LOCUS       sequence-297718-        7597 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 7597)
  TITLE     SARS-CoV-2 protein Yeast expression constructs, tagged, for protein 
            expression and/or purification
REFERENCE   2  (bases 1 to 7597)
  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-m4046454
FEATURES             Location/Qualifiers
     source          1..7597
                     /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             3224..3241
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             3242..3307
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     primer_bind     complement(3352..3370)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     CDS             3721..4524
                     /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      4607..5589
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     terminator      5667..5696
                     /label=T3Te terminator
                     /note="phage T3 early transcription terminator"
     rep_origin      5718..6305
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     terminator      complement(6317..6344)
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     CDS             complement(6368..6559)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(6560..7351,7352..7420))
                     /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     7183..7202
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7421..7523)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     terminator      complement(7524..7555)
                     /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 atggctgatg cacaatcgtt tttaaaccgg gtttgcggtg taagtgcagc
      481 ccgtcttaca ccgtgcggca caggcactag tactgatgtc gtatacaggg cttttgacat
      541 ctacaatgat aaagtagctg gttttgctaa attcctaaaa actaattgtt gtcgcttcca
      601 agaaaaggac gaagatgaca atttaattga ttcttacttt gtagttaaga gacacacttt
      661 ctctaactac caacatgaag aaacaattta taatttactt aaggattgtc cagctgttgc
      721 taaacatgac ttctttaagt ttagaataga cggtgacatg gtaccacata tatcacgtca
      781 acgtcttact aaatacacaa tggcagacct cgtctatgct ttaaggcatt ttgatgaagg
      841 taattgtgac acattaaaag aaatacttgt cacatacaat tgttgtgatg atgattattt
      901 caataaaaag gactggtatg attttgtaga aaacccagat atattacgcg tatacgccaa
      961 cttaggtgaa cgtgtacgcc aagctttgtt aaaaacagta caattctgtg atgccatgcg
     1021 aaatgctggt attgttggtg tactgacatt agataatcaa gatctcaatg gtaactggta
     1081 tgatttcggt gatttcatac aaaccacgcc aggtagtgga gttcctgttg tagattctta
     1141 ttattcattg ttaatgccta tattaacctt gaccagggct ttaactgcag agtcacatgt
     1201 tgacactgac ttaacaaagc cttacattaa gtgggatttg ttaaaatatg acttcacgga
     1261 agagaggtta aaactctttg accgttattt taaatattgg gatcagacat accacccaaa
     1321 ttgtgttaac tgtttggatg acagatgcat tctgcattgt gcaaacttta atgttttatt
     1381 ctctacagtg ttcccaccta caagttttgg accactagtg agaaaaatat ttgttgatgg
     1441 tgttccattt gtagtttcaa ctggatacca cttcagagag ctaggtgttg tacataatca
     1501 ggatgtaaac ttacatagct ctagacttag ttttaaggaa ttacttgtgt atgctgctga
     1561 ccctgctatg cacgctgctt ctggtaatct attactagat aaacgcacta cgtgcttttc
     1621 agtagctgca cttactaaca atgttgcttt tcaaactgtc aaacccggta attttaacaa
     1681 agacttctat gactttgctg tgtctaaggg tttctttaag gaaggaagtt ctgttgaatt
     1741 aaaacacttc ttctttgctc aggatggtaa tgctgctatc agcgattatg actactatcg
     1801 ttataatcta ccaacaatgt gtgatatcag acaactacta tttgtagttg aagttgttga
     1861 taagtacttt gattgttacg atggtggctg tattaatgct aaccaagtca tcgtcaacaa
