LOCUS       sequence-295888-        7187 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Mammalian expression plasmid for SARS-CoV-2 nsp8.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7187)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 7187)
  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-m4133751
FEATURES             Location/Qualifiers
     source          1..7187
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             596..616
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             617..658
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     polyA_signal    727..848
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(738..757)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     792..811
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      963..1009
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1039..1175
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1220..1291
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(1517..2105)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1597..1616)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2605..3814
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     3651..3670
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             3818..4846
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     polyA_signal    4873..4921
                     /note="synthetic polyadenylation signal"
     CDS             join(5187..5255,5256..6047)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(5405..5424)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      6060..6196
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    6341..6412
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(6417..6463)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    6481..6529
                     /note="synthetic polyadenylation signal"
     enhancer        6610..6913
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6915..7117
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     7068..7088
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
ORIGIN
        1 caccatgata gcctcagagt ttagttccct tccatcatat gcagcttttg ctactgctca
       61 agaagcttat gagcaggctg ttgctaatgg tgattctgaa gttgttctta aaaagttgaa
      121 gaagtctttg aatgtggcta aatctgaatt tgaccgtgat gcagccatgc aacgtaagtt
      181 ggaaaagatg gctgatcaag ctatgaccca aatgtataaa caggctagat ctgaggacaa
      241 gagggcaaaa gttactagtg ctatgcagac aatgcttttc actatgctta gaaagttgga
      301 taatgatgca ctcaacaaca ttatcaacaa tgcaagagat ggttgtgttc ccttgaacat
      361 aatacctctt acaacagcag ccaaactaat ggttgtcata ccagactata acacatataa
      421 aaatacgtgt gatggtacaa catttactta tgcatcagca ttgtgggaaa tccaacaggt
      481 tgtagatgca gatagtaaaa ttgttcaact tagtgaaatt agtatggaca attcacctaa
      541 tttagcatgg cctcttattg taacagcttt aagggccaat tctgctgtca aattagaaaa
      601 cctttacttt cagtcgggga aacctatacc caatccactt ctcggcttag attctaccta
      661 aggatccgca ggcctctgct agcttgactg actgagatac agcgtacctt cagctcacag
      721 acatgataag atacattgat gagtttggac aaaccacaac tagaatgcag tgaaaaaaat
      781 gctttatttg tgaaatttgt gatgctattg ctttatttgt aaccattata agctgcaata
      841 aacaagttaa caacaacaat tgcattcatt ttatgtttca ggttcagggg gaggtgtggg
      901 aggtttttta aagcaagtaa aacctctaca aatgtggtat tggcccatct ctatcggtat
      961 cgtagcataa ccccttgggg cctctaaacg ggtcttgagg ggttttttgt gcccctcggg
     1021 ccggattgct atctaccggc attggcgcag aaaaaaatgc ctgatgcgac gctgcgcgtc
     1081 ttatactccc acatatgcca gattcagcaa cggatacggc ttccccaact tgcccacttc
     1141 catacgtgtc ctccttacca gaaatttatc cttaaggtcg tcagctatcc tgcaggcgat
     1201 ctctcgattt cgatcaagac attcctttaa tggtcttttc tggacaccac taggggtcag
