LOCUS       sequence-295697-        7364 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Mammalian expression plasmid for SARS-CoV-2 nsp10.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7364)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 7364)
  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-m4133904
FEATURES             Location/Qualifiers
     source          1..7364
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             425..445
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             794..811
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     polyA_signal    904..1025
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(915..934)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     969..988
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      1140..1186
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1216..1352
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1397..1468
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(1694..2282)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1774..1793)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2782..3991
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     3828..3847
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             3995..5023
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     polyA_signal    5050..5098
                     /note="synthetic polyadenylation signal"
     CDS             join(5364..5432,5433..6224)
                     /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(5582..5601)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      6237..6373
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    6518..6589
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(6594..6640)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    6658..6706
                     /note="synthetic polyadenylation signal"
     enhancer        6787..7090
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        7092..7294
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     7245..7265
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      join(7363..7364,1..8)
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
ORIGIN
        1 caccatggca gggaatgcga cagaggttcc tgccaactca acggtcttgt ctttttgtgc
       61 tttcgccgtg gacgctgcaa aagcctacaa ggattatcta gcctcgggtg gacagccaat
      121 cactaactgc gtaaagatgc tttgtaccca caccggcact ggccaagcaa ttacagtgac
      181 ccccgaagct aatatggacc aggagtcctt cggaggtgcg agctgctgcc tgtactgtcg
      241 ttgtcatata gatcacccga atcccaaagg gttttgcgac ctcaagggca aatatgtcca
      301 gatcccaacg acatgcgcca acgatcctgt gggattcact ctgaagaaca ccgtatgtac
      361 cgtttgcggg atgtggaagg ggtatggctg tagttgcgac caattacggg aaccgatgct
      421 gcaagaaaac ctttactttc agtcgatgag tgacaaaatc attcatctaa ccgatgactc
      481 ctttgataca gacgtgttga aggccgatgg tgcgatactg gtagatttct gggctgaatg
      541 gtgtggcccg tgcaagatga tcgcacccat tctggacgag atcgccgacg agtatcaagg
      601 aaaattaact gtcgctaagc ttaatataga tcagaaccca gggacggcac ctaaatacgg
      661 catcagggga atacctaccc tcctgctctt taagaatggg gaagttgccg cgacaaaagt
      721 gggagccctg tctaagggcc agctaaaaga gttcttggac gctaacctgg caggttcagg
      781 ctcggggcac atgcaccatc accatcacca tagcagcggg ctcgtgccac gcggctaagg
      841 atccgcaggc ctctgctagc ttgactgact gagatacagc gtaccttcag ctcacagaca
      901 tgataagata cattgatgag tttggacaaa ccacaactag aatgcagtga aaaaaatgct
      961 ttatttgtga aatttgtgat gctattgctt tatttgtaac cattataagc tgcaataaac
     1021 aagttaacaa caacaattgc attcatttta tgtttcaggt tcagggggag gtgtgggagg
     1081 ttttttaaag caagtaaaac ctctacaaat gtggtattgg cccatctcta tcggtatcgt
     1141 agcataaccc cttggggcct ctaaacgggt cttgaggggt tttttgtgcc cctcgggccg
