LOCUS       sequence-49550-e        4882 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 4882)
  AUTHORS   Bahler J, Wu JQ, Longtine MS, Shah NG, McKenzie A 3rd, Steever AB, 
            Wach A, Philippsen P, Pringle JR
  TITLE     Heterologous modules for efficient and versatile PCR-based gene 
            targeting in Schizosaccharomyces pombe.
  JOURNAL   Yeast. 1998 Jul;14(10):943-51.
  PUBMED    9717240
REFERENCE   2  (bases 1 to 4882)
  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: "1998-07"; volume: "14(10)"; pages: "943-51"
COMMENT     Sequence Label: pFA6a-GFP(S65T)-kanMX6
FEATURES             Location/Qualifiers
     source          1..4882
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             61..771
                     /codon_start=1
                     /product="S65T variant of Aequorea victoria green 
                     fluorescent protein (Heim et al., 1995)"
                     /label=GFP (S65T)
                     /note="excitable with blue light"
                     /translation="SKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKF
                     ICTTGKLPVPWPTLVTTFTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTIFFKDDGN
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKVN
                     FKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLEF
                     VTAAGITHGMDELYK"
     primer_bind     complement(88..116)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     711..731
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     terminator      795..982
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(850..874)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     gene            1057..2413
                     /label=kanMX
                     /note="yeast selectable marker conferring kanamycin 
                     resistance (Wach et al., 1994)"
     promoter        1057..1400
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             1401..2210
                     /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(1468..1487)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      2216..2413
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(2517..2536)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2518..2536)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2623..2640)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2794..3382)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2874..2893)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3553..4344,4345..4413))
                     /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     4176..4195
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4414..4518)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4586..4604
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(4642..4664)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4764..4783
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        4864..4882
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     4864..4881
                     /label=SP6
                     /note="SP6 promoter, forward primer"
ORIGIN
        1 acgcggccgc cagctgaagc ttcgtacgct gcaggtcgac ggatccccgg gttaattaac
       61 agtaaaggag aagaactttt cactggagtt gtcccaattc ttgttgaatt agatggtgat
      121 gttaatgggc acaaattttc tgtcagtgga gagggtgaag gtgatgcaac atacggaaaa
      181 cttaccctta aatttatttg cactactgga aaactacctg ttccatggcc aacacttgtc
      241 actactttca cttatggtgt tcaatgcttt tcaagatacc cagatcatat gaaacggcat
      301 gactttttca agagtgccat gcccgaaggt tatgtacagg aaagaactat atttttcaaa
      361 gatgacggga actacaagac acgtgctgaa gtcaagtttg aaggtgatac ccttgttaat
      421 agaatcgagt taaaaggtat tgattttaaa gaagatggaa acattcttgg acacaaattg
      481 gaatacaact ataactcaca caatgtatac atcatggcag acaaacaaaa gaatggaatc
      541 aaagttaact tcaaaattag acacaacatt gaagatggaa gcgttcaact agcagaccat
      601 tatcaacaaa atactccaat tggcgatggc cctgtccttt taccagacaa ccattacctg
      661 tccacacaat ctgccctttc gaaagatccc aacgaaaaga gagaccacat ggtccttctt
      721 gagtttgtaa cagctgctgg gattacacat ggcatggatg aactatacaa atagggcgcg
      781 ccacttctaa ataagcgaat ttcttatgat ttatgatttt tattattaaa taagttataa
      841 aaaaaataag tgtatacaaa ttttaaagtg actcttaggt tttaaaacga aaattcttat
      901 tcttgagtaa ctctttcctg taggtcaggt tgctttctca ggtatagtat gaggtcgctc
      961 ttattgacca cacctctacc ggcagatccg ctagggataa cagggtaata tagatctgtt
     1021 tagcttgcct cgtccccgcc gggtcacccg gccagcgaca tggaggccca gaataccctc