     1261 aagtagttca tcaaactttc ttccctccct aatctcattg gttaccttgg gctatcgaaa
     1321 cttaattaac cagtcaagtc agctacttgg cgagatcgac ttgtctgggt ttcgactacg
     1381 ctcagaattg cgtcagtcaa gttcgatctg gtccttgcta ttgcacccgt tctccgatta
     1441 cgagtttcat ttaaatcatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
     1501 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
     1561 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
     1621 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
     1681 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
     1741 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
     1801 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
     1861 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
     1921 tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc
     1981 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
     2041 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
     2101 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
     2161 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt
     2221 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc
     2281 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
     2341 catttaaatt tccgaactct ccaaggccct cgtcggaaaa tcttcaaacc tttcgtccga
     2401 tccatcttgc aggctacctc tcgaacgaac tatcgcaagt ctcttggccg gccttgcgcc
     2461 ttggctattg cttggcagcg cctatcgcca ggtattactc caatcccgaa tatccgagat
     2521 cgggatcacc cgagagaagt tcaacctaca tcctcaatcc cgatctatcc gagatccgag
     2581 gaatatcgaa atcggggcgc gcctggcctc cgcgccgggt tttggcgcct cccgcgggcg
     2641 cccccctcgt cacggcgagc gctgccacgt cagacgaagg gcgcaggagc gtcctgatcc
     2701 ttccgcccgg acgctcagga cagcggcccg ctgctcataa gactcggcct tagaacccca
     2761 gtatcagcag aaggacattt taggacggga cttgggtgac tctagggcac tggttttctt
     2821 tccagagagc ggaacaggcg aggaaaagta gtcccttctc ggcgattctg cggagggatc
     2881 tccgtggggc ggtgaacgcc gatgattata taaggacgcg ccgggtgtgg cacagctagt
     2941 tccgtcgcag ccgggatttg ggtcgcggtt cttgtttgtg gatcgctgtg atcgtcactt
     3001 ggtgagtagc gggctgctgg gctggccggg gctttcgtgg ccgccgggcc gctcggtggg
     3061 acggaagcgt gtggagagac cgccaagggc tgtagtctgg gtccgcgagc aaggttgccc
     3121 tgaactgggg gttgggggga gcgcagcaaa atggcggctg ttcccgagtc ttgaatggaa
     3181 gacgcttgtg aggcgggctg tgaggtcgtt gaaacaaggt ggggggcatg gtgggcggca
     3241 agaacccaag gtcttgagcc cttcgctaat gcgggaaagc tcttattcgg gtgagatggg
     3301 ctgggcacca tctggggacc ctgacgtgaa gtttgtcact gactggagaa ctcggtttgt
     3361 cgtctgttgc gggggcggca gttatggcgg tgccgttggg cagtgcaccc gtacctttgg
     3421 gagcgcgcgc cctcgtcgtg tcgtgacgtc acccgttctg ttggcttata atgcagggtg
     3481 gggccacctg ccggtaggtg tgcggtaggc ttttctccgt cgcaggacgc agggttcggg
     3541 cctagggtag gctctcctga atcgacaggc gccggacctc tggtgagggg agggataagt
     3601 gaggcgtcag tttctttggt cggttttatg tacctatctt cttaagtagc tgaagctccg
     3661 gttttgaact atgcgctcgg ggttggcgag tgtgttttgt gaagtttttt aggcaccttt
     3721 tgaaatgtaa tcatttgggt caatatgtaa ttttcagtgt tagacttgta aattgtccgc
     3781 taaattctgg ccgtttttgg cttttttgtt agacaacatg ggtaaaaagc ctgaactcac
     3841 cgcgacgtct gtcgagaagt ttctgatcga aaagttcgac agcgtctccg acctgatgca
     3901 gctctcggag ggcgaagaat ctcgtgcttt cagcttcgat gtaggagggc gtggatatgt
     3961 cctgcgggta aatagctgcg ccgatggttt ctacaaagat cgttatgttt atcggcactt
     4021 tgcatccgcc gcgctcccga ttccggaagt gcttgacatt ggggagttca gcgagagcct
     4081 gacctattgc atctcccgcc gtgcacaggg tgtcacgttg caagacctgc ctgaaaccga
     4141 actgcccgct gttctgcagc cggtcgcgga ggcaatggat gccatcgctg ccgccgatct
     4201 tagccagacg agcgggttcg gcccattcgg accgcaagga atcggtcaat acactacatg
