LOCUS       sequence-171078-        4285 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  N-ICE plasmid pH3F.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4285)
  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 4285)
  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: pH3F
FEATURES             Location/Qualifiers
     source          1..4285
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(35..52)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(206..794)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(286..305)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(965..1756,1757..1825))
                     /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     1588..1607
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1826..1930)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     1998..2016
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(2054..2076)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     2176..2195
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        2276..2294
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     2276..2293
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     CDS             complement(2332..2403)
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags"
                     /label=3xFLAG
                     /translation="DYKDDDDKDYKDDDDKDYKDDDDK"
     protein_bind    2426..2459
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     gene            complement(2511..4086)
                     /label=hphMX6
                     /note="yeast selectable marker conferring hygromycin 
                     resistance"
     terminator      complement(2511..2708)
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     CDS             complement(2714..3742)
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     CDS             complement(2714..3742)
                     /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        complement(3743..4086)
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     protein_bind    4128..4161
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     complement(4214..4233)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4215..4233)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg
       61 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag
      121 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
      181 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
      241 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
      301 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
      361 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
      421 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
      481 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
      541 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
      601 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta
      661 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
      721 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
      781 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
      841 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc
      901 ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg
      961 acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat
     1021 ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg
     1081 gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa
     1141 taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca
     1201 tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc
     1261 gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt
     1321 cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa
     1381 aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat
     1441 cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct
     1501 tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga
     1561 gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag
     1621 tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga
     1681 gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca
     1741 ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg
     1801 cgacacggaa atgttgaata ctcatactct tcctttttca atattattga agcatttatc
     1861 agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag
     1921 gggttccgcg cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc attattatca
     1981 tgacattaac ctataaaaat aggcgtatca cgaggccctt tcgtctcgcg cgtttcggtg
     2041 atgacggtga aaacctctga cacatgcagc tcccggagac ggtcacagct tgtctgtaag
     2101 cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc gggtgtcggg
     2161 gctggcttaa ctatgcggca tcagagcaga ttgtactgag agtgcaccat atggacatat
     2221 tgtcgttaga acgcggctac aattaataca taaccttatg tatcatacac atacgattta
     2281 ggtgacacta tagaacgcgg ccgccaggat ctcgcggccg cccatggatc ctttatcgtc
     2341 gtcgtccttg taatccttat cgtcgtcgtc cttataatct ttgtcgtcgt cgtctttata
     2401 gtccatggtg tcgacaaccc ttaatataac ttcgtataat gtatgctata cgaagttatt
     2461 aggtctagag atctcgacac tggatggcgg cgttagtatc gaatcgacag cagtatagcg
     2521 accagcattc acatacgatt gacgcatgat attactttct gcgcacttaa cttcgcatct
     2581 gggcagatga tgtcgaggcg aaaaaaaata taaatcacgc taacatttga ttaaaataga
     2641 acaactacaa tataaaaaaa ctatacaaat gacaagttct tgaaaacaag aatcttttta
     2701 ttgtcagtac tgattattcc tttgccctcg gacgagtgct ggggcgtcgg tttccactat
     2761 cggcgagtac ttctacacag ccatcggtcc agacggccgc gcttctgcgg gcgatttgtg
     2821 tacgcccgac agtcccggct ccggatcgga cgattgcgtc gcatcgaccc tgcgcccaag
     2881 ctgcatcatc gaaattgccg tcaaccaagc tctgatagag ttggtcaaga ccaatgcgga
     2941 gcatatacgc ccggagccgc ggcgatcctg caagctccgg atgcctccgc tcgaagtagc
     3001 gcgtctgctg ctccatacaa gccaaccacg gcctccagaa gaagatgttg gcgacctcgt
     3061 attgggaatc cccgaacatc gcctcgctcc agtcaatgac cgctgttatg cggccattgt
     3121 ccgtcaggac attgttggag ccgaaatccg cgtgcacgag gtgccggact tcggggcagt
     3181 cctcggccca aagcatcagc tcatcgagag cctgcgcgac ggacgcactg acggtgtcgt
     3241 ccatcacagt ttgccagtga tacacatggg gatcagcaat cgcgcatatg aaatcacgcc
     3301 atgtagtgta ttgaccgatt ccttgcggtc cgaatgggcc gaacccgctc gtctggctaa
     3361 gatcggccgc agcgatcgca tccatggcct ccgcgaccgg ctgcagaaca gcgggcagtt
     3421 cggtttcagg caggtcttgc aacgtgacac cctgtgcacg gcgggagatg caataggtca
     3481 ggctctcgct gaattcccca atgtcaagca cttccggaat cgggagcgcg gccgatgcaa
     3541 agtgccgata aacataacga tctttgtaga aaccatcggc gcagctattt acccgcagga
     3601 catatccacg ccctcctaca tcgaagctga aagcacgaga ttcttcgccc tccgagagct
     3661 gcatcaggtc ggagacgctg tcgaactttt cgatcagaaa cttctcgaca gacgtcgcgg
     3721 tgagttcagg ctttttaccc atggttgttt atgttcggat gtgatgtgag aactgtatcc
     3781 tagcaagatt ttaaaaggaa gtatatgaaa gaagaacctc agtggcaaat cctaaccttt
     3841 tatatttctc tacaggggcg cggcgtgggg acaattcaac gcgtctgtga ggggagcgtt
     3901 tccctgctcg caggtccgca gcgaggagcc gtaatttttg cttcgcgccg tgcggccatc
     3961 aaaatgtatg gatgcaaatg attatacatg gggatgtatg ggctaaatgt acgggcgaca
     4021 gtcacatcat gcccctgagc tgcgcacgtc aagactgtca aggagggtat tctgggcctc
     4081 catgtcgctg gccgggtgac ccggcgggga cgtcgagaac ccttaatata acttcgtata
     4141 atgtatgcta tacgaagtta ttaggtgata tcagatccac tagtggccta tgcggccgcg
     4201 gatctgccgg tctccctata gtgagtcgta ttaatttcga taagccaggt taacctgcat
     4261 taatgaatcg gccaacgcgc gggga
//
