LOCUS       sequence-297853-        7339 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Yeast Expression plasmid for SARS-CoV-2 nsp4.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7339)
  TITLE     SARS-CoV-2 protein Yeast expression constructs, tagged, for protein 
            expression and/or purification
REFERENCE   2  (bases 1 to 7339)
  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-m4046478
FEATURES             Location/Qualifiers
     source          1..7339
                     /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             1934..1951
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     primer_bind     complement(3094..3112)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     CDS             3463..4266
                     /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
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGXXX"
     rep_origin      4349..5331
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     terminator      5409..5438
                     /label=T3Te terminator
                     /note="phage T3 early transcription terminator"
     rep_origin      5460..6047
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     terminator      complement(6059..6086)
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     CDS             complement(6110..6301)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(6302..7093,7094..7162))
                     /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     6925..6944
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7163..7265)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     terminator      complement(7266..7297)
                     /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 atgaagattg tcaacaattg gcttaaacag ttgataaagg ttactttagt
      481 atttttgttc gttgcagcca tcttctatct aattacacct gtccacgtga tgagtaagca
      541 taccgacttt tcctcagaga tcattggtta caaagctatc gatggcggag ttactagaga
      601 tattgcttcg acggacacct gtttcgctaa caagcacgcg gactttgata cttggttctc
      661 tcaaaggggt ggttcctaca caaacgacaa ggcctgcccc ttgattgcag ctgttatcac
      721 tagagaagtt ggtttcgtcg tgccaggttt accagggact atactgcgta ccaccaatgg
      781 tgattttttg catttcttac caagagtctt ctctgctgtt ggcaatattt gttatacccc
      841 tagcaaattg atcgaataca ctgattttgc cacctctgct tgtgtattgg ctgctgaatg
      901 cactattttc aaggatgcaa gtggtaagcc agttccatac tgttatgaca caaacgtcct
      961 cgaaggttct gttgcttacg aatccttgcg accggacact agatatgtgt taatggatgg
     1021 atcaatcatt caatttccaa acacatacct tgaaggttct gttcgggttg tcaccacctt
     1081 cgactccgag tactgtcgcc acggtacttg tgaaagatca gaagccggtg tctgtgtttc
     1141 taccagcggt agatgggtgt tgaataacga ttattaccgt tctcttccag gtgtcttctg
     1201 cggtgtcgat gctgttaatc tattgactaa catgtttacc cctttgatcc aaccaatagg
     1261 tgcgctggac atttccgcat ctattgtcgc cggtggaatc gttgctatcg ttgttacttg
     1321 tttggcttac tacttcatga gatttagaag agcctttggg gagtattcgc atgtcgtggc
     1381 tttcaacact ctattatttt tgatgtcgtt cactgttttg tgtcttacac cagtatactc
     1441 tttcttgcct ggtgtatact ccgttatcta tttatacctg acattctacc tgactaacga
     1501 tgtcagcttt ttggcacaca ttcaatggat ggtaatgttc accccattgg ttcccttctg
     1561 gataacgatc gcttacatca tttgcataag tactaaacat ttctattggt ttttctcaaa
     1621 ttatctaaag agaagagttg tctttaatgg tgtcagcttt tctaccttcg aagaagctgc
     1681 cttgtgtacc tttttgctca acaaagaaat gtacttgaag ttgagatctg acgttttact
     1741 cccgctaacc caatataaca gatatttagc tttatacaac aagtacaaat acttctcggg
