LOCUS       sequence-287570-        5395 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Yeast Expression plasmid for SARS-CoV-2 nsp8.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5395)
  TITLE     SARS-CoV-2 protein Yeast expression constructs, tagged, for protein 
            expression and/or purification
REFERENCE   2  (bases 1 to 5395)
  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-m4046257
FEATURES             Location/Qualifiers
     source          1..5395
                     /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             1022..1039
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             1040..1105
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     primer_bind     complement(1150..1168)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     CDS             1519..2322
                     /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      2405..3387
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     terminator      3465..3494
                     /label=T3Te terminator
                     /note="phage T3 early transcription terminator"
     rep_origin      3516..4103
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     terminator      complement(4115..4142)
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     CDS             complement(4166..4357)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(4358..5149,5150..5218))
                     /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     4981..5000
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5219..5321)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     terminator      complement(5322..5353)
                     /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 atgatagcct cagagtttag ttcccttcca tcatatgcag cttttgctac
      481 tgctcaagaa gcttatgagc aggctgttgc taatggtgat tctgaagttg ttcttaaaaa
      541 gttgaagaag tctttgaatg tggctaaatc tgaatttgac cgtgatgcag ccatgcaacg
      601 taagttggaa aagatggctg atcaagctat gacccaaatg tataaacagg ctagatctga
      661 ggacaagagg gcaaaagtta ctagtgctat gcagacaatg cttttcacta tgcttagaaa
      721 gttggataat gatgcactca acaacattat caacaatgca agagatggtt gtgttccctt
      781 gaacataata cctcttacaa cagcagccaa actaatggtt gtcataccag actataacac
      841 atataaaaat acgtgtgatg gtacaacatt tacttatgca tcagcattgt gggaaatcca
      901 acaggttgta gatgcagata gtaaaattgt tcaacttagt gaaattagta tggacaattc
      961 acctaattta gcatggcctc ttattgtaac agctttaagg gccaattctg ctgtcaaatt
     1021 actcgtgccg agaggttctg attataaaga tcatgatggt gattataaag atcatgatat
     1081 tgattataaa gatgatgatg ataaataaac aggccccttt tcctttgtcg atatcatgta
     1141 attagttatg tcacgcttac attcacgccc tcctcccaca tccgctctaa ccgaaaagga
     1201 aggagttaga caacctgaag tctaggtccc tatttatttt ttttaatagt tatgttagta
     1261 ttaagaacgt tatttatatt tcaaattttt cttttttttc tgtacaaagc ggccgccgaa
     1321 ccactagtgc actgcagtac acggtttcct tgaaattttt ttgattcggt aatctccgaa
     1381 cagaaggaag aacgaaggaa ggagcacaga cttagattgg tatatatacg catatgtagt
     1441 gttgaagaaa catgaaattg cccagtattc ttaacccaac tgcacagaac aaaaacgtgc
     1501 aggaaacgaa gataaatcat gtcgaaagct acatataagg aacgtgctgc tactcatcct
     1561 agtcctgttg ctgccaagct atttaatatc atgcacgaaa agcaaacaaa cttgtgtgct
     1621 tcattggatg ttcgtaccac caaggaatta ctggagttag ttgaagcatt aggtcccaaa
     1681 atttgtttac taaaaacaca tgtggatatc ttgactgatt tttccatgga gggcacagtt
     1741 aagccgctaa aggcattatc cgccaagtac aattttttac tcttcgagga cagaaaattt
     1801 gctgacattg gtaatacagt caaattgcag tactctgcgg gtgtatacag aatagcagaa
     1861 tgggcagaca ttacgaatgc acacggtgtg gtgggcccag gtattgttag cggtttgaag
     1921 caggcggcag aagaagtaac aaaggaacct agaggccttt tgatgttagc agaattgtca
     1981 tgcaagggct ccctatctac tggagaatat actaagggta ctgttgacat tgcgaagagc
     2041 gacaaagatt ttgttatcgg ctttattgct caaagagaca tgggtggaag agatgaaggt
     2101 tacgattggt tgattatgac acccggtgtg ggtttagatg acaagggaga tgcattgggt
