LOCUS       sequence-8995-ex        5992 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 5992)
  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 5992)
  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-his3-P41nmt1-3FLAG
FEATURES             Location/Qualifiers
     source          1..5992
                     /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"
     CDS             complement(join(2738..3200,3260..3871,4057..4068,
                     4124..4191))
                     /codon_start=1
                     /gene="Schizosaccharomyces pombe his3"
                     /product="imidazole acetol phosphate transaminase, required
                     for histidine biosynthesis"
                     /label=S. pombe his3
                     /translation="MFDLNTCLRKNILELQPYRCARDDFSEGVLLDANECAYGSVISVD
                     GVEFNRYPDPRQIEVKQRLCDLRNKELSITKPLTPDNICMGVGSDEIIDSLIRISCIPG
                     KDKILMCPPSYGMYTVSAKINDVEVVKVLLEPDFNLNVDAICETLSKDSAIKVFFACSP
                     GNPTAKALKLEDIKKILEHPTWNGIVVVDEAYIDFSAPDMSALTLVNEYPNLAVCQTLS
                     KSFGLAGIRIGFCLTSKPIATIMNSLKAPYNISEPTSRLALDALSPQSIDKMHTYRDAI
                     IQQRVRLCKELTTIKGMGKIIGGYDANFILIQVLDRPEGGKPSNDAAKYLYLQMATMHK
                     VVVRFRGTEPLCEGALRITVGTEEENTILLKTIKLVLQEYYTKK"
     promoter        4456..5617
                     /label=nmt1 P41 promoter
                     /note="mutant nmt1 promoter from Schizosaccharomyces pombe,
                     conferring medium strength thiamine-repressible expression"
     CDS             5635..5706
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags"
                     /label=3xFLAG
                     /translation="DYKDDDDKDYKDDDDKDYKDDDDK"
     terminator      5730..5917
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(5785..5809)
                     /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 ctatgcaaag ctaacgaatc
     2461 tttaattctt ttattacacg ggttataatc ctttaaatta gcgtaactcc tttacaaaat
     2521 taaaaataac atcttttcat tgcttatcga actatacgga aacacaccaa aaaggaagaa
     2581 aacgtatttt aataaagaaa gaaggaatgc aagcgcaatt agcgaacaca aaatatattg
     2641 ggtacatcag aaaagaaatg tgacgagtcc ggaaggggaa aggaaagagt tataatttgt
     2701 ctaattgaaa cgttctaata ggccagaaaa ggaaacctca tttttttgta tagtattcct
     2761 gcaatacaag tttaatagtt tttagtaata tcgtattctc ttcttctgtg ccaacagtga
     2821 tacgcaatgc tccttcgcaa agaggctctg tacctctaaa tctaacaacg actttgtgca
     2881 tcgtggccat ttgtaaataa agatattttg cggcatcgtt actaggctta cctccttctg
     2941 gtctatctaa aacctgaatg aggataaaat ttgcatcata accacctata atcttgccca
     3001 ttcctttaat ggtagtaagc tctttgcaaa gacgaactct ttgctgtata atagcatctc
     3061 tatatgtatg cattttatca atcgactgag gggataaagc atccaaagct aaacgagagg
     3121 tgggttcgct aatgttataa ggagccttta atgagttcat aatggtagca ataggtttac
     3181 ttgtcaaaca aaatccgata ctaaacatca tagttagtaa agaaataata ttaatgctta
     3241 aaagcttaga ttaacttacc gaattcctgc tagaccgaaa gatttcgaaa gggtttggca
     3301 aacagcaagg tttggatatt cattgacaag agttaaggca gacatatcgg gagcagagaa
     3361 atcaatatag gcctcatcga cgacaacgat tccattccat gtagggtgct ccaaaatctt
     3421 cttaatatcc tcgagtttaa gagccttagc tgttggatta ccaggggaac aagcaaaaaa
