LOCUS       sequence-258429-        3845 bp    DNA     circular SYN 15-APR-2025
DEFINITION  PCR-based C-terminal S6-FLAG tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3845)
  AUTHORS   Nemec AA, Tomko RJ Jr
  TITLE     A suite of PCR-based peptide tagging plasmids to permit 
            epitope-targeted enzymatic functionalization of yeast proteins.
  JOURNAL   Yeast. 2020 May 13. doi: 10.1002/yea.3471.
  PUBMED    32401365
REFERENCE   2  (bases 1 to 3845)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Yeast. 2020
            May 13. doi: 10.1002/yea.3471."
COMMENT     Sequence Label: pFA6a-S6-FLAG:HIS3MX6
FEATURES             Location/Qualifiers
     source          1..3845
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             98..121
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     gene            178..1378
                     /label=HIS3MX6
                     /note="yeast selectable marker encoding the S. pombe his5 
                     gene, which corresponds to S. cerevisiae HIS3"
     promoter        178..521
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             522..1175
                     /codon_start=1
                     /gene="Schizosaccharomyces pombe his5"
                     /product="imidazoleglycerol-phosphate dehydratase, required
                     for histidine biosynthesis"
                     /label=S. pombe his5
                     /note="yeast auxotrophic marker; corresponds to S. 
                     cerevisiae HIS3"
                     /translation="MGRRAFVERNTNETKISVAIALDKAPLPEESNFIDELITSKHANQ
                     KGEQVIQVDTGIGFLDHMYHALAKHAGWSLRLYSRGDLIIDDHHTAEDTAIALGIAFKQ
                     AMGNFAGVKRFGHAYCPLDEALSRSVVDLSGRPYAVIDLGLKREKVGELSCEMIPHLLY
                     SFSVAAGITLHVTCLYGSNDHHRAESAFKSLAVAMRAATSLTGSSEVPSTKGVL"
     terminator      1181..1378
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(1482..1501)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1483..1501)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1588..1605)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1759..2347)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1839..1858)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2518..3309,3310..3378))
                     /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     3141..3160
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3379..3483)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3551..3569
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(3607..3629)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     3729..3748
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        join(3829..3845,1..2)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     join(3829..3845,1)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
ORIGIN
        1 gaacgcggcc gccagctgaa gcttcgtacg ctgcaggtcg acggatcccc gggttaatta
       61 aggggatagc ttatcctggt tacttagact gttaaacgat tacaaggacg acgatgacaa
      121 gtaaggcgcg ccagatctgt ttagcttgcc tcgtccccgc cgggtcaccc ggccagcgac
      181 atggaggccc agaataccct ccttgacagt cttgacgtgc gcagctcagg ggcatgatgt
      241 gactgtcgcc cgtacattta gcccatacat ccccatgtat aatcatttgc atccatacat
      301 tttgatggcc gcacggcgcg aagcaaaaat tacggctcct cgctgcagac ctgcgagcag
      361 ggaaacgctc ccctcacaga cgcgttgaat tgtccccacg ccgcgcccct gtagagaaat
      421 ataaaaggtt aggatttgcc actgaggttc ttctttcata tacttccttt taaaatcttg
      481 ctaggataca gttctcacat cacatccgaa cataaacaac catgggtagg agggcttttg
      541 tagaaagaaa tacgaacgaa acgaaaatca gcgttgccat cgctttggac aaagctccct
      601 tacctgaaga gtcgaatttt attgatgaac ttataacttc caagcatgca aaccaaaagg
      661 gagaacaagt aatccaagta gacacgggaa ttggattctt ggatcacatg tatcatgcac
      721 tggctaaaca tgcaggctgg agcttacgac tttactcaag aggtgattta atcatcgatg
      781 atcatcacac tgcagaagat actgctattg cacttggtat tgcattcaag caggctatgg
      841 gtaactttgc cggcgttaaa agatttggac atgcttattg tccacttgac gaagctcttt
      901 ctagaagcgt agttgacttg tcgggacggc cctatgctgt tatcgatttg ggattaaagc
      961 gtgaaaaggt tggggaattg tcctgtgaaa tgatccctca cttactatat tccttttcgg
     1021 tagcagctgg aattactttg catgttacct gcttatatgg tagtaatgac catcatcgtg
     1081 ctgaaagcgc ttttaaatct ctggctgttg ccatgcgcgc ggctactagt cttactggaa
     1141 gttctgaagt cccaagcacg aagggagtgt tgtaaagagt actgacaata aaaagattct
     1201 tgttttcaag aacttgtcat ttgtatagtt tttttatatt gtagttgttc tattttaatc
     1261 aaatgttagc gtgatttata ttttttttcg cctcgacatc atctgcccag atgcgaagtt
     1321 aagtgcgcag aaagtaatat catgcgtcaa tcgtatgtga atgctggtcg ctatactgct
     1381 gtcgattcga tactaacgcc gccatccagt ttaaacgagc tcgaattcat cgatgatatc
     1441 agatccacta gtggcctatg cggccgcgga tctgccggtc tccctatagt gagtcgtatt
     1501 aatttcgata agccaggtta acctgcatta atgaatcggc caacgcgcgg ggagaggcgg
     1561 tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg
     1621 gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
     1681 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     1741 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     1801 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     1861 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     1921 ctttctccct tcgggaagcg tggcgctttc tcaatgctca cgctgtaggt atctcagttc
     1981 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     2041 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     2101 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     2161 gttcttgaag tggtggccta actacggcta cactagaagg acagtatttg gtatctgcgc
     2221 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     2281 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     2341 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     2401 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
     2461 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta
     2521 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     2581 tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag
     2641 tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag caataaacca
     2701 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc
     2761 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt
     2821 tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag
     2881 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt
     2941 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat
     3001 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt
     3061 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc
     3121 ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat
     3181 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
     3241 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
     3301 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
     3361 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta
     3421 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc
     3481 gcgcacattt ccccgaaaag tgccacctga cgtctaagaa accattatta tcatgacatt
     3541 aacctataaa aataggcgta tcacgaggcc ctttcgtctc gcgcgtttcg gtgatgacgg
     3601 tgaaaacctc tgacacatgc agctcccgga gacggtcaca gcttgtctgt aagcggatgc
     3661 cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc ggggctggct
     3721 taactatgcg gcatcagagc agattgtact gagagtgcac catatggaca tattgtcgtt
     3781 agaacgcggc tacaattaat acataacctt atgtatcata cacatacgat ttaggtgaca
     3841 ctata
//
