LOCUS       sequence-61672-e        4290 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 4290)
  TITLE     Formosa lab yeast 3' tagging constructs
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 4290)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pTF268 (pFA6a-TEV-6xGly-3xFlag-KanMX)
FEATURES             Location/Qualifiers
     source          1..4290
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(199..303)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     371..389
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(427..449)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     549..568
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        649..667
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     649..666
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     CDS             728..748
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             776..841
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     terminator      870..1057
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(925..949)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     gene            1132..2488
                     /label=kanMX
                     /note="yeast selectable marker conferring kanamycin 
                     resistance (Wach et al., 1994)"
     promoter        1132..1475
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             1476..2285
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin"
                     /translation="MGKEKTHVSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(1543..1562)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      2291..2488
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(2592..2611)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2593..2611)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2698..2715)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2869..3457)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2949..2968)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3628..4290,1..129,130..198))
                     /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     4251..4270
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
       61 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
      121 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
      181 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta
      241 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc
      301 gcgcacattt ccccgaaaag tgccacctga cgtctaagaa accattatta tcatgacatt
      361 aacctataaa aataggcgta tcacgaggcc ctttcgtctc gcgcgtttcg gtgatgacgg
      421 tgaaaacctc tgacacatgc agctcccgga gacggtcaca gcttgtctgt aagcggatgc
      481 cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc ggggctggct
      541 taactatgcg gcatcagagc agattgtact gagagtgcac catatggaca tattgtcgtt
      601 agaacgcggc tacaattaat acataacctt atgtatcata cacatacgat ttaggtgaca
      661 ctatagaacg cggccgccag ctgaagcttc gtacgctgca ggtcgacgga tccccgggtt
      721 aattaacgag aatttatact tccaaggttc tggtggaggc ggtggaggct ctagagacta
      781 caaggaccac gacggtgact acaaggacca cgacattgac tacaaggatg acgatgacaa
      841 gtgactcgag gcgcgccact tctaaataag cgaatttctt atgatttatg atttttatta
      901 ttaaataagt tataaaaaaa ataagtgtat acaaatttta aagtgactct taggttttaa
      961 aacgaaaatt cttattcttg agtaactctt tcctgtaggt caggttgctt tctcaggtat
     1021 agtatgaggt cgctcttatt gaccacacct ctaccggcag atccgctagg gataacaggg
     1081 taatatagat ctgtttagct tgcctcgtcc ccgccgggtc acccggccag cgacatggag
     1141 gcccagaata ccctccttga cagtcttgac gtgcgcagct caggggcatg atgtgactgt
     1201 cgcccgtaca tttagcccat acatccccat gtataatcat ttgcatccat acattttgat
     1261 ggccgcacgg cgcgaagcaa aaattacggc tcctcgctgc agacctgcga gcagggaaac
     1321 gctcccctca cagacgcgtt gaattgtccc cacgccgcgc ccctgtagag aaatataaaa
     1381 ggttaggatt tgccactgag gttcttcttt catatacttc cttttaaaat cttgctagga
     1441 tacagttctc acatcacatc cgaacataaa caaccatggg taaggaaaag actcacgttt
     1501 cgaggccgcg attaaattcc aacatggatg ctgatttata tgggtataaa tgggctcgcg
     1561 ataatgtcgg gcaatcaggt gcgacaatct atcgattgta tgggaagccc gatgcgccag
     1621 agttgtttct gaaacatggc aaaggtagcg ttgccaatga tgttacagat gagatggtca
     1681 gactaaactg gctgacggaa tttatgcctc ttccgaccat caagcatttt atccgtactc
     1741 ctgatgatgc atggttactc accactgcga tccccggcaa aacagcattc caggtattag
     1801 aagaatatcc tgattcaggt gaaaatattg ttgatgcgct ggcagtgttc ctgcgccggt
     1861 tgcattcgat tcctgtttgt aattgtcctt ttaacagcga tcgcgtattt cgtctcgctc
     1921 aggcgcaatc acgaatgaat aacggtttgg ttgatgcgag tgattttgat gacgagcgta
     1981 atggctggcc tgttgaacaa gtctggaaag aaatgcataa gcttttgcca ttctcaccgg
     2041 attcagtcgt cactcatggt gatttctcac ttgataacct tatttttgac gaggggaaat
     2101 taataggttg tattgatgtt ggacgagtcg gaatcgcaga ccgataccag gatcttgcca
     2161 tcctatggaa ctgcctcggt gagttttctc cttcattaca gaaacggctt tttcaaaaat
     2221 atggtattga taatcctgat atgaataaat tgcagtttca tttgatgctc gatgagtttt
     2281 tctaatcagt actgacaata aaaagattct tgttttcaag aacttgtcat ttgtatagtt
     2341 tttttatatt gtagttgttc tattttaatc aaatgttagc gtgatttata ttttttttcg
     2401 cctcgacatc atctgcccag atgcgaagtt aagtgcgcag aaagtaatat catgcgtcaa
     2461 tcgtatgtga atgctggtcg ctatactgct gtcgattcga tactaacgcc gccatccagt
     2521 ttaaacgagc tcgaattcat cgatgatatc agatccacta gtggcctatg cggccgcgga
     2581 tctgccggtc tccctatagt gagtcgtatt aatttcgata agccaggtta acctgcatta
     2641 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc
     2701 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa
     2761 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
     2821 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     2881 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     2941 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     3001 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     3061 tcaatgctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     3121 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     3181 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     3241 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     3301 cactagaagg acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
     3361 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
     3421 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
     3481 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
     3541 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag
     3601 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc
     3661 agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac
     3721 gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc
     3781 accggctcca gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg
     3841 tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag
     3901 tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc
     3961 acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac
     4021 atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag
     4081 aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac
     4141 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg
     4201 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc
     4261 gccacatagc agaactttaa aagtgctcat
//
