LOCUS       sequence-313646-        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-m4046327
FEATURES             Location/Qualifiers
     source          1..6259
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..408
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
     CDS             1919..1936
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             1937..1960
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     primer_bind     complement(2005..2023)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     CDS             2374..3177
                     /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      3260..4242
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     terminator      4320..4349
                     /label=T3Te terminator
                     /note="phage T3 early transcription terminator"
     rep_origin      4371..4958
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     terminator      complement(4970..4997)
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     CDS             complement(5021..5212)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(5213..6004,6005..6073))
                     /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     5836..5855
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6074..6176)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     terminator      complement(6177..6208)
                     /label=tonB terminator
                     /note="bidirectional E. coli tonB-P14 transcription 
                     terminator"
ORIGIN
        1 ccacacacca tagcttcaaa atgtttctac tcctttttta ctcttccaga ttttctcgga
       61 ctccgcgcat cgccgtacca cttcaaaaca cccaagcaca gcatactaaa tttcccctct
      121 ttcttcctct agggtgtcgt taattacccg tactaaaggt ttggaaaaga aaaaagagac
      181 cgcctcgttt ctttttcttc gtcgaaaaag gcaataaaaa tttttatcac gtttcttttt
      241 cttgaaaatt tttttttttg atttttttct ctttcgatga cctcccattg atatttaagt
      301 taataaacgg tcttcaattt ctcaagtttc agtttcattt ttcttgttct attacaactt
      361 tttttacttc ttgctcatta gaaagaaagc atagcaatct aatctaagtt ttaattacaa
      421 aatgattgtt aataattggt tgaagcagtt aattaaagtt acacttgtgt tcctttttgt
      481 tgctgctatt ttctatttaa taacacctgt tcatgtcatg tctaaacata ctgacttttc
      541 aagtgaaatc ataggataca aggctattga tggtggtgtc actcgtgaca tagcatctac
      601 agatacttgt tttgctaaca aacatgctga ttttgacaca tggtttagcc agcgtggtgg
      661 tagttatact aatgacaaag cttgcccatt gattgctgca gtcataacaa gagaagtggg
      721 ttttgtcgtg cctggtttgc ctggcacgat attacgcaca actaatggtg actttttgca
      781 tttcttacct agagttttta gtgcagttgg taacatctgt tacacaccat caaaacttat
      841 agagtacact gactttgcaa catcagcttg tgttttggct gctgaatgta caatttttaa
      901 agatgcttct ggtaagccag taccatattg ttatgatacc aatgtactag aaggttctgt
      961 tgcttatgaa agtttacgcc ctgacacacg ttatgtgctc atggatggct ctattattca
     1021 atttcctaac acctaccttg aaggttctgt tagagtggta acaacttttg attctgagta
     1081 ctgtaggcac ggcacttgtg aaagatcaga agctggtgtt tgtgtatcta ctagtggtag
     1141 atgggtactt aacaatgatt attacagatc tttaccagga gttttctgtg gtgtagatgc
     1201 tgtaaattta cttactaata tgtttacacc actaattcaa cctattggtg ctttggacat
     1261 atcagcatct atagtagctg gtggtattgt agctatcgta gtaacatgcc ttgcctacta
     1321 ttttatgagg tttagaagag cttttggtga atacagtcat gtagttgcct ttaatacttt
     1381 actattcctt atgtcattca ctgtactctg tttaacacca gtttactcat tcttacctgg
     1441 tgtttattct gttatttact tgtacttgac attttatctt actaatgatg tttctttttt
     1501 agcacatatt cagtggatgg ttatgttcac acctttagta cctttctgga taacaattgc
     1561 ttatatcatt tgtatttcca caaagcattt ctattggttc tttagtaatt acctaaagag
     1621 acgtgtagtc tttaatggtg tttcctttag tacttttgaa gaagctgcgc tgtgcacctt
     1681 tttgttaaat aaagaaatgt atctaaagtt gcgtagtgat gtgctattac ctcttacgca
     1741 atataataga tacttagctc tttataataa gtacaagtat tttagtggag caatggatac
     1801 aactagctac agagaagctg cttgttgtca tctcgcaaag gctctcaatg acttcagtaa
     1861 ctcaggttct gatgttcttt accaaccacc acaaacctct atcacctcag ctgttttgct
     1921 cgtgccgaga ggttcttgga gccatccgca gtttgaaaaa taaacaggcc ccttttcctt
     1981 tgtcgatatc atgtaattag ttatgtcacg cttacattca cgccctcctc ccacatccgc
     2041 tctaaccgaa aaggaaggag ttagacaacc tgaagtctag gtccctattt atttttttta
     2101 atagttatgt tagtattaag aacgttattt atatttcaaa tttttctttt ttttctgtac
     2161 aaagcggccg ccgaaccact agtgcactgc agtacacggt ttccttgaaa tttttttgat
     2221 tcggtaatct ccgaacagaa ggaagaacga aggaaggagc acagacttag attggtatat
     2281 atacgcatat gtagtgttga agaaacatga aattgcccag tattcttaac ccaactgcac
     2341 agaacaaaaa cgtgcaggaa acgaagataa atcatgtcga aagctacata taaggaacgt
     2401 gctgctactc atcctagtcc tgttgctgcc aagctattta atatcatgca cgaaaagcaa
