LOCUS       sequence-146277-        4280 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  fission yeast epitope tagging.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4280)
  AUTHORS   Amelina H, Moiseeva V, Collopy LC, Pearson SR, Armstrong CA, Tomita 
            K
  TITLE     Sequential and counter-selectable cassettes for fission yeast.
  JOURNAL   BMC Biotechnol. 2016 Nov 8;16(1):76.
  PUBMED    27825338
REFERENCE   2  (bases 1 to 4280)
  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: "BMC 
            Biotechnol."; date: "2016-11-8"; volume: "16(1)"; pages: "76"
COMMENT     Sequence Label: pNX3a-PK6
FEATURES             Location/Qualifiers
     source          1..4280
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     43..61
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(99..121)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     221..240
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        321..339
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     321..338
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     CDS             406..447
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             451..492
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             493..534
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             541..582
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             586..627
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             628..669
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     terminator      699..886
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(754..778)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     gene            945..2301
                     /label=kanMX
                     /note="yeast selectable marker conferring kanamycin 
                     resistance (Wach et al., 1994)"
     promoter        945..1288
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             1289..2098
                     /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(1356..1375)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      2104..2301
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(2360..2377)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2531..3119)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2611..2630)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3290..4081,4082..4150))
                     /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     3913..3932
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4151..4255)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacgtctaag aaaccattat tatcatgaca ttaacctata aaaataggcg tatcacgagg
       61 ccctttcgtc tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg
      121 gagacggtca cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg
      181 tcagcgggtg ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta
      241 ctgagagtgc accatatgga catattgtcg ttagaacgcg gctacaatta atacataacc
      301 ttatgtatca tacacatacg atttaggtga cactatagaa cgcggccgcc agctgaagct
      361 tcgtacgctg caggtcgacg gatccccggg ttaattaacg ctagcggtaa gcctattcct
      421 aaccctcttt tgggtctcga ttctacaggt ggaaaaccaa tccccaaccc actcctcggc
      481 cttgactcaa ctggcaaacc cataccaaat cctttacttg gattagatag tacctctagc
      541 ggtaagccta ttcctaaccc tcttttgggt ctcgattcta caggtggaaa accaatcccc
      601 aacccactcc tcggccttga ctcaactggc aaacccatac caaatccttt acttggatta
      661 gatagtacct ctagatgagg cgcgccactt ctaaataagc gaatttctta tgatttatga
      721 tttttattat taaataagtt ataaaaaaaa taagtgtata caaattttaa agtgactctt
      781 aggttttaaa acgaaaattc ttattcttga gtaactcttt cctgtaggtc aggttgcttt
      841 ctcaggtata gtatgaggtc gctcttattg accacacctc taccggcaga tccgctagga
      901 gatctgttta gcttgcctcg tccccgccgg gtcacccggc cagcgacatg gaggcccaga
      961 ataccctcct tgacagtctt gacgtgcgca gctcaggggc atgatgtgac tgtcgcccgt
     1021 acatttagcc catacatccc catgtataat catttgcatc catacatttt gatggccgca
     1081 cggcgcgaag caaaaattac ggctcctcgc tgcagacctg cgagcaggga aacgctcccc
     1141 tcacagacgc gttgaattgt ccccacgccg cgcccctgta gagaaatata aaaggttagg
     1201 atttgccact gaggttcttc tttcatatac ttccttttaa aatcttgcta ggatacagtt
     1261 ctcacatcac atccgaacat aaacaaccat gggtaaggaa aagactcacg tttcgaggcc
     1321 gcgattaaat tccaacatgg atgctgattt atatgggtat aaatgggctc gcgataatgt
     1381 cgggcaatca ggtgcgacaa tctatcgatt gtatgggaag cccgatgcgc cagagttgtt
     1441 tctgaaacat ggcaaaggta gcgttgccaa tgatgttaca gatgagatgg tcagactaaa
     1501 ctggctgacg gaatttatgc ctcttccgac catcaagcat tttatccgta ctcctgatga
     1561 tgcatggtta ctcaccactg cgatccccgg caaaacagca ttccaggtat tagaagaata
     1621 tcctgattca ggtgaaaata ttgttgatgc gctggcagtg ttcctgcgcc ggttgcattc
     1681 gattcctgtt tgtaattgtc cttttaacag cgatcgcgta tttcgtctcg ctcaggcgca
     1741 atcacgaatg aataacggtt tggttgatgc gagtgatttt gatgacgagc gtaatggctg
     1801 gcctgttgaa caagtctgga aagaaatgca taagcttttg ccattctcac cggattcagt
     1861 cgtcactcat ggtgatttct cacttgataa ccttattttt gacgagggga aattaatagg
     1921 ttgtattgat gttggacgag tcggaatcgc agaccgatac caggatcttg ccatcctatg
     1981 gaactgcctc ggtgagtttt ctccttcatt acagaaacgg ctttttcaaa aatatggtat
     2041 tgataatcct gatatgaata aattgcagtt tcatttgatg ctcgatgagt ttttctaatc
     2101 agtactgaca ataaaaagat tcttgttttc aagaacttgt catttgtata gtttttttat
     2161 attgtagttg ttctatttta atcaaatgtt agcgtgattt atattttttt tcgcctcgac
     2221 atcatctgcc cagatgcgaa gttaagtgcg cagaaagtaa tatcatgcgt caatcgtatg
     2281 tgaatgctgg tcgctatact gctgtcgatt cgatactaac gccgccatcc agtttaaacg
     2341 agctcgaatt ctaggcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
     2401 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     2461 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     2521 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     2581 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     2641 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     2701 cttcgggaag cgtggcgctt tctcaatgct cacgctgtag gtatctcagt tcggtgtagg
     2761 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     2821 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     2881 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     2941 agtggtggcc taactacggc tacactagaa ggacagtatt tggtatctgc gctctgctga
     3001 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     3061 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     3121 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     3181 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     3241 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     3301 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     3361 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     3421 tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     3481 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     3541 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     3601 ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     3661 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     3721 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     3781 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     3841 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     3901 cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     3961 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     4021 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     4081 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     4141 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     4201 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
     4261 ttccccgaaa agtgccacct
//
