LOCUS       sequence-8997-ex        4531 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4531)
  AUTHORS   Noguchi C, Garabedian MV, Malik M, Noguchi E
  TITLE     A vector system for genomic FLAG epitope-tagging in 
            Schizosaccharomyces pombe.
  JOURNAL   Biotechnol J. 2008 Oct;3(9-10):1280-5. doi: 10.1002/biot.200800140.
  PUBMED    18729046
REFERENCE   2  (bases 1 to 4531)
  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: "Biotechnol 
            J."; date: "2008-10"; volume: "3(9-10):1280-5. doi"; pages: " 
            10.1002/biot.200800140"
COMMENT     Sequence Label: pFA6a-5FLAG-hphMX6
FEATURES             Location/Qualifiers
     source          1..4531
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             69..200
                     /codon_start=1
                     /product="five tandem FLAG(R) epitope tags"
                     /label=5xFLAG
                     /translation="DYKDDDDKDYKDDDDKLMDYKDDDDKDYKDDDDKLMDYKDDDDK"
     terminator      227..414
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(282..306)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     gene            489..2064
                     /label=hphMX6
                     /note="yeast selectable marker conferring hygromycin 
                     resistance"
     promoter        489..832
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             833..1861
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     CDS             833..1861
                     /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      1867..2064
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(2168..2187)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2169..2187)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2274..2291)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2445..3033)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2525..2544)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3204..3995,3996..4064))
                     /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     3827..3846
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4065..4169)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4237..4255
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(4293..4315)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4415..4434
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        join(4515..4531,1..2)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     join(4515..4531,1)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
ORIGIN
        1 gaacgcggcc gccagctgaa gcttcgtacg ctgcaggtcg acggatcccc gggttaatta
       61 atcatatgga ctacaaggac gacgatgaca aggattacaa agatgacgac gataagctta
      121 tggactacaa ggacgacgat gacaaggatt acaaagatga cgacgataag cttatggact
      181 acaaggacga cgatgacaag catatgggcg cgccacttct aaataagcga atttcttatg
      241 atttatgatt tttattatta aataagttat aaaaaaaata agtgtataca aattttaaag
      301 tgactcttag gttttaaaac gaaaattctt attcttgagt aactctttcc tgtaggtcag
      361 gttgctttct caggtatagt atgaggtcgc tcttattgac cacacctcta ccggcagatc
      421 cgctagggat aacagggtaa tatagatctg tttagcttgc ctcgtccccg ccgggtcacc
      481 cggccagcga catggaggcc cagaataccc tccttgacag tcttgacgtg cgcagctcag
      541 gggcatgatg tgactgtcgc ccgtacattt agcccataca tccccatgta taatcatttg
      601 catccataca ttttgatggc cgcacggcgc gaagcaaaaa ttacggctcc tcgctgcaga
      661 cctgcgagca gggaaacgct cccctcacag acgcgttgaa ttgtccccac gccgcgcccc
      721 tgtagagaaa tataaaaggt taggatttgc cactgaggtt cttctttcat atacttcctt
      781 ttaaaatctt gctaggatac agttctcaca tcacatccga acataaacaa ccatgggtaa
      841 aaagcctgaa ctcaccgcga cgtctgtcga gaagtttctg atcgaaaagt tcgacagcgt
      901 ctccgacctg atgcagctct cggagggcga agaatctcgt gctttcagct tcgatgtagg
      961 agggcgtgga tatgtcctgc gggtaaatag ctgcgccgat ggtttctaca aagatcgtta
     1021 tgtttatcgg cactttgcat cggccgcgct cccgattccg gaagtgcttg acattgggga
     1081 attcagcgag agcctgacct attgcatctc ccgccgtgca cagggtgtca cgttgcaaga
     1141 cctgcctgaa accgaactgc ccgctgttct gcagccggtc gcggaggcca tggatgcgat
     1201 cgctgcggcc gatcttagcc agacgagcgg gttcggccca ttcggaccgc aaggaatcgg
     1261 tcaatacact acatggcgtg atttcatatg cgcgattgct gatccccatg tgtatcactg
     1321 gcaaactgtg atggacgaca ccgtcagtgc gtccgtcgcg caggctctcg atgagctgat
     1381 gctttgggcc gaggactgcc ccgaagtccg gcacctcgtg cacgcggatt tcggctccaa
     1441 caatgtcctg acggacaatg gccgcataac agcggtcatt gactggagcg aggcgatgtt
     1501 cggggattcc caatacgagg tcgccaacat cttcttctgg aggccgtggt tggcttgtat
     1561 ggagcagcag acgcgctact tcgagcggag gcatccggag cttgcaggat cgccgcggct
     1621 ccgggcgtat atgctccgca ttggtcttga ccaactctat cagagcttgg ttgacggcaa
     1681 tttcgatgat gcagcttggg cgcagggtcg atgcgacgca atcgtccgat ccggagccgg
     1741 gactgtcggg cgtacacaaa tcgcccgcag aagcgcggcc gtctggaccg atggctgtgt
     1801 agaagtactc gccgatagtg gaaaccgacg ccccagcact cgtccgaggg caaaggaata
     1861 atcagtactg acaataaaaa gattcttgtt ttcaagaact tgtcatttgt atagtttttt
     1921 tatattgtag ttgttctatt ttaatcaaat gttagcgtga tttatatttt ttttcgcctc
     1981 gacatcatct gcccagatgc gaagttaagt gcgcagaaag taatatcatg cgtcaatcgt
     2041 atgtgaatgc tggtcgctat actgctgtcg attcgatact aacgccgcca tccagtttaa
     2101 acgagctcga attcatcgat gatatcagat ccactagtgg cctatgcggc cgcggatctg
     2161 ccggtctccc tatagtgagt cgtattaatt tcgataagcc aggttaacct gcattaatga
     2221 atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc
     2281 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg
     2341 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc
     2401 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     2461 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     2521 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     2581 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcaa
     2641 tgctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     2701 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     2761 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
     2821 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     2881 agaaggacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     2941 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     3001 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     3061 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     3121 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
     3181 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
     3241 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
     3301 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
     3361 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
     3421 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
     3481 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
     3541 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
     3601 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
     3661 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
     3721 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
     3781 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
     3841 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
     3901 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
     3961 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
     4021 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
     4081 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
     4141 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc
     4201 taagaaacca ttattatcat gacattaacc tataaaaata ggcgtatcac gaggcccttt
     4261 cgtctcgcgc gtttcggtga tgacggtgaa aacctctgac acatgcagct cccggagacg
     4321 gtcacagctt gtctgtaagc ggatgccggg agcagacaag cccgtcaggg cgcgtcagcg
     4381 ggtgttggcg ggtgtcgggg ctggcttaac tatgcggcat cagagcagat tgtactgaga
     4441 gtgcaccata tggacatatt gtcgttagaa cgcggctaca attaatacat aaccttatgt
     4501 atcatacaca tacgatttag gtgacactat a
//