     2161 caacagtata gaaccgtgga tgatgtggtt tctacaggat ctgacattat tattgttgga
     2221 agaggactat ttgcaaaggg aagggatgct aaggtagagg gtgaacgtta cagaaaagca
     2281 ggctgggaag catatttgag aagatgcggc cagcaaaact aaaaaactgt attataagta
     2341 aatgcatgta tactaaactc acaaattaga gcttcaattt aattatatca gttattaccc
     2401 gagtaacgaa gcatctgtgc ttcattttgt agaacaaaaa tgcaacgcga gagcgctaat
     2461 ttttcaaaca aagaatctga gctgcatttt tacagaacag aaatgcaacg cgaaagcgct
     2521 attttaccaa cgaagaatct gtgcttcatt tttgtaaaac aaaaatgcaa cgcgagagcg
     2581 ctaatttttc aaacaaagaa tctgagctgc atttttacag aacagaaatg caacgcgaga
     2641 gcgctatttt accaacaaag aatctatact tcttttttgt tctacaaaaa tgcatcccga
     2701 gagcgctatt tttctaacaa agcatcttag attacttttt ttctcctttg tgcgctctat
     2761 aatgcagtct cttgataact ttttgcactg taggtccgtt aaggttagaa gaaggctact
     2821 ttggtgtcta ttttctcttc cataaaaaaa gcctgactcc acttcccgcg tttactgatt
     2881 actagcgaag ctgcgggtgc attttttcaa gataaaggca tccccgatta tattctatac
     2941 cgatgtggat tgcgcatact ttgtgaacag aaagtgatag cgttgatgat tcttcattgg
     3001 tcagaaaatt atgaacggtt tcttctattt tgtctctata tactacgtat aggaaatgtt
     3061 tacattttcg tattgttttc gattcactct atgaatagtt cttactacaa tttttttgtc
     3121 taaagagtaa tactagagat aaacataaaa aatgtagagg tcgagtttag atgcaagttc
     3181 aaggagcgaa aggtggatgg gtaggttata tagggatata gcacagagat atatagcaaa
     3241 gagatacttt tgagcaatgt ttgtggaagc ggtattcgca atattttagt agctcgttac
     3301 agtccggtgc gtttttggtt ttttgaaagt gcgtcatcag agcgcttttg gttttcaaaa
     3361 gcgctctgaa gttcctatac tttctagcta gagaatagga acttcccgag cggccgcgtg
     3421 ttacaagctt gactccagcg taactggact gcaatcaact cactggctca ccttcacggg
     3481 tgggcctttc ttcggtagaa aatcaaagga tcttcttgag atcctttttt tctgcgcgta
     3541 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     3601 gagctaccaa ctctttttcc gaggtaactg gcttcagcag agcgcagata ccaaatactg
     3661 ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat
     3721 acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta
     3781 ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg
     3841 gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc
     3901 gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa
     3961 gcggcagggt cggaacagga gagcgcacga gggagcttcc aggggtaaac gcctggtatc
     4021 tttatagtcc tgtcgggttt cgccacctct gacttgagca tcgatttttg tgatgctcgt
     4081 caggggggcg gagcctatgg aaaaacgcca gcaacgcaga aaggcccacc cgaaggtgag
     4141 ccaggtgatt acatttgggc cctcatcaga ggttttcacc gtcatcaccg aaacgcgcga
     4201 ggcagctgcg gtaaagctca tcagcgtggt cgtgaagcga ttcacagatg tctgcctgtt
     4261 catccgcgtc cagctcgttg agtttctcca gaagcgttaa tgtctggctt ctgataaagc
     4321 gggccatgtt aagggcggtt ttttcctgtt tggtcactta ccaatgctta atcagtgagg
     4381 cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt
     4441 agataactac gatgcgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag
     4501 acccacgctc accggctcca gatttatcag caataaacca gccagccggg agggccgagc
     4561 gcagaagtga tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag
     4621 ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca
     4681 tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa
     4741 ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga
     4801 tcgttgccag aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata
     4861 attctcttac tgtcatgcca tccgtgagat gcttttctgt gactggtgag tactcaacca
     4921 agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg
     4981 ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg
     5041 ggcgtaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg
     5101 cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag
     5161 gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcattt
     5221 tagcttcctt agctcctgaa aatctcgata actcaaaaaa tacgcccggt agtgatctta
     5281 tttcattatg gtgaaagttg gaacctctta cgtgccgatc aagtcaaaag cctccggtcg
     5341 gaggcttttg actttctgct atggaggtca ggtatgattt aaatggtcag ttaga
//
