LOCUS       sequence-6741-ex        4738 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 4738)
  AUTHORS   Sheff MA, Thorn KS
  TITLE     Optimized cassettes for fluorescent protein tagging in Saccharomyces
            cerevisiae.
  JOURNAL   Yeast 2004 Jun;21(8):661-70.
  PUBMED    15197731
REFERENCE   2  (bases 1 to 4738)
  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"; 
            date: "2004-06"; volume: "21(8)"; pages: "661-70"
COMMENT     Sequence Label: pKT0128
FEATURES             Location/Qualifiers
     source          1..4738
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             5..721
                     /codon_start=1
                     /product="yeast-enhanced green fluorescent protein"
                     /label=yeGFP
                     /note="codons optimized for translation in S. cerevisiae 
                     and C. albicans"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     CDS             5..718
                     /codon_start=1
                     /product="green fluorescent protein"
                     /label=GFP
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     terminator      742..929
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(797..821)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     gene            1004..2204
                     /label=HIS3MX6
                     /note="yeast selectable marker encoding the S. pombe his5 
                     gene, which corresponds to S. cerevisiae HIS3"
     promoter        1004..1347
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             1348..2001
                     /codon_start=1
                     /gene="Schizosaccharomyces pombe his5"
                     /product="imidazoleglycerol-phosphate dehydratase, required
                     for histidine biosynthesis"
                     /label=S. pombe his5
                     /note="yeast auxotrophic marker; corresponds to S. 
                     cerevisiae HIS3"
                     /translation="MGRRAFVERNTNETKISVAIALDKAPLPEESNFIDELITSKHANQ
                     KGEQVIQVDTGIGFLDHMYHALAKHAGWSLRLYSRGDLIIDDHHTAEDTAIALGIAFKQ
                     AMGNFAGVKRFGHAYCPLDEALSRSVVDLSGRPYAVIDLGLKREKVGELSCEMIPHLLY
                     SFSVAAGITLHVTCLYGSNDHHRAESAFKSLAVAMRAATSLTGSSEVPSTKGVL"
     terminator      2007..2204
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(2308..2327)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2309..2327)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2414..2431)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2585..3173)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2665..2684)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3344..4135,4136..4204))
                     /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     3967..3986
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4205..4309)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4377..4395
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(4433..4455)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4555..4574
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        4655..4673
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     4655..4672
                     /label=SP6
                     /note="SP6 promoter, forward primer"
ORIGIN
        1 taacatgtct aaaggtgaag aattattcac tggtgttgtc ccaattttgg ttgaattaga
       61 tggtgatgtt aatggtcaca aattttctgt ctccggtgaa ggtgaaggtg atgctactta
      121 cggtaaattg accttaaaat ttatttgtac tactggtaaa ttgccagttc catggccaac
      181 cttagtcact actttaactt atggtgttca atgtttttct agatacccag atcatatgaa
      241 acaacatgac tttttcaagt ctgccatgcc agaaggttat gttcaagaaa gaactatttt
      301 tttcaaagat gacggtaact acaagaccag agctgaagtc aagtttgaag gtgatacctt
      361 agttaataga atcgaattaa aaggtattga ttttaaagaa gatggtaaca ttttaggtca
      421 caaattggaa tacaactata actctcacaa tgtttacatc atggctgaca aacaaaagaa
      481 tggtatcaaa gttaacttca aaattagaca caacattgaa gatggttctg ttcaattagc
      541 tgaccattat caacaaaata ctccaattgg tgatggtcca gtcttgttac cagacaacca
      601 ttacttatcc actcaatctg ccttatccaa agatccaaac gaaaagagag accacatggt
      661 cttgttagaa tttgttactg ctgctggtat tacccatggt atggatgaat tgtacaaata
      721 aggcgcgcca cttctaaata agcgaatttc ttatgattta tgatttttat tattaaataa
      781 gttataaaaa aaataagtgt atacaaattt taaagtgact cttaggtttt aaaacgaaaa
      841 ttcttattct tgagtaactc tttcctgtag gtcaggttgc tttctcaggt atagtatgag
      901 gtcgctctta ttgaccacac ctctaccggc agatccgcta gggataacag ggtaatatag
