LOCUS       sequence-149544-        4316 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 4316)
  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 4316)
  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: pNX3c-PK12
FEATURES             Location/Qualifiers
     source          1..4316
                     /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      972..1159
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(1027..1051)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     gene            1218..2337
                     /label=natMX6
                     /note="yeast selectable marker conferring nourseothricin 
                     resistance"
     promoter        1218..1561
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             1562..2134
                     /codon_start=1
                     /gene="Streptomyces noursei nat1"
                     /product="nourseothricin acetyltransferase"
                     /label=NrsR
                     /note="confers resistance to nourseothricin"
                     /translation="MGTTLDDTAYRYRTSVPGDAEAIEALDGSFTTDTVFRVTATGDGF
                     TLREVPVDPPLTKVFPDDESDDESDDGEDGDPDSRTFVAYGDDGDLAGFVVVSYSGWNR
                     RLTVEDIEVAPEHRGHGVGRALMGLATEFARERGAGHLWLEVTNVNAPAIHAYRRMGFT
                     LCGLDTALYDGTASDGEQALYMSMPCP"
     terminator      2140..2337
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(2396..2413)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2567..3155)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2647..2666)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3326..4117,4118..4186))
                     /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     3949..3968
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4187..4291)
                     /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 ctagagattg aggcgcgcca
      961 cttctaaata agcgaatttc ttatgattta tgatttttat tattaaataa gttataaaaa
     1021 aaataagtgt atacaaattt taaagtgact cttaggtttt aaaacgaaaa ttcttattct
     1081 tgagtaactc tttcctgtag gtcaggttgc tttctcaggt atagtatgag gtcgctctta
     1141 ttgaccacac ctctaccggc agatccgcta ggagatctgt ttagcttgcc tcgtccccgc
     1201 cgggtcaccc ggccagcgac atggaggccc agaataccct ccttgacagt cttgacgtgc
     1261 gcagctcagg ggcatgatgt gactgtcgcc cgtacattta gcccatacat ccccatgtat
     1321 aatcatttgc atccatacat tttgatggcc gcacggcgcg aagcaaaaat tacggctcct
     1381 cgctgcagac ctgcgagcag ggaaacgctc ccctcacaga cgcgttgaat tgtccccacg
     1441 ccgcgcccct gtagagaaat ataaaaggtt aggatttgcc actgaggttc ttctttcata
     1501 tacttccttt taaaatcttg ctaggataca gttctcacat cacatccgaa cataaacaac
     1561 catgggtacc actcttgacg acacggctta ccggtaccgc accagtgtcc cgggggacgc
     1621 cgaggccatc gaggcactgg atgggtcctt caccaccgac accgtcttcc gcgtcaccgc
     1681 caccggggac ggcttcaccc tgcgggaggt gccggtggac ccgcccctga ccaaggtgtt
     1741 ccccgacgac gaatcggacg acgaatcgga cgacggggag gacggcgacc cggactcccg
     1801 gacgttcgtc gcgtacgggg acgacggcga cctggcgggc ttcgtggtcg tctcgtactc
     1861 cggctggaac cgccggctga ccgtcgagga catcgaggtc gccccggagc accgggggca
     1921 cggggtcggg cgcgcgttga tggggctcgc gacggagttc gcccgcgagc ggggcgccgg
     1981 gcacctctgg ctggaggtca ccaacgtcaa cgcaccggcg atccacgcgt accggcggat
     2041 ggggttcacc ctctgcggcc tggacaccgc cctgtacgac ggcaccgcct cggacggcga
     2101 gcaggcgctc tacatgagca tgccctgccc ctaatcagta ctgacaataa aaagattctt
     2161 gttttcaaga acttgtcatt tgtatagttt ttttatattg tagttgttct attttaatca
     2221 aatgttagcg tgatttatat tttttttcgc ctcgacatca tctgcccaga tgcgaagtta
     2281 agtgcgcaga aagtaatatc atgcgtcaat cgtatgtgaa tgctggtcgc tatactgctg
     2341 tcgattcgat actaacgccg ccatccagtt taaacgagct cgaattctag gcttcctcgc
     2401 tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg
     2461 cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag
     2521 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc
     2581 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
     2641 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
     2701 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
     2761 aatgctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
     2821 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
     2881 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
     2941 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
     3001 ctagaaggac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
     3061 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
     3121 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg
     3181 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa
     3241 aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta
     3301 tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag
     3361 cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga
     3421 tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac
     3481 cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc
     3541 ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta
     3601 gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac
     3661 gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat
     3721 gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa
     3781 gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg
     3841 tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag
     3901 aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc
     3961 cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct
     4021 caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat
     4081 cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg
     4141 ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc
     4201 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
     4261 tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacct
//
