LOCUS       sequence-299994-        6334 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Yeast expression plasmid for SARS-CoV-2 3'-to-5' exonuclease.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6334)
  TITLE     SARS-CoV-2 protein Yeast expression constructs, tagged, for protein 
            expression and/or purification
REFERENCE   2  (bases 1 to 6334)
  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-m4046334
FEATURES             Location/Qualifiers
     source          1..6334
                     /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             2009..2026
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             2027..2044
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2089..2107)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     CDS             2458..3261
                     /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      3344..4326
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     terminator      4404..4433
                     /label=T3Te terminator
                     /note="phage T3 early transcription terminator"
     rep_origin      4455..5042
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     terminator      complement(5054..5081)
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     CDS             complement(5105..5296)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(5297..6088,6089..6157))
                     /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     5920..5939
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6158..6260)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     terminator      complement(6261..6292)
                     /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 atggaaaatg taacaggact ctttaaagat tgtagtaagg taatcactgg
      481 gttacatcct acacaggcac ctacacacct cagtgttgac actaaattca aaactgaagg
      541 tttatgtgtt gacatacctg gcatacctaa ggacatgacc tatagaagac tcatctctat
      601 gatgggtttt aaaatgaatt atcaagttaa tggttaccct aacatgttta tcacccgcga
      661 agaagctata agacatgtac gtgcatggat tggcttcgat gtcgaggggt gtcatgctac
      721 tagagaagct gttggtacca atttaccttt acagctaggt ttttctacag gtgttaacct
      781 agttgctgta cctacaggtt atgttgatac acctaataat acagattttt ccagagttag
      841 tgctaaacca ccgcctggag atcaatttaa acacctcata ccacttatgt acaaaggact
      901 tccttggaat gtagtgcgta taaagattgt acaaatgtta agtgacacac ttaaaaatct
      961 ctctgacaga gtcgtatttg tcttatgggc acatggcttt gagttgacat ctatgaagta
     1021 ttttgtgaaa ataggacctg agcgcacctg ttgtctatgt gatagacgtg ccacatgctt
     1081 ttccactgct tcagacactt atgcctgttg gcatcattct attggatttg attacgtcta
     1141 taatccgttt atgattgatg ttcaacaatg gggttttaca ggtaacctac aaagcaacca
     1201 tgatctgtat tgtcaagtcc atggtaatgc acatgtagct agttgtgatg caatcatgac
     1261 taggtgtcta gctgtccacg agtgctttgt taagcgtgtt gactggacta ttgaatatcc
     1321 tataattggt gatgaactga agattaatgc ggcttgtaga aaggttcaac acatggttgt
     1381 taaagctgca ttattagcag acaaattccc agttcttcac gacattggta accctaaagc
     1441 tattaagtgt gtacctcaag ctgatgtaga atggaagttc tatgatgcac agccttgtag
     1501 tgacaaagct tataaaatag aagaattatt ctattcttat gccacacatt ctgacaaatt
     1561 cacagatggt gtatgcctat tttggaattg caatgtcgat agatatcctg ctaattccat
     1621 tgtttgtaga tttgacacta gagtgctatc taaccttaac ttgcctggtt gtgatggtgg
     1681 cagtttgtat gtaaataaac atgcattcca cacaccagct tttgataaaa gtgcttttgt
     1741 taatttaaaa caattaccat ttttctatta ctctgacagt ccatgtgagt ctcatggaaa
     1801 acaagtagtg tcagatatag attatgtacc actaaagtct gctacgtgta taacacgttg
     1861 caatttaggt ggtgctgtct gtagacatca tgctaatgag tacagattgt atctcgatgc
     1921 ttataacatg atgatctcag ctggctttag cttgtgggtt tacaaacaat ttgatactta
     1981 taacctctgg aacactttta caagacttct cgtgccgaga ggttctcacc atcatcacca
     2041 tcattaaaca ggcccctttt cctttgtcga tatcatgtaa ttagttatgt cacgcttaca
     2101 ttcacgccct cctcccacat ccgctctaac cgaaaaggaa ggagttagac aacctgaagt
     2161 ctaggtccct atttattttt tttaatagtt atgttagtat taagaacgtt atttatattt
     2221 caaatttttc ttttttttct gtacaaagcg gccgccgaac cactagtgca ctgcagtaca
     2281 cggtttcctt gaaatttttt tgattcggta atctccgaac agaaggaaga acgaaggaag
     2341 gagcacagac ttagattggt atatatacgc atatgtagtg ttgaagaaac atgaaattgc
     2401 ccagtattct taacccaact gcacagaaca aaaacgtgca ggaaacgaag ataaatcatg
     2461 tcgaaagcta catataagga acgtgctgct actcatccta gtcctgttgc tgccaagcta
     2521 tttaatatca tgcacgaaaa gcaaacaaac ttgtgtgctt cattggatgt tcgtaccacc
     2581 aaggaattac tggagttagt tgaagcatta ggtcccaaaa tttgtttact aaaaacacat
