LOCUS       sequence-146278-        4415 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 4415)
  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 4415)
  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-PK9
FEATURES             Location/Qualifiers
     source          1..4415
                     /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"
     CDS             676..717
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             721..762
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             763..804
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     terminator      834..1021
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(889..913)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     gene            1080..2436
                     /label=kanMX
                     /note="yeast selectable marker conferring kanamycin 
                     resistance (Wach et al., 1994)"
     promoter        1080..1423
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             1424..2233
                     /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(1491..1510)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      2239..2436
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(2495..2512)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2666..3254)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2746..2765)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3425..4216,4217..4285))
                     /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     4048..4067
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4286..4390)
                     /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 ctagcggtaa gcctattcct aaccctcttt tgggtctcga ttctacaggt
      721 ggaaaaccaa tccccaaccc actcctcggc cttgactcaa ctggcaaacc cataccaaat
      781 cctttacttg gattagatag tacctctaga tgaggcgcgc cacttctaaa taagcgaatt
      841 tcttatgatt tatgattttt attattaaat aagttataaa aaaaataagt gtatacaaat
      901 tttaaagtga ctcttaggtt ttaaaacgaa aattcttatt cttgagtaac tctttcctgt
      961 aggtcaggtt gctttctcag gtatagtatg aggtcgctct tattgaccac acctctaccg
     1021 gcagatccgc taggagatct gtttagcttg cctcgtcccc gccgggtcac ccggccagcg
     1081 acatggaggc ccagaatacc ctccttgaca gtcttgacgt gcgcagctca ggggcatgat
     1141 gtgactgtcg cccgtacatt tagcccatac atccccatgt ataatcattt gcatccatac
     1201 attttgatgg ccgcacggcg cgaagcaaaa attacggctc ctcgctgcag acctgcgagc
     1261 agggaaacgc tcccctcaca gacgcgttga attgtcccca cgccgcgccc ctgtagagaa
     1321 atataaaagg ttaggatttg ccactgaggt tcttctttca tatacttcct tttaaaatct
     1381 tgctaggata cagttctcac atcacatccg aacataaaca accatgggta aggaaaagac
     1441 tcacgtttcg aggccgcgat taaattccaa catggatgct gatttatatg ggtataaatg
     1501 ggctcgcgat aatgtcgggc aatcaggtgc gacaatctat cgattgtatg ggaagcccga
     1561 tgcgccagag ttgtttctga aacatggcaa aggtagcgtt gccaatgatg ttacagatga
     1621 gatggtcaga ctaaactggc tgacggaatt tatgcctctt ccgaccatca agcattttat
     1681 ccgtactcct gatgatgcat ggttactcac cactgcgatc cccggcaaaa cagcattcca
     1741 ggtattagaa gaatatcctg attcaggtga aaatattgtt gatgcgctgg cagtgttcct
     1801 gcgccggttg cattcgattc ctgtttgtaa ttgtcctttt aacagcgatc gcgtatttcg
     1861 tctcgctcag gcgcaatcac gaatgaataa cggtttggtt gatgcgagtg attttgatga
     1921 cgagcgtaat ggctggcctg ttgaacaagt ctggaaagaa atgcataagc ttttgccatt
     1981 ctcaccggat tcagtcgtca ctcatggtga tttctcactt gataacctta tttttgacga
     2041 ggggaaatta ataggttgta ttgatgttgg acgagtcgga atcgcagacc gataccagga
     2101 tcttgccatc ctatggaact gcctcggtga gttttctcct tcattacaga aacggctttt
     2161 tcaaaaatat ggtattgata atcctgatat gaataaattg cagtttcatt tgatgctcga
     2221 tgagtttttc taatcagtac tgacaataaa aagattcttg ttttcaagaa cttgtcattt
     2281 gtatagtttt tttatattgt agttgttcta ttttaatcaa atgttagcgt gatttatatt
     2341 ttttttcgcc tcgacatcat ctgcccagat gcgaagttaa gtgcgcagaa agtaatatca
     2401 tgcgtcaatc gtatgtgaat gctggtcgct atactgctgt cgattcgata ctaacgccgc
     2461 catccagttt aaacgagctc gaattctagg cttcctcgct cactgactcg ctgcgctcgg
     2521 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag
     2581 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
     2641 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
     2701 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
     2761 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     2821 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca atgctcacgc tgtaggtatc
     2881 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     2941 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     3001 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     3061 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaaggaca gtatttggta
     3121 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     3181 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
     3241 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
     3301 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc
     3361 ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg
     3421 acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat
     3481 ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg
     3541 gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa
     3601 taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca
     3661 tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc
     3721 gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt
     3781 cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa
     3841 aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat
     3901 cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct
     3961 tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga
     4021 gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag
     4081 tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga
     4141 gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca
     4201 ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg
     4261 cgacacggaa atgttgaata ctcatactct tcctttttca atattattga agcatttatc
     4321 agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag
     4381 gggttccgcg cacatttccc cgaaaagtgc cacct
//
