LOCUS       sequence-9653-ex        4444 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 4444)
  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 4444)
  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-T7-hphMX4
FEATURES             Location/Qualifiers
     source          1..4444
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             79..111
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="MASMTGGQQMG"
     terminator      135..322
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(190..214)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     gene            397..1972
                     /label=hphMX6
                     /note="yeast selectable marker conferring hygromycin 
                     resistance"
     promoter        397..740
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             741..1769
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     CDS             741..1769
                     /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      1775..1972
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(2079..2098)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2080..2098)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2185..2202)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2356..2944)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2436..2455)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3115..3906,3907..3975))
                     /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     3738..3757
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3976..4080)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4148..4166
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(4204..4226)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4326..4345
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        4426..4444
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     4426..4443
                     /label=SP6
                     /note="SP6 promoter, forward primer"
ORIGIN
        1 acgcggccgc cagctgaagc ttcgtacgct gcaggtcgac ggatccccgg gttaattaac
       61 gggggaggcg ggggtggaat ggctagcatg actggtggac agcaaatggg ttagggcgcg
      121 ccacttctaa ataagcgaat ttcttatgat ttatgatttt tattattaaa taagttataa
      181 aaaaaataag tgtatacaaa ttttaaagtg actcttaggt tttaaaacga aaattcttat
      241 tcttgagtaa ctctttcctg taggtcaggt tgctttctca ggtatagtat gaggtcgctc
      301 ttattgacca cacctctacc ggcagatccg ctagggataa cagggtaata tagatctgtt
      361 tagcttgcct tgtccccgcc gggtcacccg gccagcgaca tggaggccca gaataccctc
      421 cttgacagtc ttgacgtgcg cagctcaggg gcatgatgtg actgtcgccc gtacatttag
      481 cccatacatc cccatgtata atcatttgca tccatacatt ttgatggccg cacggcgcga
      541 agcaaaaatt acggctcctc gctgcagacc tgcgagcagg gaaacgctcc cctcacagac
      601 gcgttgaatt gtccccacgc cgcgcccctg tagagaaata taaaaggtta ggatttgcca
      661 ctgaggttct tctttcatat acttcctttt aaaatcttgc taggatacag ttctcacatc
      721 acatccgaac ataaacaacc atgggtaaaa agcctgaact caccgcgacg tctgtcgaga
      781 agtttctgat cgaaaagttc gacagcgtct ccgacctgat gcagctctcg gagggcgaag
      841 aatctcgtgc tttcagcttc gatgtaggag ggcgtggata tgtcctgcgg gtaaatagct
      901 gcgccgatgg tttctacaaa gatcgttatg tttatcggca ctttgcatcg gccgcgctcc
      961 cgattccgga agtgcttgac attggggaat tcagcgagag cctgacctat tgcatctccc
     1021 gccgtgcaca gggtgtcacg ttgcaagacc tgcctgaaac cgaactgccc gctgttctgc
     1081 agccggtcgc ggaggccatg gatgcgatcg ctgcggccga tcttagccag acgagcgggt
     1141 tcggcccatt cggaccgcaa ggaatcggtc aatacactac atggcgtgat ttcatatgcg
     1201 cgattgctga tccccatgtg tatcactggc aaactgtgat ggacgacacc gtcagtgcgt
     1261 ccgtcgcgca ggctctcgat gagctgatgc tttgggccga ggactgcccc gaagtccggc
     1321 acctcgtgca cgcggatttc ggctccaaca atgtcctgac ggacaatggc cgcataacag
     1381 cggtcattga ctggagcgag gcgatgttcg gggattccca atacgaggtc gccaacatct
     1441 tcttctggag gccgtggttg gcttgtatgg agcagcagac gcgctacttc gagcggaggc
     1501 atccggagct tgcaggatcg ccgcggctcc gggcgtatat gctccgcatt ggtcttgacc
     1561 aactctatca gagcttggtt gacggcaatt tcgatgatgc agcttgggcg cagggtcgat
     1621 gcgacgcaat cgtccgatcc ggagccggga ctgtcgggcg tacacaaatc gcccgcagaa
     1681 gcgcggccgt ctggaccgat ggctgtgtag aagtactcgc cgatagtgga aaccgacgcc
     1741 ccagcactcg tccgagggca aaggaataat cagtactgac aataaaaaga ttcttgtttt
     1801 caagaacttg tcatttgtat agttttttta tattgtagtt gttctatttt aatcaaatgt
     1861 tagcgtgatt tatatttttt ttcgcctcga catcatctgc ccagatgcga agttaagtgc
     1921 gcagaaagta atatcatgcg tcaatcgtat gtgaatgctg gtcgctatac tgctgtcgat
     1981 tcgatactaa cgccgccatc cagtgtcgaa aacgagctcg aattcatcga tgatatcaga
     2041 tccactagtg gcctatgcgg ccgcggatct gccggtctcc ctatagtgag tcgtattaat
     2101 ttcgataagc caggttaacc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt
     2161 gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct
     2221 gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga
     2281 taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc
     2341 cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg
     2401 ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg
     2461 aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt
     2521 tctcccttcg ggaagcgtgg cgctttctca atgctcacgc tgtaggtatc tcagttcggt
     2581 gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg
     2641 cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact
     2701 ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt
     2761 cttgaagtgg tggcctaact acggctacac tagaaggaca gtatttggta tctgcgctct
     2821 gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac
     2881 cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc
     2941 tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg
     3001 ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta
     3061 aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca
     3121 atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc
     3181 ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc
     3241 tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc
     3301 agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat
     3361 taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt
     3421 tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc
     3481 cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag
     3541 ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt
     3601 tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac
     3661 tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg
     3721 cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat
     3781 tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc
     3841 gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc
     3901 tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa
     3961 atgttgaata ctcatactct tcctttttca atattattga agcatttatc agggttattg
     4021 tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg
     4081 cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc attattatca tgacattaac
     4141 ctataaaaat aggcgtatca cgaggccctt tcgtctcgcg cgtttcggtg atgacggtga
     4201 aaacctctga cacatgcagc tcccggagac ggtcacagct tgtctgtaag cggatgccgg
     4261 gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc gggtgtcggg gctggcttaa
     4321 ctatgcggca tcagagcaga ttgtactgag agtgcaccat atggacatat tgtcgttaga
     4381 acgcggctac aattaataca taaccttatg tatcatacac atacgattta ggtgacacta
     4441 taga
//
