LOCUS       sequence-169914-        4478 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  N-ICE plasmid pH3FS-GAL1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4478)
  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 4478)
  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: pH3FS-GAL1
FEATURES             Location/Qualifiers
     source          1..4478
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     127..144
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     231..250
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        231..249
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    complement(307..340)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        382..725
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             726..1754
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     CDS             726..1754
                     /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        1827..2268
                     /gene="S. cerevisiae GAL1"
                     /label=GAL1 promoter
                     /note="inducible promoter, regulated by Gal4"
     protein_bind    1827..1944
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     primer_bind     2239..2262
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     protein_bind    complement(2286..2319)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             2342..2413
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags"
                     /label=3xFLAG
                     /translation="DYKDDDDKDYKDDDDKDYKDDDDK"
     primer_bind     complement(2462..2481)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2581..2603
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2641..2659)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2727..2831
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2832..2900,2901..3692)
                     /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(3050..3069)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3863..4451
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4352..4371
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct
       61 gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag ccgaacgacc
      121 gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc caatacgcaa accgcctctc
      181 cccgcgcgtt ggccgattca ttaatgcagg ttaacctggc ttatcgaaat taatacgact
      241 cactataggg agaccggcag atccgcggcc gcgtacgctg caggtcgacg gatctgatat
      301 cacctaataa cttcgtatag catacattat acgaagttat attaagggtt ctcgacgtcc
      361 ccgccgggtc acccggccag cgacatggag gcccagaata ccctccttga cagtcttgac
      421 gtgcgcagct caggggcatg atgtgactgt cgcccgtaca tttagcccat acatccccat
      481 gtataatcat ttgcatccat acattttgat ggccgcacgg cgcgaagcaa aaattacggc
      541 tcctcgctgc ggacctgcga gcagggaaac gctcccctca cagacgcgtt gaattgtccc
      601 cacgccgcgc ccctgtagag aaatataaaa ggttaggatt tgccactgag gttcttcttt
      661 catatacttc cttttaaaat cttgctagga tacagttctc acatcacatc cgaacataaa
      721 caaccatggg taaaaagcct gaactcaccg cgacgtctgt cgagaagttt ctgatcgaaa
      781 agttcgacag cgtctccgac ctgatgcagc tctcggaggg cgaagaatct cgtgctttca
      841 gcttcgatgt aggagggcgt ggatatgtcc tgcgggtaaa tagctgcgcc gatggtttct
      901 acaaagatcg ttatgtttat cggcactttg catcggccgc gctcccgatt ccggaagtgc
      961 ttgacattgg ggaattcagc gagagcctga cctattgcat ctcccgccgt gcacagggtg
     1021 tcacgttgca agacctgcct gaaaccgaac tgcccgctgt tctgcagccg gtcgcggagg
     1081 ccatggatgc gatcgctgcg gccgatctta gccagacgag cgggttcggc ccattcggac
     1141 cgcaaggaat cggtcaatac actacatggc gtgatttcat atgcgcgatt gctgatcccc
     1201 atgtgtatca ctggcaaact gtgatggacg acaccgtcag tgcgtccgtc gcgcaggctc
     1261 tcgatgagct gatgctttgg gccgaggact gccccgaagt ccggcacctc gtgcacgcgg
     1321 atttcggctc caacaatgtc ctgacggaca atggccgcat aacagcggtc attgactgga
     1381 gcgaggcgat gttcggggat tcccaatacg aggtcgccaa catcttcttc tggaggccgt
     1441 ggttggcttg tatggagcag cagacgcgct acttcgagcg gaggcatccg gagcttgcag
     1501 gatcgccgcg gctccgggcg tatatgctcc gcattggtct tgaccaactc tatcagagct
     1561 tggttgacgg caatttcgat gatgcagctt gggcgcaggg tcgatgcgac gcaatcgtcc
     1621 gatccggagc cgggactgtc gggcgtacac aaatcgcccg cagaagcgcg gccgtctgga
     1681 ccgatggctg tgtagaagta ctcgccgata gtggaaaccg acgccccagc actcgtccga
     1741 gggcaaagga ataatgcccg ctattaacgc tttgtaatgt atagctttta atgtgtgatc
     1801 gcctgacttt tagatctcta gtagtacgga ttagaagccg ccgagcgggc gacagccctc
     1861 cgacggaaga ctctcctccg tgcgtcctcg tcttcaccgg tcgcgttcct gaaacgcaga
     1921 tgtgcctcgc gccgcactgc tccgaacaat aaagattcta caatactagc ttttatggtt
     1981 atgaagagga aaaattggca gtaacctggc cccacaaacc ttcaaattaa cgaatcaaat
     2041 taacaaccat aggatgataa tgcgattagt ttcttagcct tatttctggg gtaattaatc
     2101 agcgaagcga tgatttttga tctattaaca gatatataaa tggaaaagct gcataaccac
     2161 tttaactaat actttcaaca ttttcagttt gtattacttc ttattcaaat gtcataaaag
     2221 tatcaacaaa aaattgttaa tatacctcta tactttaacg tcaaggagaa aaaactctag
     2281 acctaataac ttcgtatagc atacattata cgaagttata ttaagggttg tcgacaccat
     2341 ggactataaa gacgacgacg acaaagatta taaggacgac gacgataagg attacaagga
     2401 cgacgacgat aaaggatcca tggcgacaga gaattcatcg atgctcgagg tgcactctca
     2461 gtacaatctg ctctgatgcc gcatagttaa gccagccccg acacccgcca acacccgctg
     2521 acgcgccctg acgggcttgt ctgctcccgg catccgctta cagacaagct gtgaccgtct
     2581 ccgggagctg catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg agacgaaagg
     2641 gcctcgtgat acgcctattt ttataggtta atgtcatgat aataatggtt tcttagacgt
     2701 caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac
     2761 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa
     2821 aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat
     2881 tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc
     2941 agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga
     3001 gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg
     3061 cggtattatc ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc
     3121 agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag
     3181 taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc
     3241 tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg
     3301 taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg
     3361 acaccacgat gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac
     3421 ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac
     3481 cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg
     3541 agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg
     3601 tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg
     3661 agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac
     3721 tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg
     3781 ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg
     3841 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc
     3901 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc
     3961 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt
     4021 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
     4081 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact
     4141 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac
     4201 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag
     4261 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg
     4321 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg
     4381 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga
     4441 gcctatggaa aaacgccagc aacgcggcct ttttacgg
//