     1201 gattgctatc taccggcatt ggcgcagaaa aaaatgcctg atgcgacgct gcgcgtctta
     1261 tactcccaca tatgccagat tcagcaacgg atacggcttc cccaacttgc ccacttccat
     1321 acgtgtcctc cttaccagaa atttatcctt aaggtcgtca gctatcctgc aggcgatctc
     1381 tcgatttcga tcaagacatt cctttaatgg tcttttctgg acaccactag gggtcagaag
     1441 tagttcatca aactttcttc cctccctaat ctcattggtt accttgggct atcgaaactt
     1501 aattaaccag tcaagtcagc tacttggcga gatcgacttg tctgggtttc gactacgctc
     1561 agaattgcgt cagtcaagtt cgatctggtc cttgctattg cacccgttct ccgattacga
     1621 gtttcattta aatcatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     1681 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     1741 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     1801 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     1861 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     1921 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     1981 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     2041 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     2101 tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc
     2161 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     2221 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     2281 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     2341 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     2401 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     2461 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcat
     2521 ttaaatttcc gaactctcca aggccctcgt cggaaaatct tcaaaccttt cgtccgatcc
     2581 atcttgcagg ctacctctcg aacgaactat cgcaagtctc ttggccggcc ttgcgccttg
     2641 gctattgctt ggcagcgcct atcgccaggt attactccaa tcccgaatat ccgagatcgg
     2701 gatcacccga gagaagttca acctacatcc tcaatcccga tctatccgag atccgaggaa
     2761 tatcgaaatc ggggcgcgcc tggcctccgc gccgggtttt ggcgcctccc gcgggcgccc
     2821 ccctcgtcac ggcgagcgct gccacgtcag acgaagggcg caggagcgtc ctgatccttc
     2881 cgcccggacg ctcaggacag cggcccgctg ctcataagac tcggccttag aaccccagta
     2941 tcagcagaag gacattttag gacgggactt gggtgactct agggcactgg ttttctttcc
     3001 agagagcgga acaggcgagg aaaagtagtc ccttctcggc gattctgcgg agggatctcc
     3061 gtggggcggt gaacgccgat gattatataa ggacgcgccg ggtgtggcac agctagttcc
     3121 gtcgcagccg ggatttgggt cgcggttctt gtttgtggat cgctgtgatc gtcacttggt
     3181 gagtagcggg ctgctgggct ggccggggct ttcgtggccg ccgggccgct cggtgggacg
     3241 gaagcgtgtg gagagaccgc caagggctgt agtctgggtc cgcgagcaag gttgccctga
     3301 actgggggtt ggggggagcg cagcaaaatg gcggctgttc ccgagtcttg aatggaagac
     3361 gcttgtgagg cgggctgtga ggtcgttgaa acaaggtggg gggcatggtg ggcggcaaga
     3421 acccaaggtc ttgagccctt cgctaatgcg ggaaagctct tattcgggtg agatgggctg
     3481 ggcaccatct ggggaccctg acgtgaagtt tgtcactgac tggagaactc ggtttgtcgt
     3541 ctgttgcggg ggcggcagtt atggcggtgc cgttgggcag tgcacccgta cctttgggag
     3601 cgcgcgccct cgtcgtgtcg tgacgtcacc cgttctgttg gcttataatg cagggtgggg
     3661 ccacctgccg gtaggtgtgc ggtaggcttt tctccgtcgc aggacgcagg gttcgggcct
     3721 agggtaggct ctcctgaatc gacaggcgcc ggacctctgg tgaggggagg gataagtgag
     3781 gcgtcagttt ctttggtcgg ttttatgtac ctatcttctt aagtagctga agctccggtt
     3841 ttgaactatg cgctcggggt tggcgagtgt gttttgtgaa gttttttagg caccttttga
     3901 aatgtaatca tttgggtcaa tatgtaattt tcagtgttag acttgtaaat tgtccgctaa
     3961 attctggccg tttttggctt ttttgttaga caacatgggt aaaaagcctg aactcaccgc
     4021 gacgtctgtc gagaagtttc tgatcgaaaa gttcgacagc gtctccgacc tgatgcagct
     4081 ctcggagggc gaagaatctc gtgctttcag cttcgatgta ggagggcgtg gatatgtcct
     4141 gcgggtaaat agctgcgccg atggtttcta caaagatcgt tatgtttatc ggcactttgc
     4201 atccgccgcg ctcccgattc cggaagtgct tgacattggg gagttcagcg agagcctgac