      961 atctgtttag cttgcctcgt ccccgccggg tcacccggcc agcgacatgg aggcccagaa
     1021 taccctcctt gacagtcttg acgtgcgcag ctcaggggca tgatgtgact gtcgcccgta
     1081 catttagccc atacatcccc atgtataatc atttgcatcc atacattttg atggccgcac
     1141 ggcgcgaagc aaaaattacg gctcctcgct gcagacctgc gagcagggaa acgctcccct
     1201 cacagacgcg ttgaattgtc cccacgccgc gcccctgtag agaaatataa aaggttagga
     1261 tttgccactg aggttcttct ttcatatact tccttttaaa atcttgctag gatacagttc
     1321 tcacatcaca tccgaacata aacaaccatg ggtaggaggg cttttgtaga aagaaatacg
     1381 aacgaaacga aaatcagcgt tgccatcgct ttggacaaag ctcccttacc tgaagagtcg
     1441 aattttattg atgaacttat aacttccaag catgcaaacc aaaagggaga acaagtaatc
     1501 caagtagaca cgggaattgg attcttggat cacatgtatc atgcactggc taaacatgca
     1561 ggctggagct tacgacttta ctcaagaggt gatttaatca tcgatgatca tcacactgca
     1621 gaagatactg ctattgcact tggtattgca ttcaagcagg ctatgggtaa ctttgccggc
     1681 gttaaaagat ttggacatgc ttattgtcca cttgacgaag ctctttctag aagcgtagtt
     1741 gacttgtcgg gacggcccta tgctgttatc gatttgggat taaagcgtga aaaggttggg
     1801 gaattgtcct gtgaaatgat ccctcactta ctatattcct tttcggtagc agctggaatt
     1861 actttgcatg ttacctgctt atatggtagt aatgaccatc atcgtgctga aagcgctttt
     1921 aaatctctgg ctgttgccat gcgcgcggct actagtctta ctggaagttc tgaagtccca
     1981 agcacgaagg gagtgttgta aagagtactg acaataaaaa gattcttgtt ttcaagaact
     2041 tgtcatttgt atagtttttt tatattgtag ttgttctatt ttaatcaaat gttagcgtga
     2101 tttatatttt ttttcgcctc gacatcatct gcccagatgc gaagttaagt gcgcagaaag
     2161 taatatcatg cgtcaatcgt atgtgaatgc tggtcgctat actgctgtcg attcgatact
     2221 aacgccgcca tccagtttaa acgagctcga attcatcgat gatatcagat ccactagtgg
     2281 cctatgcggc cgcggatctg ccggtctccc tatagtgagt cgtattaatt tcgataagcc
     2341 aggttaacct gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc
     2401 gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg
     2461 tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa
     2521 agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg
     2581 cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga
     2641 ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg
     2701 tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg
     2761 gaagcgtggc gctttctcaa tgctcacgct gtaggtatct cagttcggtg taggtcgttc
     2821 gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg
     2881 gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca
     2941 ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt
     3001 ggcctaacta cggctacact agaaggacag tatttggtat ctgcgctctg ctgaagccag
     3061 ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg
     3121 gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc
     3181 ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt
     3241 tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt
     3301 ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca
     3361 gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg
     3421 tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac
     3481 cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg
     3541 ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc
     3601 gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta
     3661 caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac
     3721 gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc
     3781 ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac
     3841 tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact
     3901 caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa
     3961 tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt
     4021 cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca
     4081 ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa
     4141 aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac
     4201 tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg
     4261 gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc
     4321 gaaaagtgcc acctgacgtc taagaaacca ttattatcat gacattaacc tataaaaata
     4381 ggcgtatcac gaggcccttt cgtctcgcgc gtttcggtga tgacggtgaa aacctctgac
     4441 acatgcagct cccggagacg gtcacagctt gtctgtaagc ggatgccggg agcagacaag
     4501 cccgtcaggg cgcgtcagcg ggtgttggcg ggtgtcgggg ctggcttaac tatgcggcat
     4561 cagagcagat tgtactgaga gtgcaccata tggacatatt gtcgttagaa cgcggctaca
     4621 attaatacat aaccttatgt atcatacaca tacgatttag gtgacactat agaacgcggc
     4681 cgccagctga agcttcgtac gctgcaggtc gacggatcgg tgacggtgct ggtttaat
//
