LOCUS       sequence-271104-        3981 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  PCR-based C-terminal SRS-FLAG tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3981)
  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 3981)
  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: "Yeast. 2020
            May 13. doi: 10.1002/yea.3471."
COMMENT     Sequence Label: pFA6a-SRS-FLAG:natMX4
FEATURES             Location/Qualifiers
     source          1..3981
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     28..47
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        128..146
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     128..145
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     CDS             227..250
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     terminator      274..461
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(329..353)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     gene            536..1655
                     /label=natMX6
                     /note="yeast selectable marker conferring nourseothricin 
                     resistance"
     promoter        536..879
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             880..1452
                     /codon_start=1
                     /gene="Streptomyces noursei nat1"
                     /product="nourseothricin acetyltransferase"
                     /label=NrsR
                     /note="confers resistance to nourseothricin"
                     /translation="MGTTLDDTAYRYRTSVPGDAEAIEALDGSFTTDTVFRVTATGDGF
                     TLREVPVDPPLTKVFPDDESDDESDDGEDGDPDSRTFVAYGDDGDLAGFVVVSYSGWNR
                     RLTVEDIEVAPEHRGHGVGRALMGLATEFARERGAGHLWLEVTNVNAPAIHAYRRMGFT
                     LCGLDTALYDGTASDGEQALYMSMPCP"
     terminator      1458..1655
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(1762..1781)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1763..1781)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1868..1885)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2039..2627)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2119..2138)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2798..3589,3590..3658))
                     /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     3421..3440
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3659..3763)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3831..3849
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(3887..3909)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 gcgggtgtcg gggctggctt aactatgcgg catcagagca gattgtactg agagtgcacc
       61 atatggacat attgtcgtta gaacgcggct acaattaata cataacctta tgtatcatac
      121 acatacgatt taggtgacac tatagaacgc ggccgccagc tgaagcttcg tacgctgcag
      181 gtcgacggat ccccgggtta attaactgcc cgaaaccggt ggtggcgatt acaaggacga
      241 cgatgacaag taaggcgcgc cacttctaaa taagcgaatt tcttatgatt tatgattttt
      301 attattaaat aagttataaa aaaaataagt gtatacaaat tttaaagtga ctcttaggtt
      361 ttaaaacgaa aattcttatt cttgagtaac tctttcctgt aggtcaggtt gctttctcag
      421 gtatagtatg aggtcgctct tattgaccac acctctaccg gcagatccgc tagggataac
      481 agggtaatat agatctgttt agcttgcctc gtccccgccg ggtcacccgg ccagcgacat
      541 ggaggcccag aataccctcc ttgacagtct tgacgtgcgc agctcagggg catgatgtga
      601 ctgtcgcccg tacatttagc ccatacatcc ccatgtataa tcatttgcat ccatacattt
      661 tgatggccgc acggcgcgaa gcaaaaatta cggctcctcg ctgcagacct gcgagcaggg
      721 aaacgctccc ctcacagacg cgttgaattg tccccacgcc gcgcccctgt agagaaatat
      781 aaaaggttag gatttgccac tgaggttctt ctttcatata cttcctttta aaatcttgct
      841 aggatacagt tctcacatca catccgaaca taaacaacca tgggtaccac tcttgacgac
      901 acggcttacc ggtaccgcac cagtgtcccg ggggacgccg aggccatcga ggcactggat
      961 gggtccttca ccaccgacac cgtcttccgc gtcaccgcca ccggggacgg cttcaccctg
     1021 cgggaggtgc cggtggaccc gcccctgacc aaggtgttcc ccgacgacga atcggacgac
     1081 gaatcggacg acggggagga cggcgacccg gactcccgga cgttcgtcgc gtacggggac
     1141 gacggcgacc tggcgggctt cgtggtcgtc tcgtactccg gctggaaccg ccggctgacc
     1201 gtcgaggaca tcgaggtcgc cccggagcac cgggggcacg gggtcgggcg cgcgttgatg
     1261 gggctcgcga cggagttcgc ccgcgagcgg ggcgccgggc acctctggct ggaggtcacc
     1321 aacgtcaacg caccggcgat ccacgcgtac cggcggatgg ggttcaccct ctgcggcctg
     1381 gacaccgccc tgtacgacgg caccgcctcg gacggcgagc aggcgctcta catgagcatg
     1441 ccctgcccct aatcagtact gacaataaaa agattcttgt tttcaagaac ttgtcatttg
     1501 tatagttttt ttatattgta gttgttctat tttaatcaaa tgttagcgtg atttatattt
     1561 tttttcgcct cgacatcatc tgcccagatg cgaagttaag tgcgcagaaa gtaatatcat
     1621 gcgtcaatcg tatgtgaatg ctggtcgcta tactgctgtc gattcgatac taacgccgcc
     1681 atccagtgtc gaaaacgagc tcgaattcat cgatgatatc agatccacta gtggcctatg
     1741 cggccgcgga tctgccggtc tccctatagt gagtcgtatt aatttcgata agccaggtta
     1801 acctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt
     1861 ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag
     1921 ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca
     1981 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
     2041 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
     2101 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
     2161 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
     2221 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
     2281 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
     2341 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
     2401 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta
     2461 actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct
     2521 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt
     2581 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga
     2641 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca
     2701 tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat
     2761 caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg
     2821 cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt
     2881 agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag
     2941 acccacgctc accggctcca gatttatcag caataaacca gccagccgga agggccgagc
     3001 gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag
     3061 ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca
     3121 tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa
     3181 ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga
     3241 tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata
     3301 attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca
     3361 agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg
     3421 ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg
     3481 ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg
     3541 cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag
     3601 gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac
     3661 tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca
     3721 tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag
     3781 tgccacctga cgtctaagaa accattatta tcatgacatt aacctataaa aataggcgta
     3841 tcacgaggcc ctttcgtctc gcgcgtttcg gtgatgacgg tgaaaacctc tgacacatgc
     3901 agctcccgga gacggtcaca gcttgtctgt aagcggatgc cgggagcaga caagcccgtc
     3961 agggcgcgtc agcgggtgtt g
//
