LOCUS       sequence-9647-ex        4477 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 4477)
  AUTHORS   Funakoshi M, Hochstrasser M
  TITLE     Small epitope-linker modules for PCR-based C-terminal tagging in 
            Saccharomyces cerevisiae.
  JOURNAL   Yeast. 2009 Feb 25. 26(3):185-192.
  PUBMED    19243080
REFERENCE   2  (bases 1 to 4477)
  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. 2009
            Feb 25. 26(3):185-192."
COMMENT     Sequence Label: pFA6a-6xGLY-3xFLAG-hphMX4
FEATURES             Location/Qualifiers
     source          1..4477
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             79..144
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     terminator      168..355
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(223..247)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     gene            430..2005
                     /label=hphMX6
                     /note="yeast selectable marker conferring hygromycin 
                     resistance"
     promoter        430..773
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             774..1802
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     CDS             774..1802
                     /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      1808..2005
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(2112..2131)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2113..2131)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2218..2235)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2389..2977)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2469..2488)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3148..3939,3940..4008))
                     /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     3771..3790
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4009..4113)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4181..4199
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(4237..4259)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4359..4378
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        4459..4477
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     4459..4476
                     /label=SP6
                     /note="SP6 promoter, forward primer"
ORIGIN
        1 acgcggccgc cagctgaagc ttcgtacgct gcaggtcgac ggatccccgg gttaattaac
       61 gggggaggcg ggggtggaga ctacaaagac catgacggtg attataaaga tcatgacatc
      121 gactacaagg atgacgatga caagtagggc gcgccacttc taaataagcg aatttcttat
      181 gatttatgat ttttattatt aaataagtta taaaaaaaat aagtgtatac aaattttaaa
      241 gtgactctta ggttttaaaa cgaaaattct tattcttgag taactctttc ctgtaggtca
      301 ggttgctttc tcaggtatag tatgaggtcg ctcttattga ccacacctct accggcagat
      361 ccgctaggga taacagggta atatagatct gtttagcttg ccttgtcccc gccgggtcac
      421 ccggccagcg acatggaggc ccagaatacc ctccttgaca gtcttgacgt gcgcagctca
      481 ggggcatgat gtgactgtcg cccgtacatt tagcccatac atccccatgt ataatcattt
      541 gcatccatac attttgatgg ccgcacggcg cgaagcaaaa attacggctc ctcgctgcag
      601 acctgcgagc agggaaacgc tcccctcaca gacgcgttga attgtcccca cgccgcgccc
      661 ctgtagagaa atataaaagg ttaggatttg ccactgaggt tcttctttca tatacttcct
      721 tttaaaatct tgctaggata cagttctcac atcacatccg aacataaaca accatgggta
      781 aaaagcctga actcaccgcg acgtctgtcg agaagtttct gatcgaaaag ttcgacagcg
      841 tctccgacct gatgcagctc tcggagggcg aagaatctcg tgctttcagc ttcgatgtag
      901 gagggcgtgg atatgtcctg cgggtaaata gctgcgccga tggtttctac aaagatcgtt
      961 atgtttatcg gcactttgca tcggccgcgc tcccgattcc ggaagtgctt gacattgggg
     1021 aattcagcga gagcctgacc tattgcatct cccgccgtgc acagggtgtc acgttgcaag
     1081 acctgcctga aaccgaactg cccgctgttc tgcagccggt cgcggaggcc atggatgcga
     1141 tcgctgcggc cgatcttagc cagacgagcg ggttcggccc attcggaccg caaggaatcg
     1201 gtcaatacac tacatggcgt gatttcatat gcgcgattgc tgatccccat gtgtatcact
     1261 ggcaaactgt gatggacgac accgtcagtg cgtccgtcgc gcaggctctc gatgagctga
     1321 tgctttgggc cgaggactgc cccgaagtcc ggcacctcgt gcacgcggat ttcggctcca
     1381 acaatgtcct gacggacaat ggccgcataa cagcggtcat tgactggagc gaggcgatgt
     1441 tcggggattc ccaatacgag gtcgccaaca tcttcttctg gaggccgtgg ttggcttgta
     1501 tggagcagca gacgcgctac ttcgagcgga ggcatccgga gcttgcagga tcgccgcggc
     1561 tccgggcgta tatgctccgc attggtcttg accaactcta tcagagcttg gttgacggca
     1621 atttcgatga tgcagcttgg gcgcagggtc gatgcgacgc aatcgtccga tccggagccg
     1681 ggactgtcgg gcgtacacaa atcgcccgca gaagcgcggc cgtctggacc gatggctgtg
     1741 tagaagtact cgccgatagt ggaaaccgac gccccagcac tcgtccgagg gcaaaggaat
     1801 aatcagtact gacaataaaa agattcttgt tttcaagaac ttgtcatttg tatagttttt
     1861 ttatattgta gttgttctat tttaatcaaa tgttagcgtg atttatattt tttttcgcct
     1921 cgacatcatc tgcccagatg cgaagttaag tgcgcagaaa gtaatatcat gcgtcaatcg
     1981 tatgtgaatg ctggtcgcta tactgctgtc gattcgatac taacgccgcc atccagtgtc
     2041 gaaaacgagc tcgaattcat cgatgatatc agatccacta gtggcctatg cggccgcgga
     2101 tctgccggtc tccctatagt gagtcgtatt aatttcgata agccaggtta acctgcatta
     2161 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc
     2221 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa
     2281 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
     2341 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     2401 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     2461 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     2521 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     2581 tcaatgctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     2641 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     2701 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     2761 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     2821 cactagaagg acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
     2881 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
     2941 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
     3001 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
     3061 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag
     3121 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc
     3181 agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac
     3241 gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc
     3301 accggctcca gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg
     3361 tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag
     3421 tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc
     3481 acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac
     3541 atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag
     3601 aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac
     3661 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg
     3721 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc
     3781 gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact
     3841 ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg
     3901 atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa
     3961 tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt
     4021 tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg
     4081 tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga
     4141 cgtctaagaa accattatta tcatgacatt aacctataaa aataggcgta tcacgaggcc
     4201 ctttcgtctc gcgcgtttcg gtgatgacgg tgaaaacctc tgacacatgc agctcccgga
     4261 gacggtcaca gcttgtctgt aagcggatgc cgggagcaga caagcccgtc agggcgcgtc
     4321 agcgggtgtt ggcgggtgtc ggggctggct taactatgcg gcatcagagc agattgtact
     4381 gagagtgcac catatggaca tattgtcgtt agaacgcggc tacaattaat acataacctt
     4441 atgtatcata cacatacgat ttaggtgaca ctataga
//
