LOCUS       sequence-61681-e        4308 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 4308)
  TITLE     Formosa lab yeast 3' tagging constructs
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 4308)
  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            "Unpublished"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: pTF277 (pFA6a-TEV-6xGly-2xStrep-KanMX)
FEATURES             Location/Qualifiers
     source          1..4308
                     /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..799
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             836..859
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     terminator      888..1075
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(943..967)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     gene            1150..2506
                     /label=kanMX
                     /note="yeast selectable marker conferring kanamycin 
                     resistance (Wach et al., 1994)"
     promoter        1150..1493
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             1494..2303
                     /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(1561..1580)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      2309..2506
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(2610..2629)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2611..2629)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2716..2733)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2887..3475)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2967..2986)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3646..4308,1..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     4269..4288
                     /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 ctagatggtc
      781 tcacccacag ttcgagaagg gtggaggctc tggcggtgga tccggaggcg gttcatggtc
      841 tcacccacag ttcgagaagt gactcgaggc gcgccacttc taaataagcg aatttcttat
      901 gatttatgat ttttattatt aaataagtta taaaaaaaat aagtgtatac aaattttaaa
      961 gtgactctta ggttttaaaa cgaaaattct tattcttgag taactctttc ctgtaggtca
     1021 ggttgctttc tcaggtatag tatgaggtcg ctcttattga ccacacctct accggcagat
     1081 ccgctaggga taacagggta atatagatct gtttagcttg cctcgtcccc gccgggtcac
     1141 ccggccagcg acatggaggc ccagaatacc ctccttgaca gtcttgacgt gcgcagctca
     1201 ggggcatgat gtgactgtcg cccgtacatt tagcccatac atccccatgt ataatcattt
     1261 gcatccatac attttgatgg ccgcacggcg cgaagcaaaa attacggctc ctcgctgcag
     1321 acctgcgagc agggaaacgc tcccctcaca gacgcgttga attgtcccca cgccgcgccc
     1381 ctgtagagaa atataaaagg ttaggatttg ccactgaggt tcttctttca tatacttcct
     1441 tttaaaatct tgctaggata cagttctcac atcacatccg aacataaaca accatgggta
     1501 aggaaaagac tcacgtttcg aggccgcgat taaattccaa catggatgct gatttatatg
     1561 ggtataaatg ggctcgcgat aatgtcgggc aatcaggtgc gacaatctat cgattgtatg
     1621 ggaagcccga tgcgccagag ttgtttctga aacatggcaa aggtagcgtt gccaatgatg
     1681 ttacagatga gatggtcaga ctaaactggc tgacggaatt tatgcctctt ccgaccatca
     1741 agcattttat ccgtactcct gatgatgcat ggttactcac cactgcgatc cccggcaaaa
     1801 cagcattcca ggtattagaa gaatatcctg attcaggtga aaatattgtt gatgcgctgg
     1861 cagtgttcct gcgccggttg cattcgattc ctgtttgtaa ttgtcctttt aacagcgatc
     1921 gcgtatttcg tctcgctcag gcgcaatcac gaatgaataa cggtttggtt gatgcgagtg
     1981 attttgatga cgagcgtaat ggctggcctg ttgaacaagt ctggaaagaa atgcataagc
     2041 ttttgccatt ctcaccggat tcagtcgtca ctcatggtga tttctcactt gataacctta
     2101 tttttgacga ggggaaatta ataggttgta ttgatgttgg acgagtcgga atcgcagacc
     2161 gataccagga tcttgccatc ctatggaact gcctcggtga gttttctcct tcattacaga
     2221 aacggctttt tcaaaaatat ggtattgata atcctgatat gaataaattg cagtttcatt
     2281 tgatgctcga tgagtttttc taatcagtac tgacaataaa aagattcttg ttttcaagaa
     2341 cttgtcattt gtatagtttt tttatattgt agttgttcta ttttaatcaa atgttagcgt
     2401 gatttatatt ttttttcgcc tcgacatcat ctgcccagat gcgaagttaa gtgcgcagaa
     2461 agtaatatca tgcgtcaatc gtatgtgaat gctggtcgct atactgctgt cgattcgata
     2521 ctaacgccgc catccagttt aaacgagctc gaattcatcg atgatatcag atccactagt
     2581 ggcctatgcg gccgcggatc tgccggtctc cctatagtga gtcgtattaa tttcgataag
     2641 ccaggttaac ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg
     2701 gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc
     2761 ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg
     2821 aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct
     2881 ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca
     2941 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct
     3001 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc
     3061 gggaagcgtg gcgctttctc aatgctcacg ctgtaggtat ctcagttcgg tgtaggtcgt
     3121 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc
     3181 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc
     3241 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg
     3301 gtggcctaac tacggctaca ctagaaggac agtatttggt atctgcgctc tgctgaagcc
     3361 agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag
     3421 cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga
     3481 tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat
     3541 tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag
     3601 ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat
     3661 cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc
     3721 cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat
     3781 accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag
     3841 ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg
     3901 ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc
     3961 tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca
     4021 acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg
     4081 tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc
     4141 actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta
     4201 ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc
     4261 aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcat
//