     1081 cttgacagtc ttgacgtgcg cagctcaggg gcatgatgtg actgtcgccc gtacatttag
     1141 cccatacatc cccatgtata atcatttgca tccatacatt ttgatggccg cacggcgcga
     1201 agcaaaaatt acggctcctc gctgcagacc tgcgagcagg gaaacgctcc cctcacagac
     1261 gcgttgaatt gtccccacgc cgcgcccctg tagagaaata taaaaggtta ggatttgcca
     1321 ctgaggttct tctttcatat acttcctttt aaaatcttgc taggatacag ttctcacatc
     1381 acatccgaac ataaacaacc atgggtaagg aaaagactca cgtttcgagg ccgcgattaa
     1441 attccaacat ggatgctgat ttatatgggt ataaatgggc tcgcgataat gtcgggcaat
     1501 caggtgcgac aatctatcga ttgtatggga agcccgatgc gccagagttg tttctgaaac
     1561 atggcaaagg tagcgttgcc aatgatgtta cagatgagat ggtcagacta aactggctga
     1621 cggaatttat gcctcttccg accatcaagc attttatccg tactcctgat gatgcatggt
     1681 tactcaccac tgcgatcccc ggcaaaacag cattccaggt attagaagaa tatcctgatt
     1741 caggtgaaaa tattgttgat gcgctggcag tgttcctgcg ccggttgcat tcgattcctg
     1801 tttgtaattg tccttttaac agcgatcgcg tatttcgtct cgctcaggcg caatcacgaa
     1861 tgaataacgg tttggttgat gcgagtgatt ttgatgacga gcgtaatggc tggcctgttg
     1921 aacaagtctg gaaagaaatg cataagcttt tgccattctc accggattca gtcgtcactc
     1981 atggtgattt ctcacttgat aaccttattt ttgacgaggg gaaattaata ggttgtattg
     2041 atgttggacg agtcggaatc gcagaccgat accaggatct tgccatccta tggaactgcc
     2101 tcggtgagtt ttctccttca ttacagaaac ggctttttca aaaatatggt attgataatc
     2161 ctgatatgaa taaattgcag tttcatttga tgctcgatga gtttttctaa tcagtactga
     2221 caataaaaag attcttgttt tcaagaactt gtcatttgta tagttttttt atattgtagt
     2281 tgttctattt taatcaaatg ttagcgtgat ttatattttt tttcgcctcg acatcatctg
     2341 cccagatgcg aagttaagtg cgcagaaagt aatatcatgc gtcaatcgta tgtgaatgct
     2401 ggtcgctata ctgctgtcga ttcgatacta acgccgccat ccagtttaaa cgagctcgaa
     2461 ttcatcgatg atatcagatc cactagtggc ctatgcggcc gcggatctgc cggtctccct
     2521 atagtgagtc gtattaattt cgataagcca ggttaacctg cattaatgaa tcggccaacg
     2581 cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct
     2641 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     2701 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     2761 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     2821 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     2881 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     2941 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcaat gctcacgctg
     3001 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     3061 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     3121 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     3181 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaaggacagt
     3241 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     3301 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     3361 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     3421 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     3481 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     3541 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     3601 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     3661 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     3721 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     3781 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     3841 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     3901 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     3961 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     4021 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     4081 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     4141 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     4201 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     4261 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     4321 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     4381 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     4441 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     4501 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtct aagaaaccat
     4561 tattatcatg acattaacct ataaaaatag gcgtatcacg aggccctttc gtctcgcgcg
     4621 tttcggtgat gacggtgaaa acctctgaca catgcagctc ccggagacgg tcacagcttg
     4681 tctgtaagcg gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg
     4741 gtgtcggggc tggcttaact atgcggcatc agagcagatt gtactgagag tgcaccatat
     4801 ggacatattg tcgttagaac gcggctacaa ttaatacata accttatgta tcatacacat
     4861 acgatttagg tgacactata ga
//
