LOCUS       sequence-299913-        7603 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Yeast Expression plasmid for SARS-CoV-2 RNA-dependent RNA 
            polymerase.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7603)
  TITLE     SARS-CoV-2 protein Yeast expression constructs, tagged, for protein 
            expression and/or purification
REFERENCE   2  (bases 1 to 7603)
  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"
COMMENT     Sequence Label: pGBW-m4046567
FEATURES             Location/Qualifiers
     source          1..7603
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        10..417
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
     CDS             3230..3247
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             3248..3313
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     primer_bind     complement(3358..3376)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     CDS             3727..4530
                     /codon_start=1
                     /gene="S. cerevisiae URA3"
                     /product="orotidine-5'-phosphate decarboxylase, required 
                     for uracil biosynthesis"
                     /label=URA3
                     /note="yeast auxotrophic marker, counterselectable with 
                     5-fluoroorotic acid (5-FOA)"
                     /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
                     ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     rep_origin      4613..5595
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     terminator      5673..5702
                     /label=T3Te terminator
                     /note="phage T3 early transcription terminator"
     rep_origin      5724..6311
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     terminator      complement(6323..6350)
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     CDS             complement(6374..6565)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(6566..7357,7358..7426))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESLRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLATIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGSLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     7189..7208
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7427..7529)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     terminator      complement(7530..7561)
                     /label=tonB terminator
                     /note="bidirectional E. coli tonB-P14 transcription 
                     terminator"
ORIGIN
        1 gcggccgccc cacacaccat agcttcaaaa tgtttctact ccttttttac tcttccagat
       61 tttctcggac tccgcgcatc gccgtaccac ttcaaaacac ccaagcacag catactaaat
      121 ttcccctctt tcttcctcta gggtgtcgtt aattacccgt actaaaggtt tggaaaagaa
      181 aaaagagacc gcctcgtttc tttttcttcg tcgaaaaagg caataaaaat ttttatcacg
      241 tttctttttc ttgaaaattt ttttttttga tttttttctc tttcgatgac ctcccattga
      301 tatttaagtt aataaacggt cttcaatttc tcaagtttca gtttcatttt tcttgttcta
      361 ttacaacttt ttttacttct tgctcattag aaagaaagca tagcaatcta atctaagttt
      421 taattacaaa atgtccgccg atgcacaatc tttcttaaat agagtctgcg gcgtgagtgc
      481 tgctcgtcta accccttgtg gtacgggtac ttcaacagac gtagtttaca gggcgtttga
      541 tatttataac gacaaggtcg ctggtttcgc taaatttttg aagaccaact gttgtagatt
      601 ccaggagaag gatgaagacg acaatcttat cgattcgtac ttcgttgtta aaagacatac
      661 tttttccaac tatcaacacg aagaaacaat atacaatttg ctcaaggact gtccagccgt
      721 cgctaaacac gattttttca agttcagaat tgatggggac atggtgccac atatcagcag
      781 acaacggttg accaagtaca ctatggctga tttagtttac gccttgcgac actttgacga
      841 aggaaactgt gatactctga aggaaatttt ggttacctat aactgctgtg acgatgatta
      901 cttcaacaaa aaggactggt acgacttcgt agagaatccc gacatcttga gggtttacgc
      961 taacttaggt gaaagagtca gacaagcttt gttaaagaca gtccaatttt gcgatgctat
     1021 gagaaatgcg ggtattgttg gtgttttgac tttggataat caagatttga acggtaactg
     1081 gtatgacttc ggtgatttta tccagactac cccgggttct ggggtaccag ttgtcgattc