     2641 gtggatatct tgactgattt ttccatggag ggcacagtta agccgctaaa ggcattatcc
     2701 gccaagtaca attttttact cttcgaggac agaaaatttg ctgacattgg taatacagtc
     2761 aaattgcagt actctgcggg tgtatacaga atagcagaat gggcagacat tacgaatgca
     2821 cacggtgtgg tgggcccagg tattgttagc ggtttgaagc aggcggcaga agaagtaaca
     2881 aaggaaccta gaggcctttt gatgttagca gaattgtcat gcaagggctc cctatctact
     2941 ggagaatata ctaagggtac tgttgacatt gcgaagagcg acaaagattt tgttatcggc
     3001 tttattgctc aaagagacat gggtggaaga gatgaaggtt acgattggtt gattatgaca
     3061 cccggtgtgg gtttagatga caagggagat gcattgggtc aacagtatag aaccgtggat
     3121 gatgtggttt ctacaggatc tgacattatt attgttggaa gaggactatt tgcaaaggga
     3181 agggatgcta aggtagaggg tgaacgttac agaaaagcag gctgggaagc atatttgaga
     3241 agatgcggcc agcaaaacta aaaaactgta ttataagtaa atgcatgtat actaaactca
     3301 caaattagag cttcaattta attatatcag ttattacccg agtaacgaag catctgtgct
     3361 tcattttgta gaacaaaaat gcaacgcgag agcgctaatt tttcaaacaa agaatctgag
     3421 ctgcattttt acagaacaga aatgcaacgc gaaagcgcta ttttaccaac gaagaatctg
     3481 tgcttcattt ttgtaaaaca aaaatgcaac gcgagagcgc taatttttca aacaaagaat
     3541 ctgagctgca tttttacaga acagaaatgc aacgcgagag cgctatttta ccaacaaaga
     3601 atctatactt cttttttgtt ctacaaaaat gcatcccgag agcgctattt ttctaacaaa
     3661 gcatcttaga ttactttttt tctcctttgt gcgctctata atgcagtctc ttgataactt
     3721 tttgcactgt aggtccgtta aggttagaag aaggctactt tggtgtctat tttctcttcc
     3781 ataaaaaaag cctgactcca cttcccgcgt ttactgatta ctagcgaagc tgcgggtgca
     3841 ttttttcaag ataaaggcat ccccgattat attctatacc gatgtggatt gcgcatactt
     3901 tgtgaacaga aagtgatagc gttgatgatt cttcattggt cagaaaatta tgaacggttt
     3961 cttctatttt gtctctatat actacgtata ggaaatgttt acattttcgt attgttttcg
     4021 attcactcta tgaatagttc ttactacaat ttttttgtct aaagagtaat actagagata
     4081 aacataaaaa atgtagaggt cgagtttaga tgcaagttca aggagcgaaa ggtggatggg
     4141 taggttatat agggatatag cacagagata tatagcaaag agatactttt gagcaatgtt
     4201 tgtggaagcg gtattcgcaa tattttagta gctcgttaca gtccggtgcg tttttggttt
     4261 tttgaaagtg cgtcatcaga gcgcttttgg ttttcaaaag cgctctgaag ttcctatact
     4321 ttctagctag agaataggaa cttcccgagc ggccgcgtgt tacaagcttg actccagcgt
     4381 aactggactg caatcaactc actggctcac cttcacgggt gggcctttct tcggtagaaa
     4441 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
     4501 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
     4561 aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt
     4621 taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt
     4681 taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat
     4741 agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct
     4801 tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca
     4861 cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag
     4921 agcgcacgag ggagcttcca ggggtaaacg cctggtatct ttatagtcct gtcgggtttc
     4981 gccacctctg acttgagcat cgatttttgt gatgctcgtc aggggggcgg agcctatgga
     5041 aaaacgccag caacgcagaa aggcccaccc gaaggtgagc caggtgatta catttgggcc
     5101 ctcatcagag gttttcaccg tcatcaccga aacgcgcgag gcagctgcgg taaagctcat
     5161 cagcgtggtc gtgaagcgat tcacagatgt ctgcctgttc atccgcgtcc agctcgttga
     5221 gtttctccag aagcgttaat gtctggcttc tgataaagcg ggccatgtta agggcggttt
     5281 tttcctgttt ggtcacttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     5341 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atgcgggagg
     5401 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
     5461 atttatcagc aataaaccag ccagccggga gggccgagcg cagaagtgat cctgcaactt
     5521 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
     5581 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt
     5641 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
     5701 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgccaga agtaagttgg
     5761 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
     5821 ccgtgagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
     5881 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca
     5941 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgtaaactc tcaaggatct
     6001 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
     6061 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
     6121 agggaataag ggcgacacgg aaatgttgaa tactcatttt agcttcctta gctcctgaaa
     6181 atctcgataa ctcaaaaaat acgcccggta gtgatcttat ttcattatgg tgaaagttgg
     6241 aacctcttac gtgccgatca agtcaaaagc ctccggtcgg aggcttttga ctttctgcta
     6301 tggaggtcag gtatgattta aatggtcagt taga
//
