LOCUS       sequence-146279-        4550 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 4550)
  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 4550)
  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-PK12
FEATURES             Location/Qualifiers
     source          1..4550
                     /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"
     CDS             811..852
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             856..897
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             898..939
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     terminator      969..1156
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(1024..1048)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     gene            1215..2571
                     /label=kanMX
                     /note="yeast selectable marker conferring kanamycin 
                     resistance (Wach et al., 1994)"
     promoter        1215..1558
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             1559..2368
                     /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(1626..1645)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      2374..2571
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(2630..2647)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2801..3389)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2881..2900)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3560..4351,4352..4420))
                     /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     4183..4202
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4421..4525)
                     /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 tacctctagc ggtaagccta ttcctaaccc tcttttgggt
      841 ctcgattcta caggtggaaa accaatcccc aacccactcc tcggccttga ctcaactggc
      901 aaacccatac caaatccttt acttggatta gatagtacct ctagatgagg cgcgccactt
      961 ctaaataagc gaatttctta tgatttatga tttttattat taaataagtt ataaaaaaaa
     1021 taagtgtata caaattttaa agtgactctt aggttttaaa acgaaaattc ttattcttga
     1081 gtaactcttt cctgtaggtc aggttgcttt ctcaggtata gtatgaggtc gctcttattg
     1141 accacacctc taccggcaga tccgctagga gatctgttta gcttgcctcg tccccgccgg
     1201 gtcacccggc cagcgacatg gaggcccaga ataccctcct tgacagtctt gacgtgcgca
     1261 gctcaggggc atgatgtgac tgtcgcccgt acatttagcc catacatccc catgtataat
     1321 catttgcatc catacatttt gatggccgca cggcgcgaag caaaaattac ggctcctcgc
     1381 tgcagacctg cgagcaggga aacgctcccc tcacagacgc gttgaattgt ccccacgccg
     1441 cgcccctgta gagaaatata aaaggttagg atttgccact gaggttcttc tttcatatac
     1501 ttccttttaa aatcttgcta ggatacagtt ctcacatcac atccgaacat aaacaaccat
     1561 gggtaaggaa aagactcacg tttcgaggcc gcgattaaat tccaacatgg atgctgattt
     1621 atatgggtat aaatgggctc gcgataatgt cgggcaatca ggtgcgacaa tctatcgatt
     1681 gtatgggaag cccgatgcgc cagagttgtt tctgaaacat ggcaaaggta gcgttgccaa
     1741 tgatgttaca gatgagatgg tcagactaaa ctggctgacg gaatttatgc ctcttccgac
     1801 catcaagcat tttatccgta ctcctgatga tgcatggtta ctcaccactg cgatccccgg
     1861 caaaacagca ttccaggtat tagaagaata tcctgattca ggtgaaaata ttgttgatgc
     1921 gctggcagtg ttcctgcgcc ggttgcattc gattcctgtt tgtaattgtc cttttaacag
     1981 cgatcgcgta tttcgtctcg ctcaggcgca atcacgaatg aataacggtt tggttgatgc
     2041 gagtgatttt gatgacgagc gtaatggctg gcctgttgaa caagtctgga aagaaatgca
     2101 taagcttttg ccattctcac cggattcagt cgtcactcat ggtgatttct cacttgataa
     2161 ccttattttt gacgagggga aattaatagg ttgtattgat gttggacgag tcggaatcgc
     2221 agaccgatac caggatcttg ccatcctatg gaactgcctc ggtgagtttt ctccttcatt
     2281 acagaaacgg ctttttcaaa aatatggtat tgataatcct gatatgaata aattgcagtt
     2341 tcatttgatg ctcgatgagt ttttctaatc agtactgaca ataaaaagat tcttgttttc
     2401 aagaacttgt catttgtata gtttttttat attgtagttg ttctatttta atcaaatgtt
     2461 agcgtgattt atattttttt tcgcctcgac atcatctgcc cagatgcgaa gttaagtgcg
     2521 cagaaagtaa tatcatgcgt caatcgtatg tgaatgctgg tcgctatact gctgtcgatt
     2581 cgatactaac gccgccatcc agtttaaacg agctcgaatt ctaggcttcc tcgctcactg
     2641 actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa
     2701 tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc
     2761 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc
     2821 ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat
     2881 aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc
     2941 cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcaatgct
     3001 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg
     3061 aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc
     3121 cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga
     3181 ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa
     3241 ggacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta
     3301 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc
     3361 agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg
     3421 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga
     3481 tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg
     3541 agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct
     3601 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg
     3661 agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc
     3721 cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa
     3781 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc
     3841 cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt
     3901 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc
     3961 ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt
     4021 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc
     4081 catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt
     4141 gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata
     4201 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga
     4261 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag
     4321 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa
     4381 aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt
     4441 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga
     4501 aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
//