     1141 ttactattct ctgttaatgc caatactcac cctaactaga gcattgactg ccgaatccca
     1201 tgttgacact gatttaacta agccatacat taaatgggac cttctaaagt acgatttcac
     1261 cgaagaacgt ttaaagttgt tcgacagata ttttaaatac tgggaccaaa cctaccaccc
     1321 taattgtgtc aactgtttgg acgacagatg tattcttcac tgcgctaact ttaatgtgtt
     1381 gttctcgact gtttttccac caacctcttt cggtccgctg gttcgaaaga tttttgttga
     1441 cggtgttcca ttcgtggtca gtacgggata tcatttcaga gaattagggg tcgttcataa
     1501 ccaagacgtg aacttgcact catcaagatt gtccttcaag gaattgttgg tctacgcagc
     1561 tgatcctgca atgcacgccg cttctggaaa cttattgctg gataagagaa caacctgttt
     1621 cagcgttgcc gctcttacta ataacgtcgc gttccaaact gtaaagccgg gtaactttaa
     1681 taaagacttc tacgattttg ctgttagcaa gggttttttc aaagaaggtt ctagtgttga
     1741 gttgaaacac tttttctttg ctcaagacgg caatgctgct atcagtgatt atgactatta
     1801 ccgttacaac ttaccaacaa tgtgtgatat cagacagttg cttttcgtcg tcgaagttgt
     1861 tgataaatat tttgactgtt atgatggtgg ttgcatcaat gcaaatcaag tcattgtcaa
     1921 caacttagat aagtctgctg gcttcccatt caacaagtgg ggtaaagcca gattgtacta
     1981 tgattccatg tcctacgagg accaagatgc tttgttcgcg tacaccaaga gaaacgttat
     2041 tcctacaatt actcaaatga atctaaagta cgctatctct gcaaaaaatc gggccagaac
     2101 cgttgctggt gtttctatct gttctacgat gactaaccgt caatttcatc agaagctatt
     2161 gaaatccata gctgctacta gaggtgctac tgtagtcatc ggtacctcaa agttctatgg
     2221 tggttggcac aatatgctca agaccgttta ctctgatgtc gaaaacccac atcttatggg
     2281 ttgggactac cctaagtgtg atagagccat gccaaatatg ctaagaatta tggcatcctt
     2341 ggtcttggct agaaagcata ccacatgctg ctcattgtcc caccgttttt acagattggc
     2401 caacgaatgt gctcaagtcc tttctgaaat ggttatgtgt ggtggtagtc tatatgtaaa
     2461 gccaggcgga acttcgtctg gagatgccac cactgcttac gctaactcgg tttttaacat
     2521 ttgtcaagcc gttaccgcca acgtaaacgc tttgctgtct acggatggta acaagatcgc
     2581 tgataagtat gtgcgcaact tgcaacatag actttatgaa tgtttgtata gaaatcgaga
     2641 tgtcgatact gacttcgtta acgaatttta cgcgtactta agaaagcact tctccatgat
     2701 gattttgtca gacgacgctg ttgtttgttt caattctact tacgctagcc aaggtttggt
     2761 cgcttcaatc aaaaacttca agtctgtttt atactaccaa aacaatgttt ttatgtccga
     2821 ggcaaaatgc tggacggaaa cagatttgac taagggtcca catgaatttt gtagtcaaca
     2881 cacaatgtta gtgaaacaag gggacgacta cgtttaccta ccctacccag atccaagcag
     2941 aattttgggt gccggttgtt tcgtggacga catagtcaag accgacggta ccttgatgat
     3001 cgaacggttt gtctctctgg ccatagacgc ttacccattg actaagcacc ctaaccaaga
     3061 atatgctgat gtttttcatt tatatcttca atacattaga aaattacatg acgaattgac
     3121 cggtcacatg ttagatatgt actctgtgat gttgacgaac gataatacgt cacgttactg
     3181 ggaacccgaa ttttacgaag cgatgtacac tccacacaca gtattgcagc tcgtgccgag
     3241 aggttctgat tataaagatc atgatggtga ttataaagat catgatattg attataaaga
     3301 tgatgatgat aaataaacag gccccttttc ctttgtcgat atcatgtaat tagttatgtc
     3361 acgcttacat tcacgccctc ctcccacatc cgctctaacc gaaaaggaag gagttagaca
     3421 acctgaagtc taggtcccta tttatttttt ttaatagtta tgttagtatt aagaacgtta
     3481 tttatatttc aaatttttct tttttttctg tacaaagcgg ccgccgaacc actagtgcac
     3541 tgcagtacac ggtttccttg aaattttttt gattcggtaa tctccgaaca gaaggaagaa
     3601 cgaaggaagg agcacagact tagattggta tatatacgca tatgtagtgt tgaagaaaca
     3661 tgaaattgcc cagtattctt aacccaactg cacagaacaa aaacgtgcag gaaacgaaga
     3721 taaatcatgt cgaaagctac atataaggaa cgtgctgcta ctcatcctag tcctgttgct
     3781 gccaagctat ttaatatcat gcacgaaaag caaacaaact tgtgtgcttc attggatgtt
     3841 cgtaccacca aggaattact ggagttagtt gaagcattag gtcccaaaat ttgtttacta
     3901 aaaacacatg tggatatctt gactgatttt tccatggagg gcacagttaa gccgctaaag
     3961 gcattatccg ccaagtacaa ttttttactc ttcgaggaca gaaaatttgc tgacattggt
     4021 aatacagtca aattgcagta ctctgcgggt gtatacagaa tagcagaatg ggcagacatt
     4081 acgaatgcac acggtgtggt gggcccaggt attgttagcg gtttgaagca ggcggcagaa
     4141 gaagtaacaa aggaacctag aggccttttg atgttagcag aattgtcatg caagggctcc
     4201 ctatctactg gagaatatac taagggtact gttgacattg cgaagagcga caaagatttt
     4261 gttatcggct ttattgctca aagagacatg ggtggaagag atgaaggtta cgattggttg
