LOCUS       sequence-298222-        6204 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Yeast Expression plasmid for SARS-CoV-2 nsp4.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 6204)
  TITLE     SARS-CoV-2 protein Yeast expression constructs, tagged, for protein 
            expression and/or purification
REFERENCE   2  (bases 1 to 6204)
  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-m4046667
FEATURES             Location/Qualifiers
     source          1..6204
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     primer_bind     complement(1959..1977)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     CDS             2328..3131
                     /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      3214..4196
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     terminator      4274..4303
                     /label=T3Te terminator
                     /note="phage T3 early transcription terminator"
     rep_origin      4325..4912
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     terminator      complement(4924..4951)
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     CDS             complement(4975..5166)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(5167..5958,5959..6027))
                     /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     5790..5809
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6028..6130)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     terminator      complement(6131..6162)
                     /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 tttttttnga tttttttctc tttcgatgac ctcccattga
      301 tatttaagtt aataaacggt cttcaatttc tcaagtttca gtttcatttt tcttgttcta
      361 ttacaacttt ttttacgaaa gaaagcatag caatctaatc taagttttaa ttacaaaatg
      421 attgttaata attggttgaa gcagttaatt aaagttacac ttgtgttcct ttttgttgct
      481 gctattttct atttaataac acctgttcat gtcatgtcta aacatactga cttttcaagt
      541 gaaatcatag gatacaaggc tattgatggt ggtgtcactc gtgacatagc atctacagat
      601 acttgttttg ctaacaaaca tgctgatttt gacacatggt ttagccagcg tggtggtagt
      661 tatactaatg acaaagcttg cccattgatt gctgcagtca taacaagaga agtgggtttt
      721 gtcgtgcctg gtttgcctgg cacgatatta cgcacaacta atggtgactt tttgcatttc
      781 ttacctagag tttttagtgc agttggtaac atctgttaca caccatcaaa acttatagag
      841 tacactgact ttgcaacatc agcttgtgtt ttggctgctg aatgtacaat ttttaaagat
      901 gcttctggta agccagtacc atattgttat gataccaatg tactagaagg ttctgttgct
      961 tatgaaagtt tacgccctga cacacgttat gtgctcatgg atggctctat tattcaattt
     1021 cctaacacct accttgaagg ttctgttaga gtggtaacaa cttttgattc tgagtactgt
     1081 aggcacggca cttgtgaaag atcagaagct ggtgtttgtg tatctactag tggtagatgg
     1141 gtacttaaca atgattatta cagatcttta ccaggagttt tctgtggtgt agatgctgta
     1201 aatttactta ctaatatgtt tacaccacta attcaaccta ttggtgcttt ggacatatca
     1261 gcatctatag tagctggtgg tattgtagct atcgtagtaa catgccttgc ctactatttt
     1321 atgaggttta gaagagcttt tggtgaatac agtcatgtag ttgcctttaa tactttacta
     1381 ttccttatgt cattcactgt actctgttta acaccagttt actcattctt acctggtgtt
     1441 tattctgtta tttacttgta cttgacattt tatcttacta atgatgtttc ttttttagca
     1501 catattcagt ggatggttat gttcacacct ttagtacctt tctggataac aattgcttat
     1561 atcatttgta tttccacaaa gcatttctat tggttcttta gtaattacct aaagagacgt
     1621 gtagtcttta atggtgtttc ctttagtact tttgaagaag ctgcgctgtg cacctttttg
     1681 ttaaataaag aaatgtatct aaagttgcgt agtgatgtgc tattacctct tacgcaatat
     1741 aatagatact tagctcttta taataagtac aagtatttta gtggagcaat ggatacaact
     1801 agctacagag aagctgcttg ttgtcatctc gcaaaggctc tcaatgactt cagtaactca
     1861 ggttctgatg ttctttacca accaccacaa acctctatca cctcagctgt tttgtaaaca
     1921 ggcccctttt cctttgtcga tatcatgtaa ttagttatgt cacgcttaca ttcacgccct
     1981 cctcccacat ccgctctaac cgaaaaggaa ggagttagac aacctgaagt ctaggtccct
     2041 atttattttt tttaatagtt atgttagtat taagaacgtt atttatattt caaatttttc
     2101 ttttttttct gtacaaagcg gccgccgaac cactagtgca ctgcagtaca cggtttcctt
     2161 gaaatttttt tgattcggta atctccgaac agaaggaaga acgaaggaag gagcacagac
     2221 ttagattggt atatatacgc atatgtagtg ttgaagaaac atgaaattgc ccagtattct
     2281 taacccaact gcacagaaca aaaacgtgca ggaaacgaag ataaatcatg tcgaaagcta
     2341 catataagga acgtgctgct actcatccta gtcctgttgc tgccaagcta tttaatatca
     2401 tgcacgaaaa gcaaacaaac ttgtgtgctt cattggatgt tcgtaccacc aaggaattac
     2461 tggagttagt tgaagcatta ggtcccaaaa tttgtttact aaaaacacat gtggatatct
