LOCUS       sequence-297954-        6300 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 6300)
  TITLE     SARS-CoV-2 protein Yeast expression constructs, tagged, for protein 
            expression and/or purification
REFERENCE   2  (bases 1 to 6300)
  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-m4046179
FEATURES             Location/Qualifiers
     source          1..6300
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        25..466
                     /gene="S. cerevisiae GAL1"
                     /label=GAL1 promoter
                     /note="inducible promoter, regulated by Gal4"
     protein_bind    25..142
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     primer_bind     437..460
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     CDS             1975..1992
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             1993..2010
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2055..2073)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     CDS             2424..3227
                     /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      3310..4292
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     terminator      4370..4399
                     /label=T3Te terminator
                     /note="phage T3 early transcription terminator"
     rep_origin      4421..5008
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     terminator      complement(5020..5047)
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     CDS             complement(5071..5262)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(5263..6054,6055..6123))
                     /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     5886..5905
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6124..6226)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     terminator      complement(6227..6258)
                     /label=tonB terminator
                     /note="bidirectional E. coli tonB-P14 transcription 
                     terminator"
ORIGIN
        1 gcggccgctt gctatattga agtacggatt agaagccgcc gagcgggcga cagccctccg
       61 acggaagact ctcctccgtg cgtcctcgtc ttcaccggtc gcgttcctga aacgcagatg
      121 tgcctcgcgc cgcactgctc cgaacaataa agattctaca atactagctt ttatggttat
      181 gaagaggaaa aattggcagt aacctggccc cacaaacctt caaattaacg aatcaaatta
      241 acaaccatag gatgataatg cgattagttt tttagcctta tttctggggt aattaatcag
      301 cgaagcgatg atttttgatc tattaacaga tatataaatg gaaaagctgc ataaccactt
      361 taactaatac tttcaacatt ttcagtttgt attacttctt attcaaatgt cataaaagta
      421 tcaacaaaaa attgttaata tacctctata ctttaacgtc aaggagaaaa aactataatg
      481 attgttaata attggttgaa gcagttaatt aaagttacac ttgtgttcct ttttgttgct
      541 gctattttct atttaataac acctgttcat gtcatgtcta aacatactga cttttcaagt
      601 gaaatcatag gatacaaggc tattgatggt ggtgtcactc gtgacatagc atctacagat
      661 acttgttttg ctaacaaaca tgctgatttt gacacatggt ttagccagcg tggtggtagt
      721 tatactaatg acaaagcttg cccattgatt gctgcagtca taacaagaga agtgggtttt
      781 gtcgtgcctg gtttgcctgg cacgatatta cgcacaacta atggtgactt tttgcatttc
      841 ttacctagag tttttagtgc agttggtaac atctgttaca caccatcaaa acttatagag
      901 tacactgact ttgcaacatc agcttgtgtt ttggctgctg aatgtacaat ttttaaagat
      961 gcttctggta agccagtacc atattgttat gataccaatg tactagaagg ttctgttgct
     1021 tatgaaagtt tacgccctga cacacgttat gtgctcatgg atggctctat tattcaattt
     1081 cctaacacct accttgaagg ttctgttaga gtggtaacaa cttttgattc tgagtactgt
     1141 aggcacggca cttgtgaaag atcagaagct ggtgtttgtg tatctactag tggtagatgg
     1201 gtacttaaca atgattatta cagatcttta ccaggagttt tctgtggtgt agatgctgta
     1261 aatttactta ctaatatgtt tacaccacta attcaaccta ttggtgcttt ggacatatca
     1321 gcatctatag tagctggtgg tattgtagct atcgtagtaa catgccttgc ctactatttt
     1381 atgaggttta gaagagcttt tggtgaatac agtcatgtag ttgcctttaa tactttacta
     1441 ttccttatgt cattcactgt actctgttta acaccagttt actcattctt acctggtgtt
     1501 tattctgtta tttacttgta cttgacattt tatcttacta atgatgtttc ttttttagca
     1561 catattcagt ggatggttat gttcacacct ttagtacctt tctggataac aattgcttat
     1621 atcatttgta tttccacaaa gcatttctat tggttcttta gtaattacct aaagagacgt
     1681 gtagtcttta atggtgtttc ctttagtact tttgaagaag ctgcgctgtg cacctttttg
     1741 ttaaataaag aaatgtatct aaagttgcgt agtgatgtgc tattacctct tacgcaatat
     1801 aatagatact tagctcttta taataagtac aagtatttta gtggagcaat ggatacaact
     1861 agctacagag aagctgcttg ttgtcatctc gcaaaggctc tcaatgactt cagtaactca
     1921 ggttctgatg ttctttacca accaccacaa acctctatca cctcagctgt tttgctcgtg
     1981 ccgagaggtt ctcaccatca tcaccatcat taaacaggcc ccttttcctt tgtcgatatc
     2041 atgtaattag ttatgtcacg cttacattca cgccctcctc ccacatccgc tctaaccgaa
     2101 aaggaaggag ttagacaacc tgaagtctag gtccctattt atttttttta atagttatgt
     2161 tagtattaag aacgttattt atatttcaaa tttttctttt ttttctgtac aaagcggccg
     2221 ccgaaccact agtgcactgc agtacacggt ttccttgaaa tttttttgat tcggtaatct