     1801 tgccatggat actacttcat accgtgaagc ggcttgttgt caccttgcta aggctttgaa
     1861 tgatttctct aattccggtt ccgacgtctt gtatcaacca cctcagacgt ctattacctc
     1921 cgctgttttg caactcgtgc cgagaggttc taaaatcgag gaaggtaagc tagtcatttg
     1981 gataaatgga gataagggtt acaacggttt ggcagaagta ggcaagaaat ttgaaaagga
     2041 cacaggtatc aaggtgacgg ttgaacatcc agacaagtta gaagaaaaat tccctcaagt
     2101 tgctgctact ggtgatggtc ccgacattat tttctgggcc cacgatcgtt ttggtggata
     2161 tgctcagtca ggtttgctgg ctgagatcac cccagacaag gccttccaag ataaattgta
     2221 cccattcact tgggatgctg tcagatacaa cggtaagctt atagcatatc caattgctgt
     2281 tgaagcctta tcgttgatct acaataagga cttgttacct aacccaccaa agacatggga
     2341 agaaattcca gctttggata aagagctaaa ggccaaggga aaaagcgctt tgatgtttaa
     2401 cttacaagaa ccttatttca cctggccatt aatcgctgct gacggtggtt acgcatttaa
     2461 atacgaaaac ggtaagtatg acattaagga tgtcggcgtt gataatgctg gtgcaaaggc
     2521 tggtttgacc ttcttggttg atttaataaa gaataaacac atgaacgctg acactgacta
     2581 ctctatcgcc gaagccgctt tcaacaaggg tgaaactgct atgaccataa atggtccatg
     2641 ggcctggagt aacattgata cttctaaagt aaactatggt gtcactgttt tgcctacatt
     2701 caaaggccaa ccatccaagc cctttgtcgg tgttctatcc gctggcatca atgctgcttc
     2761 tccaaacaag gagctagcta aagaattttt ggaaaattac ttgttaactg acgaaggttt
     2821 ggaggccgtt aacaaggaca agccattggg tgctgtcgct ttgaaatctt acgaagaaga
     2881 attggctaag gatcccagaa ttgccgctac catggagaac gcccaaaagg gtgaaattat
     2941 gccaaacatc ccacaaatgt cagccttctg gtacgcagtt aggactgccg tgattaatgc
     3001 tgcttccggt agacaaaccg tcgacgaagc attaaaagat gctcaaacgt aaacaggccc
     3061 cttttccttt gtcgatatca tgtaattagt tatgtcacgc ttacattcac gccctcctcc
     3121 cacatccgct ctaaccgaaa aggaaggagt tagacaacct gaagtctagg tccctattta
     3181 ttttttttaa tagttatgtt agtattaaga acgttattta tnnttcaaat ttttcttttt
     3241 tttctgtaca aagcggccgc cgaaccacta gtgcactgca gtacacggtt tccttgaaat
     3301 ttttttgatt cggtaatctc cgaacagaag gaagaacgaa ggaaggagca cagacttaga
     3361 ttggtatata tacgcatatg tagtgttgaa gaaacatgaa attgcccagt attcttaacc
     3421 caactgcaca gaacaaaaac gtgcaggaaa cgaagataaa tcatgtcgaa agctacatat
     3481 aaggaacgtg ctgctactca tcctagtcct gttgctgcca agctatttaa tatcatgcac
     3541 gaaaagcaaa caaacttgtg tgcttcattg gatgttcgta ccaccaagga attactggag
     3601 ttagttgaag cattaggtcc caaaatttgt ttactaaaaa cacatgtgga tatcttgact
     3661 gatttttcca tggagggcac agttaagccg ctaaaggcat tatccgccaa gtacaatttt
     3721 ttactcttcg aggacagaaa atttgctgac attggtaata cagtcaaatt gcagtactct
     3781 gcgggtgtat acagaatagc agaatgggca gacattacga atgcacacgg tgtggtgggc
     3841 ccaggtattg ttagcggttt gaagcaggcg gcagaagaag taacaaagga acctagaggc
     3901 cttttgatgt tagcagaatt gtcatgcaag ggctccctat ctactggaga atatactaag
     3961 ggtactgttg acattgcgaa gagcgacaaa gattttgtta tcggctttat tgctcaaaga
     4021 gacatgggtg gaagagatga aggttacgat tggttgatta tgacacccgg tgtgggttta
     4081 gatgacaagg gagatgcatt gggtcaacag tatagaaccg tggatgatgt ggtttctaca
     4141 ggatctgaca ttattattgt tggaagagga ctatttgcaa agggaaggga tgctaaggta
     4201 gagggtgaac gttacagaaa agcaggctgg gaagcatatt tgagaagatg cggcnnnnnn
     4261 nnntaaaaaa ctgtattata agtaaatgca tgtatactaa actcacaaat tagagcttca
     4321 atttaattat atcagttatt acccgagtaa cgaagcatct gtgcttcatt ttgtagaaca
     4381 aaaatgcaac gcgagagcgc taatttttca aacaaagaat ctgagctgca tttttacaga
     4441 acagaaatgc aacgcgaaag cgctatttta ccaacgaaga atctgtgctt catttttgta
