LOCUS       sequence-300501-        6259 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 6259)
  TITLE     SARS-CoV-2 protein Yeast expression constructs, tagged, for protein 
            expression and/or purification
REFERENCE   2  (bases 1 to 6259)
  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-m4046439
FEATURES             Location/Qualifiers
     source          1..6259
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(320..1123)
                     /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"
     primer_bind     1474..1492
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     CDS             complement(1537..1554)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(1555..1572)
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     promoter        complement(3089..3502)
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
     terminator      3548..3579
                     /label=tonB terminator
                     /note="bidirectional E. coli tonB-P14 transcription 
                     terminator"
     promoter        3580..3682
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     CDS             join(3683..3751,3752..4543)
                     /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     complement(3901..3920)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     CDS             4544..4735
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     terminator      4759..4786
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     rep_origin      complement(4798..5385)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     terminator      complement(5407..5436)
                     /label=T3Te terminator
                     /note="phage T3 early transcription terminator"
     rep_origin      join(5514..6259,1..237)
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
ORIGIN
        1 ctctcgcgtt gcatttctgt tctgtaaaaa tgcagctcag attctttgtt tgaaaaatta
       61 gcgctctcgc gttgcatttt tgttttacaa aaatgaagca cagattcttc gttggtaaaa
      121 tagcgctttc gcgttgcatt tctgttctgt aaaaatgcag ctcagattct ttgtttgaaa
      181 aattagcgct ctcgcgttgc atttttgttc tacaaaatga agcacagatg cttcgttact
      241 cgggtaataa ctgatataat taaattgaag ctctaatttg tgagtttagt atacatgcat
      301 ttacttataa tacagttttt tagttttgct ggccgcatct tctcaaatat gcttcccagc
      361 ctgcttttct gtaacgttca ccctctacct tagcatccct tccctttgca aatagtcctc
      421 ttccaacaat aataatgtca gatcctgtag aaaccacatc atccacggtt ctatactgtt
      481 gacccaatgc atctcccttg tcatctaaac ccacaccggg tgtcataatc aaccaatcgt
      541 aaccttcatc tcttccaccc atgtctcttt gagcaataaa gccgataaca aaatctttgt
      601 cgctcttcgc aatgtcaaca gtacccttag tatattctcc agtagatagg gagcccttgc
      661 atgacaattc tgctaacatc aaaaggcctc taggttcctt tgttacttct tctgccgcct
      721 gcttcaaacc gctaacaata cctgggccca ccacaccgtg tgcattcgta atgtctgccc
      781 attctgctat tctgtataca cccgcagagt actgcaattt gactgtatta ccaatgtcag
      841 caaattttct gtcctcgaag agtaaaaaat tgtacttggc ggataatgcc tttagcggct
      901 taactgtgcc ctccatggaa aaatcagtca agatatccac atgtgttttt agtaaacaaa
      961 ttttgggacc taatgcttca actaactcca gtaattcctt ggtggtacga acatccaatg
     1021 aagcacacaa gtttgtttgc ttttcgtgca tgatattaaa tagcttggca gcaacaggac
     1081 taggatgagt agcagcacgt tccttatatg tagctttcga catgatttat cttcgtttcc
     1141 tgcacgtttt tgttctgtgc agttgggtta agaatactgg gcaatttcat gtttcttcaa
     1201 cactacatat gcgtatatat accaatctaa gtctgtgctc cttccttcgt tcttccttct
     1261 gttcggagat taccgaatca aaaaaatttc aaggaaaccg tgtactgcag tgcactagtg
     1321 gttcggcggc cgctttgtac agaaaaaaaa gaaaaatttg aaatataaat aacgttctta
     1381 atactaacat aactattaaa aaaaataaat agggacctag acttcaggtt gtctaactcc
     1441 ttccttttcg gttagagcgg atgtgggagg agggcgtgaa tgtaagcgtg acataactaa
     1501 ttacatgata tcgacaaagg aaaaggggcc tgtttaatga tggtgatgat ggtgagaacc
     1561 tctcggcacg agttgcaaaa cagcggaggt aatagacgtc tgaggtggtt gatacaagac
     1621 gtcggaaccg gaattagaga aatcattcaa agccttagca aggtgacaac aagccgcttc
     1681 acggtatgaa gtagtatcca tggcacccga gaagtatttg tacttgttgt ataaagctaa
     1741 atatctgtta tattgggtta gcgggagtaa aacgtcagat ctcaacttca agtacatttc
     1801 tttgttgagc aaaaaggtac acaaggcagc ttcttcgaag gtagaaaagc tgacaccatt
     1861 aaagacaact cttctcttta gataatttga gaaaaaccaa tagaaatgtt tagtacttat
     1921 gcaaatgatg taagcgatcg ttatccagaa gggaaccaat ggggtgaaca ttaccatcca
     1981 ttgaatgtgt gccaaaaagc tgacatcgtt agtcaggtag aatgtcaggt ataaatagat
     2041 aacggagtat acaccaggca agaaagagta tactggtgta agacacaaaa cagtgaacga
     2101 catcaaaaat aatagagtgt tgaaagccac gacatgcgaa tactccccaa aggctcttct
     2161 aaatctcatg aagtagtaag ccaaacaagt aacaacgata gcaacgattc caccggcgac
     2221 aatagatgcg gaaatgtcca gcgcacctat tggttggatc aaaggggtaa acatgttagt
     2281 caatagatta acagcatcga caccgcagaa gacacctgga agagaacggt aataatcgtt