     4321 attatgacac ccggtgtggg tttagatgac aagggagatg cattgggtca acagtataga
     4381 accgtggatg atgtggtttc tacaggatct gacattatta ttgttggaag aggactattt
     4441 gcaaagggaa gggatgctaa ggtagagggt gaacgttaca gaaaagcagg ctgggaagca
     4501 tatttgagaa gatgcggcca gcaaaactaa aaaactgtat tataagtaaa tgcatgtata
     4561 ctaaactcac aaattagagc ttcaatttaa ttatatcagt tattacccga gtaacgaagc
     4621 atctgtgctt cattttgtag aacaaaaatg caacgcgaga gcgctaattt ttcaaacaaa
     4681 gaatctgagc tgcattttta cagaacagaa atgcaacgcg aaagcgctat tttaccaacg
     4741 aagaatctgt gcttcatttt tgtaaaacaa aaatgcaacg cgagagcgct aatttttcaa
     4801 acaaagaatc tgagctgcat ttttacagaa cagaaatgca acgcgagagc gctattttac
     4861 caacaaagaa tctatacttc ttttttgttc tacaaaaatg catcccgaga gcgctatttt
     4921 tctaacaaag catcttagat tacttttttt ctcctttgtg cgctctataa tgcagtctct
     4981 tgataacttt ttgcactgta ggtccgttaa ggttagaaga aggctacttt ggtgtctatt
     5041 ttctcttcca taaaaaaagc ctgactccac ttcccgcgtt tactgattac tagcgaagct
     5101 gcgggtgcat tttttcaaga taaaggcatc cccgattata ttctataccg atgtggattg
     5161 cgcatacttt gtgaacagaa agtgatagcg ttgatgattc ttcattggtc agaaaattat
     5221 gaacggtttc ttctattttg tctctatata ctacgtatag gaaatgttta cattttcgta
     5281 ttgttttcga ttcactctat gaatagttct tactacaatt tttttgtcta aagagtaata
     5341 ctagagataa acataaaaaa tgtagaggtc gagtttagat gcaagttcaa ggagcgaaag
     5401 gtggatgggt aggttatata gggatatagc acagagatat atagcaaaga gatacttttg
     5461 agcaatgttt gtggaagcgg tattcgcaat attttagtag ctcgttacag tccggtgcgt
     5521 ttttggtttt ttgaaagtgc gtcatcagag cgcttttggt tttcaaaagc gctctgaagt
     5581 tcctatactt tctagctaga gaataggaac ttcccgagcg gccgcgtgtt acaagcttga
     5641 ctccagcgta actggactgc aatcaactca ctggctcacc ttcacgggtg ggcctttctt
     5701 cggtagaaaa tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg
     5761 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact
     5821 ctttttccga ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt
     5881 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
     5941 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact
     6001 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac
     6061 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag
     6121 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg
     6181 gaacaggaga gcgcacgagg gagcttccag gggtaaacgc ctggtatctt tatagtcctg
     6241 tcgggtttcg ccacctctga cttgagcatc gatttttgtg atgctcgtca ggggggcgga
     6301 gcctatggaa aaacgccagc aacgcagaaa ggcccacccg aaggtgagcc aggtgattac
     6361 atttgggccc tcatcagagg ttttcaccgt catcaccgaa acgcgcgagg cagctgcggt
     6421 aaagctcatc agcgtggtcg tgaagcgatt cacagatgtc tgcctgttca tccgcgtcca
     6481 gctcgttgag tttctccaga agcgttaatg tctggcttct gataaagcgg gccatgttaa
     6541 gggcggtttt ttcctgtttg gtcacttacc aatgcttaat cagtgaggca cctatctcag
     6601 cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga
     6661 tgcgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac
     6721 cggctccaga tttatcagca ataaaccagc cagccgggag ggccgagcgc agaagtgatc
     6781 ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta
     6841 gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac
     6901 gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat
     6961 gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgccagaa
     7021 gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg
     7081 tcatgccatc cgtgagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag
     7141 aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc
     7201 cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgtaaactct
     7261 caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat
     7321 cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg
     7381 ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatttta gcttccttag
     7441 ctcctgaaaa tctcgataac tcaaaaaata cgcccggtag tgatcttatt tcattatggt
     7501 gaaagttgga acctcttacg tgccgatcaa gtcaaaagcc tccggtcgga ggcttttgac
     7561 tttctgctat ggaggtcagg tatgatttaa atggtcagtt aga
//
