LOCUS       sequence-72612-e        4555 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  genomic G9-5FLAG epitope tagging in S. pombe.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4555)
  AUTHORS   Gadaleta MC, Iwasaki O, Noguchi C, Noma K, Noguchi E
  TITLE     New vectors for epitope tagging and gene disruption in 
            Schizosaccharomyces pombe.
  JOURNAL   Biotechniques. 2013 Nov;55(5):257-63. doi: 10.2144/000114100.
  PUBMED    24215641
REFERENCE   2  (bases 1 to 4555)
  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"; doi: "10"; journalName: 
            "Biotechniques"; date: "2013-11"; volume: "55"; issue: "5"; pages: 
            "257-63"
COMMENT     Sequence Label: pFA6a-G9-5FLAG-hphMX6
FEATURES             Location/Qualifiers
     source          1..4555
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             93..224
                     /codon_start=1
                     /product="five tandem FLAG(R) epitope tags"
                     /label=5xFLAG
                     /translation="DYKDDDDKDYKDDDDKLMDYKDDDDKDYKDDDDKLMDYKDDDDK"
     terminator      251..438
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(306..330)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     gene            513..2088
                     /label=hphMX6
                     /note="yeast selectable marker conferring hygromycin 
                     resistance"
     promoter        513..856
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             857..1885
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     CDS             857..1885
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     terminator      1891..2088
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(2192..2211)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2193..2211)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2298..2315)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2469..3057)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2549..2568)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3228..4019,4020..4088))
                     /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     3851..3870
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4089..4193)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4261..4279
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(4317..4339)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4439..4458
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        join(4539..4555,1..2)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     join(4539..4555,1)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
ORIGIN
        1 gaacgcggcc gccagctgaa gcttcgtacg ctgcaggtcg acggatcccc gggggaggtg
       61 gaggtggagg tggaggttta attaatcata tggactacaa ggacgacgat gacaaggatt
      121 acaaagatga cgacgataag cttatggact acaaggacga cgatgacaag gattacaaag
      181 atgacgacga taagcttatg gactacaagg acgacgatga caagcatatg ggcgcgccac
      241 ttctaaataa gcgaatttct tatgatttat gatttttatt attaaataag ttataaaaaa
      301 aataagtgta tacaaatttt aaagtgactc ttaggtttta aaacgaaaat tcttattctt
      361 gagtaactct ttcctgtagg tcaggttgct ttctcaggta tagtatgagg tcgctcttat
      421 tgaccacacc tctaccggca gatccgctag ggataacagg gtaatataga tctgtttagc
      481 ttgcctcgtc cccgccgggt cacccggcca gcgacatgga ggcccagaat accctccttg
      541 acagtcttga cgtgcgcagc tcaggggcat gatgtgactg tcgcccgtac atttagccca
      601 tacatcccca tgtataatca tttgcatcca tacattttga tggccgcacg gcgcgaagca
      661 aaaattacgg ctcctcgctg cagacctgcg agcagggaaa cgctcccctc acagacgcgt
      721 tgaattgtcc ccacgccgcg cccctgtaga gaaatataaa aggttaggat ttgccactga
      781 ggttcttctt tcatatactt ccttttaaaa tcttgctagg atacagttct cacatcacat
      841 ccgaacataa acaaccatgg gtaaaaagcc tgaactcacc gcgacgtctg tcgagaagtt
      901 tctgatcgaa aagttcgaca gcgtctccga cctgatgcag ctctcggagg gcgaagaatc
      961 tcgtgctttc agcttcgatg taggagggcg tggatatgtc ctgcgggtaa atagctgcgc
     1021 cgatggtttc tacaaagatc gttatgttta tcggcacttt gcatcggccg cgctcccgat
     1081 tccggaagtg cttgacattg gggaattcag cgagagcctg acctattgca tctcccgccg
     1141 tgcacagggt gtcacgttgc aagacctgcc tgaaaccgaa ctgcccgctg ttctgcagcc
     1201 ggtcgcggag gccatggatg cgatcgctgc ggccgatctt agccagacga gcgggttcgg
     1261 cccattcgga ccgcaaggaa tcggtcaata cactacatgg cgtgatttca tatgcgcgat
     1321 tgctgatccc catgtgtatc actggcaaac tgtgatggac gacaccgtca gtgcgtccgt
     1381 cgcgcaggct ctcgatgagc tgatgctttg ggccgaggac tgccccgaag tccggcacct
     1441 cgtgcacgcg gatttcggct ccaacaatgt cctgacggac aatggccgca taacagcggt
     1501 cattgactgg agcgaggcga tgttcgggga ttcccaatac gaggtcgcca acatcttctt
     1561 ctggaggccg tggttggctt gtatggagca gcagacgcgc tacttcgagc ggaggcatcc
     1621 ggagcttgca ggatcgccgc ggctccgggc gtatatgctc cgcattggtc ttgaccaact
     1681 ctatcagagc ttggttgacg gcaatttcga tgatgcagct tgggcgcagg gtcgatgcga
     1741 cgcaatcgtc cgatccggag ccgggactgt cgggcgtaca caaatcgccc gcagaagcgc
     1801 ggccgtctgg accgatggct gtgtagaagt actcgccgat agtggaaacc gacgccccag
     1861 cactcgtccg agggcaaagg aataatcagt actgacaata aaaagattct tgttttcaag
     1921 aacttgtcat ttgtatagtt tttttatatt gtagttgttc tattttaatc aaatgttagc
     1981 gtgatttata ttttttttcg cctcgacatc atctgcccag atgcgaagtt aagtgcgcag
     2041 aaagtaatat catgcgtcaa tcgtatgtga atgctggtcg ctatactgct gtcgattcga
     2101 tactaacgcc gccatccagt ttaaacgagc tcgaattcat cgatgatatc agatccacta
     2161 gtggcctatg cggccgcgga tctgccggtc tccctatagt gagtcgtatt aatttcgata
     2221 agccaggtta acctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt
     2281 gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga
     2341 gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca
     2401 ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg
     2461 ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt
     2521 cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc
     2581 ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct
     2641 tcgggaagcg tggcgctttc tcaatgctca cgctgtaggt atctcagttc ggtgtaggtc
     2701 gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta
     2761 tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca
     2821 gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag
     2881 tggtggccta actacggcta cactagaagg acagtatttg gtatctgcgc tctgctgaag
     2941 ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt
     3001 agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa
     3061 gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg
     3121 attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga
     3181 agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta
     3241 atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc
     3301 cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg
     3361 ataccgcgag acccacgctc accggctcca gatttatcag caataaacca gccagccgga
     3421 agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt
     3481 tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt
     3541 gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc
     3601 caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc
     3661 ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca
     3721 gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag
     3781 tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg
     3841 tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa
     3901 cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa
     3961 cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga
     4021 gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga
     4081 atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg
     4141 agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt
     4201 ccccgaaaag tgccacctga cgtctaagaa accattatta tcatgacatt aacctataaa
     4261 aataggcgta tcacgaggcc ctttcgtctc gcgcgtttcg gtgatgacgg tgaaaacctc
     4321 tgacacatgc agctcccgga gacggtcaca gcttgtctgt aagcggatgc cgggagcaga
     4381 caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc ggggctggct taactatgcg
     4441 gcatcagagc agattgtact gagagtgcac catatggaca tattgtcgtt agaacgcggc
     4501 tacaattaat acataacctt atgtatcata cacatacgat ttaggtgaca ctata
//