     1921 cctagacaaa tcagctggtt ttccatttaa taaatggggt aaggctagac tttattatga
     1981 ttcaatgagt tatgaggatc aagatgcact tttcgcatat acaaaacgta atgtcatccc
     2041 tactataact caaatgaatc ttaagtatgc cattagtgca aagaatagag ctcgcaccgt
     2101 agctggtgtc tctatctgta gtactatgac caatagacag tttcatcaaa aattattgaa
     2161 atcaatagcc gccactagag gagctactgt agtaattgga acaagcaaat tctatggtgg
     2221 ttggcacaac atgttaaaaa ctgtttatag tgatgtagaa aaccctcacc ttatgggttg
     2281 ggattatcct aaatgtgata gagccatgcc taacatgctt agaattatgg cctcacttgt
     2341 tcttgctcgc aaacatacaa cgtgttgtag cttgtcacac cgtttctata gattagctaa
     2401 tgagtgtgct caagtattga gtgaaatggt catgtgtggc ggttcactat atgttaaacc
     2461 aggtggaacc tcatcaggag atgccacaac tgcttatgct aatagtgttt ttaacatttg
     2521 tcaagctgtc acggccaatg ttaatgcact tttatctact gatggtaaca aaattgccga
     2581 taagtatgtc cgcaatttac aacacagact ttatgagtgt ctctatagaa atagagatgt
     2641 tgacacagac tttgtgaatg agttttacgc atatttgcgt aaacatttct caatgatgat
     2701 actctctgac gatgctgttg tgtgtttcaa tagcacttat gcatctcaag gtctagtggc
     2761 tagcataaag aactttaagt cagttcttta ttatcaaaac aatgttttta tgtctgaagc
     2821 aaaatgttgg actgagactg accttactaa aggacctcat gaattttgct ctcaacatac
     2881 aatgctagtt aaacagggtg atgattatgt gtaccttcct tacccagatc catcaagaat
     2941 cctaggggcc ggctgttttg tagatgatat cgtaaaaaca gatggtacac ttatgattga
     3001 acggttcgtg tctttagcta tagatgctta cccacttact aaacatccta atcaggagta
     3061 tgctgatgtc tttcatttgt acttacaata cataagaaag ctacatgatg agttaacagg
     3121 acacatgtta gacatgtatt ctgttatgct tactaatgat aacacttcaa ggtattggga
     3181 acctgagttt tatgaggcta tgtacacacc gcatacagtc ttactcgtgc cgagaggttc
     3241 tgattataaa gatcatgatg gtgattataa agatcatgat attgattata aagatgatga
     3301 tgataaataa acaggcccct tttcctttgt cgatatcatg taattagtta tgtcacgctt
     3361 acattcacgc cctcctccca catccgctct aaccgaaaag gaaggagtta gacaacctga
     3421 agtctaggtc cctatttatt ttttttaata gttatgttag tattaagaac gttatttata
     3481 tttcaaattt ttcttttttt tctgtacaaa gcggccgccg aaccactagt gcactgcagt
     3541 acacggtttc cttgaaattt ttttgattcg gtaatctccg aacagaagga agaacgaagg
     3601 aaggagcaca gacttagatt ggtatatata cgcatatgta gtgttgaaga aacatgaaat
     3661 tgcccagtat tcttaaccca actgcacaga acaaaaacgt gcaggaaacg aagataaatc
     3721 atgtcgaaag ctacatataa ggaacgtgct gctactcatc ctagtcctgt tgctgccaag
     3781 ctatttaata tcatgcacga aaagcaaaca aacttgtgtg cttcattgga tgttcgtacc
     3841 accaaggaat tactggagtt agttgaagca ttaggtccca aaatttgttt actaaaaaca
     3901 catgtggata tcttgactga tttttccatg gagggcacag ttaagccgct aaaggcatta
     3961 tccgccaagt acaatttttt actcttcgag gacagaaaat ttgctgacat tggtaataca
     4021 gtcaaattgc agtactctgc gggtgtatac agaatagcag aatgggcaga cattacgaat
     4081 gcacacggtg tggtgggccc aggtattgtt agcggtttga agcaggcggc agaagaagta
     4141 acaaaggaac ctagaggcct tttgatgtta gcagaattgt catgcaaggg ctccctatct
     4201 actggagaat atactaaggg tactgttgac attgcgaaga gcgacaaaga ttttgttatc
     4261 ggctttattg ctcaaagaga catgggtgga agagatgaag gttacgattg gttgattatg
     4321 acacccggtg tgggtttaga tgacaaggga gatgcattgg gtcaacagta tagaaccgtg
     4381 gatgatgtgg tttctacagg atctgacatt attattgttg gaagaggact atttgcaaag
     4441 ggaagggatg ctaaggtaga gggtgaacgt tacagaaaag caggctggga agcatatttg
     4501 agaagatgcg gccagcaaaa ctaaaaaact gtattataag taaatgcatg tatactaaac
     4561 tcacaaatta gagcttcaat ttaattatat cagttattac ccgagtaacg aagcatctgt
     4621 gcttcatttt gtagaacaaa aatgcaacgc gagagcgcta atttttcaaa caaagaatct