     3481 gactttgatg gcactatctt tagaaagtgt ttcgcaaata gcatcgacgt tcaaattgaa
     3541 atctggttca aggagaacct taacaacctc aacatcgttg atttttgcag agaccgtata
     3601 cattccgtac gaaggtggac acatcaaaat cttatctttt ccaggaatac aagaaatgcg
     3661 aattaaggag tcaataattt catcgctgcc aacacccatg caaatgttat ctggtgttag
     3721 tggcttggta atggaaagct ctttgtttct caaatcacat aatctttgct ttacttcgat
     3781 ctgtctagga tccggataac gattgaattc cacaccgtcc acggaaataa ccgaaccata
     3841 agcacattca ttggcatcaa gaaggacacc tctattaaga ttagcaaaaa atatataggc
     3901 gagaaaaggg taaaaaacaa caaaggcctg aaatcgccag ctccaacaac aacagtaaac
     3961 agggagtact tcattgcttt ccaagcagct aataaaatat ctttctagaa gtagccctca
     4021 tatcaattat tacccaaata taaagataaa acatactcag agaagtcact atattctatg
     4081 ttaatatata tcacgctttc tagtttatca agttattaca tactctctcg cacagcgata
     4141 aggctgaagt tctaagatat ttttacgaag acaagtattc aaatcaaaca tcgtatctct
     4201 ttcgatgtgt ttttgttaaa gtaagaagaa agcgatcgat gagattaatt aataattaac
     4261 ctaagatgca ggtgatatct ataaatatat agtgtgtagc aatgtgagtc tttaattcat
     4321 tacaacgtgt atcaccgtgt cattatatcc aataaattca gaaacctgaa gaggaaagag
     4381 aaataaagct ccatatgtct ttcatttcca aatacaatta taagtgcaat agtaaagaaa
     4441 acgttgaaag agatctcgcc ataaaagaca gaataagtca tcagcggttg tttcatttcc
     4501 tatatttttt ttttattttt ttatttttta ataagggaaa atttaacgtc taaggataca
     4561 gaagattgtt agcacattaa agtaataaag gcttaagtag taagtgcctt agcatgttat
     4621 tgtatttcaa aggacataat ctaaaataat aacaatatca tttctcacaa gttattcaat
     4681 tttctttttt ttttctaata atatcaagaa tgtattattt gtttgacata agtcaactaa
     4741 tttatttaat atgctggatt aatcttgcag acatgtaaat taacaagttt tagtcaaata
     4801 acgttgaagt ttcaatgaac tcaaataatt tctctttttt tttatataac catatatgtc
     4861 taatctgatt tatattttcc gcaggatcaa ctgaagttat gacatttgga ttggatcact
     4921 tataaccttg gtcgccaaat aatacaaaaa tcagcgttat aaaacaaaga aggtttttgt
     4981 taagaaatta atcctctttc ttgataagaa agttgaaccg aaattgcaga tactgatata
     5041 tgaaaataat acccacaatt ttgggaatag cgcaagcctc aatttaaaca ataggtgagg
     5101 acacatgata atgacctcaa tgattgttag aagaaaagag cctcattaca aaatcgaaaa
     5161 atgaatggtt gggtacaagt ttccaaaaca tggtaaagtg gactttgcgt atgagacgta
     5221 aatagaaaaa aacacttgtt atatgttttc tagaattatt gttgtctctt tatggttgga
     5281 tgatgcaaaa tagtaatttc ggttagttgc tgtaaaacac cacgagacaa atagatatgg
     5341 atatttatta aatcaggaaa aacgtaactc tcggctactg gatggttcag tcacccaacg
     5401 attactgggg agagaaaaca gggcaaaagc aaagcttaaa ggaatccgat tgtcattcgg
     5461 caatgtgcag cgaaactaaa aaccggataa tggacctgtt aatcgaaaca ttgaagataa
     5521 aggaagagga atcctggcat atcatcaatt gaataagttg aattaattat ttcaatctca
     5581 ttctcacttt ctgacttata gtcgctttgt taaatcatgt ctttaattaa tatggactac
     5641 aaggacgacg atgacaagga ttacaaagat gacgacgata aggactataa ggacgatgat
     5701 gataaatagg gcgcgccact tctaaataag cgaatttctt atgatttatg atttttatta
     5761 ttaaataagt tataaaaaaa ataagtgtat acaaatttta aagtgactct taggttttaa
     5821 aacgaaaatt cttattcttg agtaactctt tcctgtaggt caggttgctt tctcaggtat
     5881 agtatgaggt cgctcttatt gaccacacct ctaccggcag atccgctagg gataacaggg
     5941 taatatagat ccgtcgacct gcagcgtacg aagcttcagc tggcggccgc gt
//
