LOCUS       sequence-193168-        3855 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  N-ICE plasmid pH3FS-CYC1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3855)
  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 3855)
  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: pN3FS-CYC1
FEATURES             Location/Qualifiers
     source          1..3855
                     /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"
     CDS             complement(2244..2315)
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags"
                     /label=3xFLAG
                     /translation="DYKDDDDKDYKDDDDKDYKDDDDK"
     protein_bind    2338..2371
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(2736..3308)
                     /codon_start=1
                     /gene="Streptomyces noursei nat1"
                     /product="nourseothricin acetyltransferase"
                     /label=NrsR
                     /note="confers resistance to nourseothricin"
                     /translation="MGTTLDDTAYRYRTSVPGDAEAIEALDGSFTTDTVFRVTATGDGF
                     TLREVPVDPPLTKVFPDDESDDESDDGEDGDLDSRTFVAYGDDGDLAGFVVVSYSGWNR
                     RLTVEDIEVAPEHRGHGVGRALMGLATEFARERGAGHLWLEVTNVNAPAIHAYRRMGFT
                     LCGLDTALYDGTASDGEQALYMSMPCP"
     promoter        complement(3309..3652)
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     protein_bind    3694..3727
                     /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(3784..3803)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3785..3803)
                     /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 agtgcacctc gagcatcgat
     2221 gaattctctg tcgccatgga tcctttatcg tcgtcgtcct tgtaatcctt atcgtcgtcg
     2281 tccttataat ctttgtcgtc gtcgtcttta tagtccatgg tgtcgacaac ccttaatata
     2341 acttcgtata atgtatgcta tacgaagtta ttaggtctag atattaattt agtgtgtgta
     2401 tttgtgtttg tgtgtctata gaagtatagt aatttatgct gcaaaggtcc taatgtataa
     2461 ggaaagaata tttagagaaa agaagaaaac aagagtttta tatacataca gagcacatgc
     2521 atgccatatg atcatgtgtc gtcgcacaca tatatatatg cctgtatgtg tcagcactaa
     2581 agttgcctgg ccatccacgc tatatataca cgcctggcgg atctgctcga ggattgccta
     2641 cgcgtgggct tgatccacca accaacgcag atctatacac ttattttttt tataacttat
     2701 ttaataataa aaatcataaa tcataagaaa ttcgcttagg ggcagggcat gctcatgtag
     2761 agcgcctgct cgccgtccga ggcggtgccg tcgtacaggg cggtgtccag gccgcagagg
     2821 gtgaacccca tccgccggta cgcgtggatc gccggtgcgt tgacgttggt gacctccagc
     2881 cagaggtgcc cggcgccccg ctcgcgggcg aactccgtcg cgagccccat caacgcgcgc
     2941 ccgaccccgt gcccccggtg ctccggggcg acctcgatgt cctcgacggt cagccggcgg
     3001 ttccagccgg agtacgagac gaccacgaag cccgccaggt cgccgtcgtc cccgtacgcg
     3061 acgaacgtcc gggagtccag gtcgccgtcc tccccgtcgt ccgattcgtc gtccgattcg
     3121 tcgtcgggga acaccttggt caggggcggg tccaccggca cctcccgcag ggtgaagccg
     3181 tccccggtgg cggtgacgcg gaagacggtg tcggtggtga aggacccatc cagtgcctcg
     3241 atggcctcgg cgtcccccgg gacactggtg cggtaccggt aagccgtgtc gtcaagagtg
     3301 gtacccatgg ttgtttatgt tcggatgtga tgtgagaact gtatcctagc aagattttaa
     3361 aaggaagtat atgaaagaag aacctcagtg gcaaatccta accttttata tttctctaca
     3421 ggggcgcggc gtggggacaa ttcaacgcgt ctgtgagggg agcgtttccc tgctcgcagg
     3481 tctgcagcga ggagccgtaa tttttgcttc gcgccgtgcg gccatcaaaa tgtatggatg
     3541 caaatgatta tacatgggga tgtatgggct aaatgtacgg gcgacagtca catcatgccc
     3601 ctgagctgcg cacgtcaaga ctgtcaagga gggtattctg ggcctccatg tcgctggccg
     3661 ggtgacccgg cggggacgtc gagaaccctt aatataactt cgtataatgt atgctatacg
     3721 aagttattag gtgatatcag atccgtcgac ctgcagcgta cgcggccgcg gatctgccgg
     3781 tctccctata gtgagtcgta ttaatttcga taagccaggt taacctgcat taatgaatcg
     3841 gccaacgcgc gggga
//