     4681 gagctgcatt tttacagaac agaaatgcaa cgcgaaagcg ctattttacc aacgaagaat
     4741 ctgtgcttca tttttgtaaa acaaaaatgc aacgcgagag cgctaatttt tcaaacaaag
     4801 aatctgagct gcatttttac agaacagaaa tgcaacgcga gagcgctatt ttaccaacaa
     4861 agaatctata cttctttttt gttctacaaa aatgcatccc gagagcgcta tttttctaac
     4921 aaagcatctt agattacttt ttttctcctt tgtgcgctct ataatgcagt ctcttgataa
     4981 ctttttgcac tgtaggtccg ttaaggttag aagaaggcta ctttggtgtc tattttctct
     5041 tccataaaaa aagcctgact ccacttcccg cgtttactga ttactagcga agctgcgggt
     5101 gcattttttc aagataaagg catccccgat tatattctat accgatgtgg attgcgcata
     5161 ctttgtgaac agaaagtgat agcgttgatg attcttcatt ggtcagaaaa ttatgaacgg
     5221 tttcttctat tttgtctcta tatactacgt ataggaaatg tttacatttt cgtattgttt
     5281 tcgattcact ctatgaatag ttcttactac aatttttttg tctaaagagt aatactagag
     5341 ataaacataa aaaatgtaga ggtcgagttt agatgcaagt tcaaggagcg aaaggtggat
     5401 gggtaggtta tatagggata tagcacagag atatatagca aagagatact tttgagcaat
     5461 gtttgtggaa gcggtattcg caatatttta gtagctcgtt acagtccggt gcgtttttgg
     5521 ttttttgaaa gtgcgtcatc agagcgcttt tggttttcaa aagcgctctg aagttcctat
     5581 actttctagc tagagaatag gaacttcccg agcggccgcg tgttacaagc ttgactccag
     5641 cgtaactgga ctgcaatcaa ctcactggct caccttcacg ggtgggcctt tcttcggtag
     5701 aaaatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca
     5761 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt
     5821 ccgaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt
     5881 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc
     5941 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac
     6001 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca
     6061 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg
     6121 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag
     6181 gagagcgcac gagggagctt ccaggggtaa acgcctggta tctttatagt cctgtcgggt
     6241 ttcgccacct ctgacttgag catcgatttt tgtgatgctc gtcagggggg cggagcctat
     6301 ggaaaaacgc cagcaacgca gaaaggccca cccgaaggtg agccaggtga ttacatttgg
     6361 gccctcatca gaggttttca ccgtcatcac cgaaacgcgc gaggcagctg cggtaaagct
     6421 catcagcgtg gtcgtgaagc gattcacaga tgtctgcctg ttcatccgcg tccagctcgt
     6481 tgagtttctc cagaagcgtt aatgtctggc ttctgataaa gcgggccatg ttaagggcgg
     6541 ttttttcctg tttggtcact taccaatgct taatcagtga ggcacctatc tcagcgatct
     6601 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatgcggg
     6661 agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc
     6721 cagatttatc agcaataaac cagccagccg ggagggccga gcgcagaagt gatcctgcaa
     6781 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc
     6841 cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt
     6901 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc
     6961 ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgcc agaagtaagt
     7021 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc
     7081 catccgtgag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt
     7141 gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata
     7201 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgtaaa ctctcaagga
     7261 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag
     7321 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa
     7381 aaaagggaat aagggcgaca cggaaatgtt gaatactcat tttagcttcc ttagctcctg
     7441 aaaatctcga taactcaaaa aatacgcccg gtagtgatct tatttcatta tggtgaaagt
     7501 tggaacctct tacgtgccga tcaagtcaaa agcctccggt cggaggcttt tgactttctg
     7561 ctatggaggt caggtatgat ttaaatggtc agttaga
//
