LOCUS       sequence-9000-ex        5177 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 5177)
  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 5177)
  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-natMX6-P3nmt1-3FLAG
FEATURES             Location/Qualifiers
     source          1..5177
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(1..19)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2..19)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(100..119)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     219..241
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(279..297)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        365..469
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(470..538,539..1330)
                     /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     complement(688..707)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1501..2089
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1990..2009
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2243..2260
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     2347..2366
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2347..2365
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     gene            complement(2470..3592)
                     /label=natMX6
                     /note="yeast selectable marker conferring nourseothricin 
                     resistance"
     terminator      complement(2470..2667)
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     CDS             complement(2673..3248)
                     /codon_start=1
                     /gene="Streptomyces noursei nat1"
                     /product="nourseothricin acetyltransferase"
                     /label=NrsR
                     /note="confers resistance to nourseothricin"
                     /translation="MGTTLDDTAYRYRTSVPGDAEAIEALDGSFTTDTVFRVTATGDGF
                     TLREVPVDPPLTKVFPDDESDDESDDGEDGDPDSRTFVAYGDDGDLAGFVVISYSAWNR
                     RLTVEDIEVAPEHRGHGVGRALMGLATEFAGERGAGHLWLEVTNVNAPAIHAYRRMGFT
                     LCGLDTALYDGTASDGERQALYMSMPCP"
     promoter        complement(3249..3592)
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     promoter        3637..4802
                     /label=nmt1 promoter
                     /note="wild-type nmt1 promoter from Schizosaccharomyces 
                     pombe, conferring strong thiamine-repressible expression"
     CDS             4820..4891
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags"
                     /label=3xFLAG
                     /translation="DYKDDDDKDYKDDDDKDYKDDDDK"
     terminator      4915..5102
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(4970..4994)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
ORIGIN
        1 tctatagtgt cacctaaatc gtatgtgtat gatacataag gttatgtatt aattgtagcc
       61 gcgttctaac gacaatatgt ccatatggtg cactctcagt acaatctgct ctgatgccgc
      121 atagttaagc cagccccgac acccgccaac acccgctgac gcgccctgac gggcttgtct
      181 gctcccggca tccgcttaca gacaagctgt gaccgtctcc gggagctgca tgtgtcagag
      241 gttttcaccg tcatcaccga aacgcgcgag acgaaagggc ctcgtgatac gcctattttt
      301 ataggttaat gtcatgataa taatggtttc ttagacgtca ggtggcactt ttcggggaaa
      361 tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat
      421 gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca
      481 acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca
      541 cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta
      601 catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt
      661 tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc
      721 cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc
      781 accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc
      841 cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa
      901 ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga
      961 accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat
     1021 ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca
     1081 attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc
     1141 ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat
     1201 tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag
     1261 tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa
     1321 gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca
     1381 tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc
     1441 ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