     4501 aaacaaaaat gcaacgcgag agcgctaatt tttcaaacaa agaatctgag ctgcattttt
     4561 acagaacaga aatgcaacgc gagagcgcta ttttaccaac aaagaatcta tacttctttt
     4621 ttgttctaca aaaatgcatc ccgagagcgc tatttttcta acaaagcatc ttagattact
     4681 ttttttctcc tttgtgcgct ctataatgca gtctcttgat aactttttgc actgtaggtc
     4741 cgttaaggtt agaagaaggc tactttggtg tctattttct cttccataaa aaaagcctga
     4801 ctccacttcc cgcgtttact gattactagc gaagctgcgg gtgcattttt tcaagataaa
     4861 ggcatccccg attatattct ataccgatgt ggattgcgca tactttgtga acagaaagtg
     4921 atagcgttga tgattcttca ttggtcagaa aattatgaac ggtttcttct attttgtctc
     4981 tatatactac gtataggaaa tgtttacatt ttcgtattgt tttcgattca ctctatgaat
     5041 agttcttact acaatttttt tgtctaaaga gtaatactag agataaacat aaaaaatgta
     5101 gaggtcgagt ttagatgcaa gttcaaggag cgaaaggtgg atgggtaggt tatataggga
     5161 tatagcacag agatatatag caaagagata cttttgagca atgtttgtgg aagcggtatt
     5221 cgcaatattt tagtagctcg ttacagtccg gtgcgttttt ggttttttga aagtgcgtca
     5281 tcagagcgct tttggttttc aaaagcgctc tgaagttcct atactttcta gctagagaat
     5341 aggaacttcc cgagcggccg cgtgttacaa gcttgactcc agcgtaactg gactgcaatc
     5401 aactcactgg ctcaccttca cgggtgggcc tttcttcggt agaaaatcaa aggatcttct
     5461 tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca
     5521 gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaggta actggcttca
     5581 gcagagcgca gataccaaat actgttcttc tagtgtagcc gtagttaggc caccacttca
     5641 agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg
     5701 ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg
     5761 cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct
     5821 acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga
     5881 gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc
     5941 ttccaggggt aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg
     6001 agcatcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg
     6061 cagaaaggcc cacccgaagg tgagccaggt gattacattt gggccctcat cagaggtttt
     6121 caccgtcatc accgaaacgc gcgaggcagc tgcggtaaag ctcatcagcg tggtcgtgaa
     6181 gcgattcaca gatgtctgcc tgttcatccg cgtccagctc gttgagtttc tccagaagcg
     6241 ttaatgtctg gcttctgata aagcgggcca tgttaagggc ggttttttcc tgtttggtca
     6301 cttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca
     6361 tagttgcctg actccccgtc gtgtagataa ctacgatgcg ggagggctta ccatctggcc
     6421 ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa
     6481 accagccagc cgggagggcc gagcgcagaa gtgatcctgc aactttatcc gcctccatcc
     6541 agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca
     6601 acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat
     6661 tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag
     6721 cggttagctc cttcggtcct ccgatcgttg ccagaagtaa gttggccgca gtgttatcac
     6781 tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgtg agatgctttt
     6841 ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt
     6901 gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc
     6961 tcatcattgg aaaacgttct tcggggcgta aactctcaag gatcttaccg ctgttgagat
     7021 ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca
     7081 gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga
     7141 cacggaaatg ttgaatactc attttagctt ccttagctcc tgaaaatctc gataactcaa
     7201 aaaatacgcc cggtagtgat cttatttcat tatggtgaaa gttggaacct cttacgtgcc
     7261 gatcaagtca aaagcctccg gtcggaggct tttgactttc tgctatggag gtcaggtatg
     7321 atttaaatgg tcagttaga
//
