3301 tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca agagcaactc
     3361 ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt cacagaaaag
     3421 catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat
     3481 aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct aaccgctttt
     3541 ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa
     3601 gccataccaa acgacgagcg tgacaccacg atgcctgtag caatggcaac aacgttgcgc
     3661 aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat agactggatg
     3721 gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt
     3781 gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca
     3841 gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat
     3901 gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg gtaactgtca
     3961 gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg
     4021 atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg
     4081 ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt
     4141 ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg
     4201 ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata
     4261 ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca
     4321 ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag
     4381 tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc
     4441 tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga
     4501 tacctacagc gtgagctatg agaaa
//