     1501 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
     1561 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
     1621 cagcagagcg cagataccaa atactgtcct tctagtgtag ccgtagttag gccaccactt
     1681 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
     1741 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
     1801 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
     1861 ctacaccgaa ctgagatacc tacagcgtga gcattgagaa agcgccacgc ttcccgaagg
     1921 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
     1981 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
     2041 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
     2101 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
     2161 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
     2221 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat
     2281 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcaggttaa cctggcttat
     2341 cgaaattaat acgactcact atagggagac cggcagatcc gcggccgcat aggccactag
     2401 tggatctgat atcatcgatg aattcgagct cgtttaaact ggatggcggc gttagtatcg
     2461 aatcgacagc agtatagcga ccagcattca catacgattg acgcatgata ttactttctg
     2521 cgcacttaac ttcgcatctg ggcagatgat gtcgaggcga aaaaaaatat aaatcacgct
     2581 aacatttgat taaaatagaa caactacaat ataaaaaaac tatacaaatg acaagttctt
     2641 gaaaacaaga atctttttat tgtcagtact gattaggggc agggcatgct catgtagagc
     2701 gcctgccgct cgccgtccga ggcggtgccg tcgtacaggg cggtgtccag gccgcagagg
     2761 gtgaacccca tccgccggta cgcgtggatc gccggtgcgt tgacgttggt gacctccagc
     2821 cagaggtgcc cggcgccccg ctcgccggcg aactccgtcg cgagccccat caacgcgcgc
     2881 ccgaccccgt gcccccggtg ctccggggcg acctcgatgt cctcgacggt cagccggcgg
     2941 ttccacgccg agtacgagat gaccacgaag cccgccaggt cgccgtcgtc cccgtacgcg
     3001 acgaacgtcc gggagtccgg gtcgccgtcc tccccgtcgt ccgattcgtc gtccgattcg
     3061 tcgtcgggga acaccttggt caggggcggg tccaccggca cctcccgcag ggtgaagccg
     3121 tccccggtgg cggtgacgcg gaagacggtg tcggtggtga aggacccatc cagtgcctcg
     3181 atggcctcgg cgtcccccgg gacactggtg cggtaccggt aagccgtgtc gtcaagagtg
     3241 gtacccatgg ttgtttatgt tcggatgtga tgtgagaact gtatcctagc aagattttaa
     3301 aaggaagtat atgaaagaag aacctcagtg gcaaatccta accttttata tttctctaca
     3361 ggggcgcggc gtggggacaa ttcaacgcgt ctgtgagggg agcgtttccc tgctcgcagg
     3421 tctgcagcga ggagccgtaa tttttgcttc gcgccgtgcg gccatcaaaa tgtatggatg
     3481 caaatgatta tacatgggga tgtatgggct aaatgtacgg gcgacagtca catcatgccc
     3541 ctgagctgcg cacgtcaaga ctgtcaagga gggtattctg ggcctccatg tcgctggccg
     3601 ggtgacccgg cggggacgag gcaagctaaa cagatctcgc cataaaagac agaataagtc
     3661 atcagcggtt gtttcatttc ctatattttt tttttatttt tttatttttt aataagggaa
     3721 aatttaacgt ctaaggatac agaagattgt tagcacatta aagtaataaa ggcttaagta
     3781 gtaagtgcct tagcatgtta ttgtatttca aaggacataa tctaaaataa taacaatatc
     3841 atttctcaca agttattcaa ttttcttttt tttttctaat aatatcaaga atgtattatt
     3901 tgtttgacat aagtcaacta atttatttaa tatgctggat taatcttgca gacatgtaaa
     3961 ttaacaagtt ttagtcaaat aacgttgaag tttcaatgaa ctcaaataat ttctcttttt
     4021 ttttatataa ccatatatgt ctaatctgat ttatattttc cgcaggatca actgaagtta
     4081 tgacatttgg attggatcac ttataacctt ggtcgccaaa taatacaaaa atcagcgtta
     4141 taaaacaaag aaggtttttg ttaagaaatt aatcctcttt cttgataaga aagttgaacc
     4201 gaaattgcag atactgatat atgaaaataa tacccacaat tttgggaata gcgcaagcct
     4261 caatttaaac aataggtgag gacacatgat aatgacctca atgattgtta gaagaaaaga
     4321 gcctcattac aaaatcgaaa aatgaatggt tgggtacaag tttccaaaac atggtaaagt
     4381 ggactttgcg tatgagacgt aaatagaaaa aaacacttgt tatatgtttt ctagaattat
     4441 tgttgtctct ttatggttgg atgatgcaaa atagtaattt cggttagttg ctgtaaaaca
     4501 ccacgagaca aatagatatg gatatttatt aaatcaggaa aaacgtaact ctcggctact
     4561 ggatggttca gtcacccaac gattactggg gagagaaaac agggcaaaag caaagcttaa
     4621 aggaatccga ttgtcattcg gcaatgtgca gcgaaactaa aaaccggata atggacctgt
     4681 taatcgaaac attgaagata tataaaggaa gaggaatcct ggcatatcat caattgaata
     4741 agttgaatta attatttcaa tctcattctc actttctgac ttatagtcgc tttgttaaat
     4801 catgtcttta attaatatgg actacaagga cgacgatgac aaggattaca aagatgacga
     4861 cgataaggac tataaggacg atgatgataa atagggcgcg ccacttctaa ataagcgaat
     4921 ttcttatgat ttatgatttt tattattaaa taagttataa aaaaaataag tgtatacaaa
     4981 ttttaaagtg actcttaggt tttaaaacga aaattcttat tcttgagtaa ctctttcctg
     5041 taggtcaggt tgctttctca ggtatagtat gaggtcgctc ttattgacca cacctctacc
     5101 ggcagatccg ctagggataa cagggtaata tagatccgtc gacctgcagc gtacgaagct
     5161 tcagctggcg gccgcgt
//
