LOCUS       sequence-297778-        6265 bp    DNA     circular SYN 15-APR-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 6265)
  TITLE     SARS-CoV-2 protein Yeast expression constructs, tagged, for protein 
            expression and/or purification
REFERENCE   2  (bases 1 to 6265)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: pGBW-m4046524
FEATURES             Location/Qualifiers
     source          1..6265
                     /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"
     CDS             1952..1975
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     primer_bind     complement(2020..2038)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     CDS             2389..3192
                     /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      3275..4257
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     terminator      4335..4364
                     /label=T3Te terminator
                     /note="phage T3 early transcription terminator"
     rep_origin      4386..4973
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     terminator      complement(4985..5012)
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     CDS             complement(5036..5227)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(5228..6019,6020..6088))
                     /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     5851..5870
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6089..6191)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     terminator      complement(6192..6223)
                     /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 ttggagccat ccgcagtttg aaaaataaac
     1981 aggccccttt tcctttgtcg atatcatgta attagttatg tcacgcttac attcacgccc
     2041 tcctcccaca tccgctctaa ccgaaaagga aggagttaga caacctgaag tctaggtccc
     2101 tatttatttt ttttaatagt tatgttagta ttaagaacgt tatttatatt tcaaattttt
     2161 cttttttttc tgtacaaagc ggccgccgaa ccactagtgc actgcagtac acggtttcct
     2221 tgaaattttt ttgattcggt aatctccgaa cagaaggaag aacgaaggaa ggagcacaga
     2281 cttagattgg tatatatacg catatgtagt gttgaagaaa catgaaattg cccagtattc
     2341 ttaacccaac tgcacagaac aaaaacgtgc aggaaacgaa gataaatcat gtcgaaagct
     2401 acatataagg aacgtgctgc tactcatcct agtcctgttg ctgccaagct atttaatatc
     2461 atgcacgaaa agcaaacaaa cttgtgtgct tcattggatg ttcgtaccac caaggaatta
     2521 ctggagttag ttgaagcatt aggtcccaaa atttgtttac taaaaacaca tgtggatatc
     2581 ttgactgatt tttccatgga gggcacagtt aagccgctaa aggcattatc cgccaagtac
     2641 aattttttac tcttcgagga cagaaaattt gctgacattg gtaatacagt caaattgcag
     2701 tactctgcgg gtgtatacag aatagcagaa tgggcagaca ttacgaatgc acacggtgtg
     2761 gtgggcccag gtattgttag cggtttgaag caggcggcag aagaagtaac aaaggaacct
     2821 agaggccttt tgatgttagc agaattgtca tgcaagggct ccctatctac tggagaatat
     2881 actaagggta ctgttgacat tgcgaagagc gacaaagatt ttgttatcgg ctttattgct
     2941 caaagagaca tgggtggaag agatgaaggt tacgattggt tgattatgac acccggtgtg
     3001 ggtttagatg acaagggaga tgcattgggt caacagtata gaaccgtgga tgatgtggtt
     3061 tctacaggat ctgacattat tattgttgga agaggactat ttgcaaaggg aagggatgct
     3121 aaggtagagg gtgaacgtta cagaaaagca ggctgggaag catatttgag aagatgcggc
     3181 cagcaaaact aaaaaactgt attataagta aatgcatgta tactaaactc acaaattaga
     3241 gcttcaattt aattatatca gttattaccc gagtaacgaa gcatctgtgc ttcattttgt
     3301 agaacaaaaa tgcaacgcga gagcgctaat ttttcaaaca aagaatctga gctgcatttt
     3361 tacagaacag aaatgcaacg cgaaagcgct attttaccaa cgaagaatct gtgcttcatt
     3421 tttgtaaaac aaaaatgcaa cgcgagagcg ctaatttttc aaacaaagaa tctgagctgc
     3481 atttttacag aacagaaatg caacgcgaga gcgctatttt accaacaaag aatctatact
     3541 tcttttttgt tctacaaaaa tgcatcccga gagcgctatt tttctaacaa agcatcttag
     3601 attacttttt ttctcctttg tgcgctctat aatgcagtct cttgataact ttttgcactg
     3661 taggtccgtt aaggttagaa gaaggctact ttggtgtcta ttttctcttc cataaaaaaa
     3721 gcctgactcc acttcccgcg tttactgatt actagcgaag ctgcgggtgc attttttcaa
     3781 gataaaggca tccccgatta tattctatac cgatgtggat tgcgcatact ttgtgaacag
     3841 aaagtgatag cgttgatgat tcttcattgg tcagaaaatt atgaacggtt tcttctattt
     3901 tgtctctata tactacgtat aggaaatgtt tacattttcg tattgttttc gattcactct
     3961 atgaatagtt cttactacaa tttttttgtc taaagagtaa tactagagat aaacataaaa
     4021 aatgtagagg tcgagtttag atgcaagttc aaggagcgaa aggtggatgg gtaggttata
     4081 tagggatata gcacagagat atatagcaaa gagatacttt tgagcaatgt ttgtggaagc
     4141 ggtattcgca atattttagt agctcgttac agtccggtgc gtttttggtt ttttgaaagt
     4201 gcgtcatcag agcgcttttg gttttcaaaa gcgctctgaa gttcctatac tttctagcta
     4261 gagaatagga acttcccgag cggccgcgtg ttacaagctt gactccagcg taactggact
     4321 gcaatcaact cactggctca ccttcacggg tgggcctttc ttcggtagaa aatcaaagga
     4381 tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg
     4441 ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaggtaactg
     4501 gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc
     4561 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg
     4621 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
     4681 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
     4741 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg
     4801 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
     4861 gggagcttcc aggggtaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
     4921 gacttgagca tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca
     4981 gcaacgcaga aaggcccacc cgaaggtgag ccaggtgatt acatttgggc cctcatcaga
     5041 ggttttcacc gtcatcaccg aaacgcgcga ggcagctgcg gtaaagctca tcagcgtggt
     5101 cgtgaagcga ttcacagatg tctgcctgtt catccgcgtc cagctcgttg agtttctcca
     5161 gaagcgttaa tgtctggctt ctgataaagc gggccatgtt aagggcggtt ttttcctgtt
     5221 tggtcactta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
     5281 catccatagt tgcctgactc cccgtcgtgt agataactac gatgcgggag ggcttaccat
     5341 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag
     5401 caataaacca gccagccggg agggccgagc gcagaagtga tcctgcaact ttatccgcct
     5461 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
     5521 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg
     5581 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca
     5641 aaaaagcggt tagctccttc ggtcctccga tcgttgccag aagtaagttg gccgcagtgt
     5701 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtgagat
     5761 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac
     5821 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa
     5881 aagtgctcat cattggaaaa cgttcttcgg ggcgtaaact ctcaaggatc ttaccgctgt
     5941 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
     6001 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
     6061 gggcgacacg gaaatgttga atactcattt tagcttcctt agctcctgaa aatctcgata
     6121 actcaaaaaa tacgcccggt agtgatctta tttcattatg gtgaaagttg gaacctctta
     6181 cgtgccgatc aagtcaaaag cctccggtcg gaggcttttg actttctgct atggaggtca
     6241 ggtatgattt aaatggtcag ttaga
//
