LOCUS       sequence-55622-e        4359 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 4359)
  AUTHORS   Longtine MS, McKenzie A 3rd, Demarini DJ, Shah NG, Wach A, Brachat 
            A, Philippsen P, Pringle JR
  TITLE     Additional modules for versatile and economical PCR-based gene 
            deletion and modification in Saccharomyces cerevisiae.
  JOURNAL   Yeast. 1998 Jul;14(10):953-61.
  PUBMED    9717241
REFERENCE   2  (bases 1 to 4359)
  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: "953-61"
COMMENT     Sequence Label: pFA6a-GFP(S65T)-TRP1
FEATURES             Location/Qualifiers
     source          1..4359
                     /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"
     promoter        1018..1165
                     /gene="S. cerevisiae TRP1"
                     /label=TRP1 promoter
     CDS             1166..1840
                     /codon_start=1
                     /gene="S. cerevisiae TRP1"
                     /product="phosphoribosylanthranilate isomerase, required 
                     for tryptophan biosynthesis"
                     /label=TRP1
                     /note="yeast auxotrophic marker"
                     /translation="MSVINFTGSSGPLVKVCGLQSTEAAECALDSDADLLGIICVPNRK
                     RTIDPVIARKISSLVKAYKNSSGTPKYLVGVFRNQPKEDVLALVNDYGIDIVQLHGDES
                     WQEYQEFLGLPVIKRLVFPKDCNILLSAASQKPHSFIPLFDSEAGGTGELLDWNSISDW
                     VGRQESPESLHFMLAGGLTPENVGDALRLNGVIGVDVSGGVETNGVKDSNKIANFVKNA
                     KK"
     RBS             1914..1922
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     primer_bind     complement(1994..2013)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1995..2013)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2100..2117)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2271..2859)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2351..2370)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3030..3821,3822..3890))
                     /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     3653..3672
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3891..3995)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4063..4081
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(4119..4141)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4241..4260
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        4341..4359
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     4341..4358
                     /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 tagatctgta
     1021 caatcttgat ccggagcttt tctttttttg ccgattaaga attaattcgg tcgaaaaaag
     1081 aaaaggagag ggccaagagg gagggcattg gtgactattg agcacgtgag tatacgtgat
     1141 taagcacaca aaggcagctt ggagtatgtc tgttattaat ttcacaggta gttctggtcc
     1201 attggtgaaa gtttgcggct tgcagagcac agaggccgca gaatgtgctc tagattccga
     1261 tgctgacttg ctgggtatta tatgtgtgcc caatagaaag agaacaattg acccggttat
     1321 tgcaaggaaa atttcaagtc ttgtaaaagc atataaaaat agttcaggca ctccgaaata
     1381 cttggttggc gtgtttcgta atcaacctaa ggaggatgtt ttggctctgg tcaatgatta
     1441 cggcattgat atcgtccaac tgcatggaga tgagtcgtgg caagaatacc aagagttcct
     1501 cggtttgcca gttattaaaa gactcgtatt tccaaaagac tgcaacatac tactcagtgc
     1561 agcttcacag aaacctcatt cgtttattcc cttgtttgat tcagaagcag gtgggacagg
     1621 tgaacttttg gattggaact cgatttctga ctgggttgga aggcaagaga gccccgaaag
     1681 cttacatttt atgttagctg gtggactgac gccagaaaat gttggtgatg cgcttagatt
     1741 aaatggcgtt attggtgttg atgtaagcgg aggtgtggag acaaatggtg taaaagactc
     1801 taacaaaata gcaaatttcg tcaaaaatgc taagaaatag gttattactg agtagtattt
     1861 atttaagtat tgtttgtgca cttgcctgcg gtgtgaaata ccgcacagat gcgtaaggag
     1921 gtttaaacga gctcgaattc atcgatgata tcagatccac tagtggccta tgcggccgcg
     1981 gatctgccgg tctccctata gtgagtcgta ttaatttcga taagccaggt taacctgcat
     2041 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc
     2101 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
     2161 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     2221 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     2281 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     2341 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     2401 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     2461 tctcaatgct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     2521 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     2581 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     2641 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     2701 tacactagaa ggacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     2761 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     2821 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     2881 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     2941 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     3001 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     3061 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     3121 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     3181 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     3241 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     3301 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     3361 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     3421 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     3481 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     3541 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     3601 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     3661 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     3721 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     3781 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     3841 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     3901 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     3961 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     4021 gacgtctaag aaaccattat tatcatgaca ttaacctata aaaataggcg tatcacgagg
     4081 ccctttcgtc tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg
     4141 gagacggtca cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg
     4201 tcagcgggtg ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta
     4261 ctgagagtgc accatatgga catattgtcg ttagaacgcg gctacaatta atacataacc
     4321 ttatgtatca tacacatacg atttaggtga cactataga
//