     4261 gcgtgatttc atatgcgcga ttgctgatcc ccatgtgtat cactggcaaa ctgtgatgga
     4321 cgacaccgtc agtgcgtccg tcgcgcaggc tctcgatgag ctgatgcttt gggccgagga
     4381 ctgccccgaa gtccggcacc tcgtgcacgc ggatttcggc tccaacaatg tcctgacgga
     4441 caatggccgc ataacagcgg tcattgactg gagcgaggcg atgttcgggg attcccaata
     4501 cgaggtcgcc aacatcttct tctggaggcc gtggttggct tgtatggagc agcagacgcg
     4561 ctacttcgag cggaggcatc cggagcttgc aggatcgccc cggctccggg cgtatatgct
     4621 ccgcattggt cttgaccaac tctatcagag cttggttgac ggcaatttcg atgatgcagc
     4681 ttgggcgcag ggtcgatgcg acgcaatcgt ccgatccgga gccgggactg tcgggcgtac
     4741 acaaatcgcc cgcagaagcg ccgccgtctg gaccgatggc tgtgtagaag tactcgccga
     4801 tagtggaaac cgacgcccca gcactcgtcc gagggcaaag gaataagcta gtatgtaagc
     4861 ctagtcttag ataataaaat cgctatccat cgaagatgga tgtgtgttgg ttttttgtgt
     4921 gtgtaacgct aggcgcgcct ggtgtaccga gaacgatcct ctcagtgcga gtctcgacga
     4981 tccatatcgt tgcttggcag tcagccagtc ggaatccagc ttgggaccca ggaagtccaa
     5041 tcgtcagata ttgtactcaa gcctggtcac ggcagcgtac cgatctgttt aaacctagat
     5101 attgatagtc tgatcggtca acgtataatc gagtcctagc ttttgcaaac atctatcaag
     5161 agacaggatc agcaggaggc tttcgcatga gtattcaaca tttccgtgtc gcccttattc
     5221 ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa
     5281 aagatgctga agatcagttg ggtgcgcgag tgggttacat cgaactggat ctcaacagcg
     5341 gtaagatcct tgagagtttt cgccccgaag aacgctttcc aatgatgagc acttttaaag
     5401 ttctgctatg tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc
     5461 gcatacacta ttctcagaat gacttggttg agtattcacc agtcacagaa aagcatctta
     5521 cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg
     5581 cggccaactt acttctgaca acgattggag gaccgaagga gctaaccgct tttttgcaca
     5641 acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac
     5701 caaacgacga gcgtgacacc acgatgcctg tagcaatggc aacaaccttg cgtaaactat
     5761 taactggcga actacttact ctagcttccc ggcaacagtt gatagactgg atggaggcgg
     5821 ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata
     5881 aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta
     5941 agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa
     6001 atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaaccg attctaggtg
     6061 cattggcgca gaaaaaaatg cctgatgcga cgctgcgcgt cttatactcc cacatatgcc
     6121 agattcagca acggatacgg cttccccaac ttgcccactt ccatacgtgt cctccttacc
     6181 agaaatttat ccttaagatc gtttaaactc gactctggct ctatcgaatc tccgtcgttt
     6241 cgagcttacg cgaacagccg tggcgctcat ttgctcgtcg ggcatcgaat ctcgtcagct
     6301 atcgtcagct tacctttttg gcagcgatcg cggctcccga catcttggac cattagctcc
     6361 acaggtatct tcttccctct agtggtcata acagcagctt cagctacctc tcaattcaaa
     6421 aaacccctca agacccgttt agaggcccca aggggttatg ctatcaatcg ttgcgttaca
     6481 cacacaaaaa accaacacac atccatcttc gatggatagc gattttatta tctaactgct
     6541 gatcgagtgt agccagatct agtaatcaat tacggggtca ttagttcata gcccatatat
     6601 ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc
     6661 ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag ggactttcca
     6721 ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac atcaagtgta
     6781 tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg cctggcatta
     6841 tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg tattagtcat
     6901 cgctattacc atgctgatgc ggttttggca gtacatcaat gggcgtggat agcggtttga
     6961 ctcacgggga tttccaagtc tccaccccat tgacgtcaat gggagtttgt tttggcacca
     7021 aaatcaacgg gactttccaa aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg
     7081 taggcgtgta cggtgggagg tctatataag cagagctggt ttagtgaacc gtcagatcag
     7141 atctttgtcg atcctaccat ccactcgaca cacccgccag cggccgc
//