     2461 acaaacttgt gtgcttcatt ggatgttcgt accaccaagg aattactgga gttagttgaa
     2521 gcattaggtc ccaaaatttg tttactaaaa acacatgtgg atatcttgac tgatttttcc
     2581 atggagggca cagttaagcc gctaaaggca ttatccgcca agtacaattt tttactcttc
     2641 gaggacagaa aatttgctga cattggtaat acagtcaaat tgcagtactc tgcgggtgta
     2701 tacagaatag cagaatgggc agacattacg aatgcacacg gtgtggtggg cccaggtatt
     2761 gttagcggtt tgaagcaggc ggcagaagaa gtaacaaagg aacctagagg ccttttgatg
     2821 ttagcagaat tgtcatgcaa gggctcccta tctactggag aatatactaa gggtactgtt
     2881 gacattgcga agagcgacaa agattttgtt atcggcttta ttgctcaaag agacatgggt
     2941 ggaagagatg aaggttacga ttggttgatt atgacacccg gtgtgggttt agatgacaag
     3001 ggagatgcat tgggtcaaca gtatagaacc gtggatgatg tggtttctac aggatctgac
     3061 attattattg ttggaagagg actatttgca aagggaaggg atgctaaggt agagggtgaa
     3121 cgttacagaa aagcaggctg ggaagcatat ttgagaagat gcggccagca aaactaaaaa
     3181 actgtattat aagtaaatgc atgtatacta aactcacaaa ttagagcttc aatttaatta
     3241 tatcagttat tacccgagta acgaagcatc tgtgcttcat tttgtagaac aaaaatgcaa
     3301 cgcgagagcg ctaatttttc aaacaaagaa tctgagctgc atttttacag aacagaaatg
     3361 caacgcgaaa gcgctatttt accaacgaag aatctgtgct tcatttttgt aaaacaaaaa
     3421 tgcaacgcga gagcgctaat ttttcaaaca aagaatctga gctgcatttt tacagaacag
     3481 aaatgcaacg cgagagcgct attttaccaa caaagaatct atacttcttt tttgttctac
     3541 aaaaatgcat cccgagagcg ctatttttct aacaaagcat cttagattac tttttttctc
     3601 ctttgtgcgc tctataatgc agtctcttga taactttttg cactgtaggt ccgttaaggt
     3661 tagaagaagg ctactttggt gtctattttc tcttccataa aaaaagcctg actccacttc
     3721 ccgcgtttac tgattactag cgaagctgcg ggtgcatttt ttcaagataa aggcatcccc
     3781 gattatattc tataccgatg tggattgcgc atactttgtg aacagaaagt gatagcgttg
     3841 atgattcttc attggtcaga aaattatgaa cggtttcttc tattttgtct ctatatacta
     3901 cgtataggaa atgtttacat tttcgtattg ttttcgattc actctatgaa tagttcttac
     3961 tacaattttt ttgtctaaag agtaatacta gagataaaca taaaaaatgt agaggtcgag
     4021 tttagatgca agttcaagga gcgaaaggtg gatgggtagg ttatataggg atatagcaca
     4081 gagatatata gcaaagagat acttttgagc aatgtttgtg gaagcggtat tcgcaatatt
     4141 ttagtagctc gttacagtcc ggtgcgtttt tggttttttg aaagtgcgtc atcagagcgc
     4201 ttttggtttt caaaagcgct ctgaagttcc tatactttct agctagagaa taggaacttc
     4261 ccgagcggcc gcgtgttaca agcttgactc cagcgtaact ggactgcaat caactcactg
     4321 gctcaccttc acgggtgggc ctttcttcgg tagaaaatca aaggatcttc ttgagatcct
     4381 ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt
     4441 tgtttgccgg atcaagagct accaactctt tttccgaggt aactggcttc agcagagcgc
     4501 agataccaaa tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg
     4561 tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg
     4621 ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt
     4681 cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac
     4741 tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg
     4801 acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg
     4861 taaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcatcgat
     4921 ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcagaaaggc
     4981 ccacccgaag gtgagccagg tgattacatt tgggccctca tcagaggttt tcaccgtcat
     5041 caccgaaacg cgcgaggcag ctgcggtaaa gctcatcagc gtggtcgtga agcgattcac
     5101 agatgtctgc ctgttcatcc gcgtccagct cgttgagttt ctccagaagc gttaatgtct
     5161 ggcttctgat aaagcgggcc atgttaaggg cggttttttc ctgtttggtc acttaccaat
     5221 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     5281 gactccccgt cgtgtagata actacgatgc gggagggctt accatctggc cccagtgctg
     5341 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
     5401 ccgggagggc cgagcgcaga agtgatcctg caactttatc cgcctccatc cagtctatta
     5461 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     5521 ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     5581 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     5641 ccttcggtcc tccgatcgtt gccagaagta agttggccgc agtgttatca ctcatggtta
     5701 tggcagcact gcataattct cttactgtca tgccatccgt gagatgcttt tctgtgactg
     5761 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     5821 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     5881 gaaaacgttc ttcggggcgt aaactctcaa ggatcttacc gctgttgaga tccagttcga
     5941 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     6001 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     6061 gttgaatact cattttagct tccttagctc ctgaaaatct cgataactca aaaaatacgc
     6121 ccggtagtga tcttatttca ttatggtgaa agttggaacc tcttacgtgc cgatcaagtc
     6181 aaaagcctcc ggtcggaggc ttttgacttt ctgctatgga ggtcaggtat gatttaaatg
     6241 gtcagttaga gcggccgcc
//