     2281 ccgaacagaa ggaagaacga aggaaggagc acagacttag attggtatat atacgcatat
     2341 gtagtgttga agaaacatga aattgcccag tattcttaac ccaactgcac agaacaaaaa
     2401 cgtgcaggaa acgaagataa atcatgtcga aagctacata taaggaacgt gctgctactc
     2461 atcctagtcc tgttgctgcc aagctattta atatcatgca cgaaaagcaa acaaacttgt
     2521 gtgcttcatt ggatgttcgt accaccaagg aattactgga gttagttgaa gcattaggtc
     2581 ccaaaatttg tttactaaaa acacatgtgg atatcttgac tgatttttcc atggagggca
     2641 cagttaagcc gctaaaggca ttatccgcca agtacaattt tttactcttc gaggacagaa
     2701 aatttgctga cattggtaat acagtcaaat tgcagtactc tgcgggtgta tacagaatag
     2761 cagaatgggc agacattacg aatgcacacg gtgtggtggg cccaggtatt gttagcggtt
     2821 tgaagcaggc ggcagaagaa gtaacaaagg aacctagagg ccttttgatg ttagcagaat
     2881 tgtcatgcaa gggctcccta tctactggag aatatactaa gggtactgtt gacattgcga
     2941 agagcgacaa agattttgtt atcggcttta ttgctcaaag agacatgggt ggaagagatg
     3001 aaggttacga ttggttgatt atgacacccg gtgtgggttt agatgacaag ggagatgcat
     3061 tgggtcaaca gtatagaacc gtggatgatg tggtttctac aggatctgac attattattg
     3121 ttggaagagg actatttgca aagggaaggg atgctaaggt agagggtgaa cgttacagaa
     3181 aagcaggctg ggaagcatat ttgagaagat gcggccagca aaactaaaaa actgtattat
     3241 aagtaaatgc atgtatacta aactcacaaa ttagagcttc aatttaatta tatcagttat
     3301 tacccgagta acgaagcatc tgtgcttcat tttgtagaac aaaaatgcaa cgcgagagcg
     3361 ctaatttttc aaacaaagaa tctgagctgc atttttacag aacagaaatg caacgcgaaa
     3421 gcgctatttt accaacgaag aatctgtgct tcatttttgt aaaacaaaaa tgcaacgcga
     3481 gagcgctaat ttttcaaaca aagaatctga gctgcatttt tacagaacag aaatgcaacg
     3541 cgagagcgct attttaccaa caaagaatct atacttcttt tttgttctac aaaaatgcat
     3601 cccgagagcg ctatttttct aacaaagcat cttagattac tttttttctc ctttgtgcgc
     3661 tctataatgc agtctcttga taactttttg cactgtaggt ccgttaaggt tagaagaagg
     3721 ctactttggt gtctattttc tcttccataa aaaaagcctg actccacttc ccgcgtttac
     3781 tgattactag cgaagctgcg ggtgcatttt ttcaagataa aggcatcccc gattatattc
     3841 tataccgatg tggattgcgc atactttgtg aacagaaagt gatagcgttg atgattcttc
     3901 attggtcaga aaattatgaa cggtttcttc tattttgtct ctatatacta cgtataggaa
     3961 atgtttacat tttcgtattg ttttcgattc actctatgaa tagttcttac tacaattttt
     4021 ttgtctaaag agtaatacta gagataaaca taaaaaatgt agaggtcgag tttagatgca
     4081 agttcaagga gcgaaaggtg gatgggtagg ttatataggg atatagcaca gagatatata
     4141 gcaaagagat acttttgagc aatgtttgtg gaagcggtat tcgcaatatt ttagtagctc
     4201 gttacagtcc ggtgcgtttt tggttttttg aaagtgcgtc atcagagcgc ttttggtttt
     4261 caaaagcgct ctgaagttcc tatactttct agctagagaa taggaacttc ccgagcggcc
     4321 gcgtgttaca agcttgactc cagcgtaact ggactgcaat caactcactg gctcaccttc
     4381 acgggtgggc ctttcttcgg tagaaaatca aaggatcttc ttgagatcct ttttttctgc
     4441 gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg
     4501 atcaagagct accaactctt tttccgaggt aactggcttc agcagagcgc agataccaaa
     4561 tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc
     4621 tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg
     4681 tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac
     4741 ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct
     4801 acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc
     4861 ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg taaacgcctg
     4921 gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcatcgat ttttgtgatg
     4981 ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcagaaaggc ccacccgaag
     5041 gtgagccagg tgattacatt tgggccctca tcagaggttt tcaccgtcat caccgaaacg
     5101 cgcgaggcag ctgcggtaaa gctcatcagc gtggtcgtga agcgattcac agatgtctgc
     5161 ctgttcatcc gcgtccagct cgttgagttt ctccagaagc gttaatgtct ggcttctgat
     5221 aaagcgggcc atgttaaggg cggttttttc ctgtttggtc acttaccaat gcttaatcag
     5281 tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt
     5341 cgtgtagata actacgatgc gggagggctt accatctggc cccagtgctg caatgatacc
     5401 gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag ccgggagggc
     5461 cgagcgcaga agtgatcctg caactttatc cgcctccatc cagtctatta attgttgccg
     5521 ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac
     5581 aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg
     5641 atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc
     5701 tccgatcgtt gccagaagta agttggccgc agtgttatca ctcatggtta tggcagcact
     5761 gcataattct cttactgtca tgccatccgt gagatgcttt tctgtgactg gtgagtactc
     5821 aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat
     5881 acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc
     5941 ttcggggcgt aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac
     6001 tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa
     6061 aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact
     6121 cattttagct tccttagctc ctgaaaatct cgataactca aaaaatacgc ccggtagtga
     6181 tcttatttca ttatggtgaa agttggaacc tcttacgtgc cgatcaagtc aaaagcctcc
     6241 ggtcggaggc ttttgacttt ctgctatgga ggtcaggtat gatttaaatg gtcagttaga
//