     2521 tgactgattt ttccatggag ggcacagtta agccgctaaa ggcattatcc gccaagtaca
     2581 attttttact cttcgaggac agaaaatttg ctgacattgg taatacagtc aaattgcagt
     2641 actctgcggg tgtatacaga atagcagaat gggcagacat tacgaatgca cacggtgtgg
     2701 tgggcccagg tattgttagc ggtttgaagc aggcggcaga agaagtaaca aaggaaccta
     2761 gaggcctttt gatgttagca gaattgtcat gcaagggctc cctatctact ggagaatata
     2821 ctaagggtac tgttgacatt gcgaagagcg acaaagattt tgttatcggc tttattgctc
     2881 aaagagacat gggtggaaga gatgaaggtt acgattggtt gattatgaca cccggtgtgg
     2941 gtttagatga caagggagat gcattgggtc aacagtatag aaccgtggat gatgtggttt
     3001 ctacaggatc tgacattatt attgttggaa gaggactatt tgcaaaggga agggatgcta
     3061 aggtagaggg tgaacgttac agaaaagcag gctgggaagc atatttgaga agatgcggcc
     3121 agcaaaacta aaaaactgta ttataagtaa atgcatgtat actaaactca caaattagag
     3181 cttcaattta attatatcag ttattacccg agtaacgaag catctgtgct tcattttgta
     3241 gaacaaaaat gcaacgcgag agcgctaatt tttcaaacaa agaatctgag ctnnnnnntt
     3301 acagaacaga aatgcaacgc gaaagcgcta ttttaccaac gaagaatctg tgcttcattt
     3361 ttgtaaaaca aaaatgcaac gcgagagcgc taatttttca aacaaagaat ctgagctgca
     3421 tttttacaga acagaaatgc aacgcgagag cgctatttta ccaacaaaga atctatactt
     3481 cttttttgtt ctacaaaaat gcatcccgag agcgctattt ttctaacaaa gcatcttaga
     3541 ttactttttt tctcctttgt gcgctctata atgcagtctc ttgataactt tttgcactgt
     3601 aggtccgtta aggttagaag aaggctactt tggtgtctat tttctcttcc ataaaaaaag
     3661 cctgactcca cttcccgcgt ttactgatta ctagcgaagc tgcgggtgca ttttttcaag
     3721 ataaaggcat ccccgattat attctatacc gatgtggatt gcgcatactt tgtgaacaga
     3781 aagtgatagc gttgatgatt cttcattggt cagaaaatta tgaacggttt cttctatttt
     3841 gtctctatat actacgtata ggaaatgttt acattttcgt attgttttcg attcactcta
     3901 tgaatagttc ttactacaat ttttttgtct aaagagtaat actagagata aacataaaaa
     3961 atgtagaggt cgagtttaga tgcaagttca aggagcgaaa ggtggatggg taggttatat
     4021 agggatatag cacagagata tatagcaaag agatactttt gagcaatgtt tgtggaagcg
     4081 gtattcgcaa tattttagta gctcgttaca gtccggtgcg tttttggttt tttgaaagtg
     4141 cgtcatcaga gcgcttttgg ttttcaaaag cgctctgaag ttcctatact ttctagctag
     4201 agaataggaa cttcccgagc ggccgcgtgt tacaagcttg actccagcgt aactggactg
     4261 caatcaactc actggctcac cttcacgggt gggcctttct tcggtagaaa atcaaaggat
     4321 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
     4381 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aggtaactgg
     4441 cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca
     4501 cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc
     4561 tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga
     4621 taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac
     4681 gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga
     4741 agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag
     4801 ggagcttcca ggggtaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg
     4861 acttgagcat cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag
     4921 caacgcagaa aggcccaccc gaaggtgagc caggtgatta catttgggcc ctcatcagag
     4981 gttttcaccg tcatcaccga aacgcgcgag gcagctgcgg taaagctcat cagcgtggtc
     5041 gtgaagcgat tcacagatgt ctgcctgttc atccgcgtcc agctcgttga gtttctccag
     5101 aagcgttaat gtctggcttc tgataaagcg ggccatgtta agggcggttt tttcctgttt
     5161 ggtcacttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     5221 atccatagtt gcctgactcc ccgtcgtgta gataactacg atgcgggagg gcttaccatc
     5281 tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc
     5341 aataaaccag ccagccggga gggccgagcg cagaagtgat cctgcaactt tatccgcctc
     5401 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
     5461 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc
     5521 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa
     5581 aaaagcggtt agctccttcg gtcctccgat cgttgccaga agtaagttgg ccgcagtgtt
     5641 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtgagatg
     5701 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc
     5761 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa
     5821 agtgctcatc attggaaaac gttcttcggg gcgtaaactc tcaaggatct taccgctgtt
     5881 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
     5941 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
     6001 ggcgacacgg aaatgttgaa tactcatttt agcttcctta gctcctgaaa atctcgataa
     6061 ctcaaaaaat acgcccggta gtgatcttat ttcattatgg tgaaagttgg aacctcttac
     6121 gtgccgatca agtcaaaagc ctccggtcgg aggcttttga ctttctgcta tggaggtcag
     6181 gtatgattta aatggtcagt taga
//