     2341 attcaacacc catctaccgc tggtagaaac acagacaccg gcttctgatc tttcacaagt
     2401 accgtggcga cagtactcgg agtcgaaggt ggtgacaacc cgaacagaac cttcaaggta
     2461 tgtgtttgga aattgaatga ttgatccatc cattaacaca tatctagtgt ccggtcgcaa
     2521 ggattcgtaa gcaacagaac cttcgaggac gtttgtgtca taacagtatg gaactggctt
     2581 accacttgca tccttgaaaa tagtgcattc agcagccaat acacaagcag aggtggcaaa
     2641 atcagtgtat tcgatcaatt tgctaggggt ataacaaata ttgccaacag cagagaagac
     2701 tcttggtaag aaatgcaaaa aatcaccatt ggtggtacgc agtatagtcc ctggtaaacc
     2761 tggcacgacg aaaccaactt ctctagtgat aacagctgca atcaaggggc aggccttgtc
     2821 gtttgtgtag gaaccacccc tttgagagaa ccaagtatca aagtccgcgt gcttgttagc
     2881 gaaacaggtg tccgtcgaag caatatctct agtaactccg ccatcgatag ctttgtaacc
     2941 aatgatctct gaggaaaagt cggtatgctt actcatcacg tggacaggtg taattagata
     3001 gaagatggct gcaacgaaca aaaatactaa agtaaccttt atcaactgtt taagccaatt
     3061 gttgacaatc ttcattttgt aattaaaact tagattagat tgctatgctt tctttctaat
     3121 gagcaagaag taaaaaaagt tgtaatagaa caagaaaaat gaaactgaaa cttgagaaat
     3181 tgaagaccgt ttattaactt aaatatcaat gggaggtcat cgaaagagaa aaaaatcaaa
     3241 aaaaaaaatt ttcaagaaaa agaaacgtga taaaaatttt tattgccttt ttcgacgaag
     3301 aaaaagaaac gaggcggtct cttttttctt ttccaaacct ttagtacggg taattaacga
     3361 caccctagag gaagaaagag gggaaattta gtatgctgtg cttgggtgtt ttgaagtggt
     3421 acggcgatgc gcggagtccg agaaaatctg gaagagtaaa aaaggagtag aaacattttg
     3481 aagctatggt gtgtggggcg gccgctctaa ctgaccattt aaatcatacc tgacctccat
     3541 agcagaaagt caaaagcctc cgaccggagg cttttgactt gatcggcacg taagaggttc
     3601 caactttcac cataatgaaa taagatcact accgggcgta ttttttgagt tatcgagatt
     3661 ttcaggagct aaggaagcta aaatgagtat tcaacatttc cgtgtcgccc ttattccctt
     3721 ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga
     3781 tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa
     3841 gatccttgag agtttacgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct
     3901 gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat
     3961 acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc atctcacgga
     4021 tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc
     4081 caacttactt ctggcaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat
     4141 gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa
     4201 cgacgagcgt gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac
     4261 tggcgaacta cttactctag cttcccggca acaattaata gactggatgg aggcggataa
     4321 agttgcagga tcacttctgc gctcggccct cccggctggc tggtttattg ctgataaatc
     4381 tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc
     4441 ctcccgcatc gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag
     4501 acagatcgct gagataggtg cctcactgat taagcattgg taagtgacca aacaggaaaa
     4561 aaccgccctt aacatggccc gctttatcag aagccagaca ttaacgcttc tggagaaact
     4621 caacgagctg gacgcggatg aacaggcaga catctgtgaa tcgcttcacg accacgctga
     4681 tgagctttac cgcagctgcc tcgcgcgttt cggtgatgac ggtgaaaacc tctgatgagg
     4741 gcccaaatgt aatcacctgg ctcaccttcg ggtgggcctt tctgcgttgc tggcgttttt
     4801 ccataggctc cgcccccctg acgagcatca caaaaatcga tgctcaagtc agaggtggcg
     4861 aaacccgaca ggactataaa gataccaggc gtttacccct ggaagctccc tcgtgcgctc
     4921 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     4981 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     5041 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     5101 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     5161 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     5221 ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctc
     5281 ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt
     5341 tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatt
     5401 ttctaccgaa gaaaggccca cccgtgaagg tgagccagtg agttgattgc agtccagtta
     5461 cgctggagtc aagcttgtaa cacgcggccg ctcgggaagt tcctattctc tagctagaaa
     5521 gtataggaac ttcagagcgc ttttgaaaac caaaagcgct ctgatgacgc actttcaaaa
     5581 aaccaaaaac gcaccggact gtaacgagct actaaaatat tgcgaatacc gcttccacaa
     5641 acattgctca aaagtatctc tttgctatat atctctgtgc tatatcccta tataacctac
     5701 ccatccacct ttcgctcctt gaacttgcat ctaaactcga cctctacatt ttttatgttt
     5761 atctctagta ttactcttta gacaaaaaaa ttgtagtaag aactattcat agagtgaatc
     5821 gaaaacaata cgaaaatgta aacatttcct atacgtagta tatagagaca aaatagaaga
     5881 aaccgttcat aattttctga ccaatgaaga atcatcaacg ctatcacttt ctgttcacaa
     5941 agtatgcgca atccacatcg gtatagaata taatcgggga tgcctttatc ttgaaaaaat
     6001 gcacccgcag cttcgctagt aatcagtaaa cgcgggaagt ggagtcaggc tttttttatg
     6061 gaagagaaaa tagacaccaa agtagccttc ttctaacctt aacggaccta cagtgcaaaa
     6121 agttatcaag agactgcatt atagagcgca caaaggagaa aaaaagtaat ctaagatgct
     6181 ttgttagaaa aatagcgctc tcgggatgca tttttgtaga acaaaaaaga agtatagatt
     6241 ctttgttggt aaaatagcg
//