     4261 ctattgcatc tcccgccgtg cacagggtgt cacgttgcaa gacctgcctg aaaccgaact
     4321 gcccgctgtt ctgcagccgg tcgcggaggc aatggatgcc atcgctgccg ccgatcttag
     4381 ccagacgagc gggttcggcc cattcggacc gcaaggaatc ggtcaataca ctacatggcg
     4441 tgatttcata tgcgcgattg ctgatcccca tgtgtatcac tggcaaactg tgatggacga
     4501 caccgtcagt gcgtccgtcg cgcaggctct cgatgagctg atgctttggg ccgaggactg
     4561 ccccgaagtc cggcacctcg tgcacgcgga tttcggctcc aacaatgtcc tgacggacaa
     4621 tggccgcata acagcggtca ttgactggag cgaggcgatg ttcggggatt cccaatacga
     4681 ggtcgccaac atcttcttct ggaggccgtg gttggcttgt atggagcagc agacgcgcta
     4741 cttcgagcgg aggcatccgg agcttgcagg atcgccccgg ctccgggcgt atatgctccg
     4801 cattggtctt gaccaactct atcagagctt ggttgacggc aatttcgatg atgcagcttg
     4861 ggcgcagggt cgatgcgacg caatcgtccg atccggagcc gggactgtcg ggcgtacaca
     4921 aatcgcccgc agaagcgccg ccgtctggac cgatggctgt gtagaagtac tcgccgatag
     4981 tggaaaccga cgccccagca ctcgtccgag ggcaaaggaa taagctagta tgtaagccta
     5041 gtcttagata ataaaatcgc tatccatcga agatggatgt gtgttggttt tttgtgtgtg
     5101 taacgctagg cgcgcctggt gtaccgagaa cgatcctctc agtgcgagtc tcgacgatcc
     5161 atatcgttgc ttggcagtca gccagtcgga atccagcttg ggacccagga agtccaatcg
     5221 tcagatattg tactcaagcc tggtcacggc agcgtaccga tctgtttaaa cctagatatt
     5281 gatagtctga tcggtcaacg tataatcgag tcctagcttt tgcaaacatc tatcaagaga
     5341 caggatcagc aggaggcttt cgcatgagta ttcaacattt ccgtgtcgcc cttattccct
     5401 tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag
     5461 atgctgaaga tcagttgggt gcgcgagtgg gttacatcga actggatctc aacagcggta
     5521 agatccttga gagttttcgc cccgaagaac gctttccaat gatgagcact tttaaagttc
     5581 tgctatgtgg cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca
     5641 tacactattc tcagaatgac ttggttgagt attcaccagt cacagaaaag catcttacgg
     5701 atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg
     5761 ccaacttact tctgacaacg attggaggac cgaaggagct aaccgctttt ttgcacaaca
     5821 tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa
     5881 acgacgagcg tgacaccacg atgcctgtag caatggcaac aaccttgcgt aaactattaa
     5941 ctggcgaact acttactcta gcttcccggc aacagttgat agactggatg gaggcggata
     6001 aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat
     6061 ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc
     6121 cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata
     6181 gacagatcgc tgagataggt gcctcactga ttaagcattg gtaaccgatt ctaggtgcat
     6241 tggcgcagaa aaaaatgcct gatgcgacgc tgcgcgtctt atactcccac atatgccaga
     6301 ttcagcaacg gatacggctt ccccaacttg cccacttcca tacgtgtcct ccttaccaga
     6361 aatttatcct taagatcgtt taaactcgac tctggctcta tcgaatctcc gtcgtttcga
     6421 gcttacgcga acagccgtgg cgctcatttg ctcgtcgggc atcgaatctc gtcagctatc
     6481 gtcagcttac ctttttggca gcgatcgcgg ctcccgacat cttggaccat tagctccaca
     6541 ggtatcttct tccctctagt ggtcataaca gcagcttcag ctacctctca attcaaaaaa
     6601 cccctcaaga cccgtttaga ggccccaagg ggttatgcta tcaatcgttg cgttacacac
     6661 acaaaaaacc aacacacatc catcttcgat ggatagcgat tttattatct aactgctgat
     6721 cgagtgtagc cagatctagt aatcaattac ggggtcatta gttcatagcc catatatgga
     6781 gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca acgacccccg
     6841 cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga ctttccattg
     6901 acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc aagtgtatca
     6961 tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc
     7021 ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat tagtcatcgc
     7081 tattaccatg ctgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc
     7141 acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa
     7201 tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag
     7261 gcgtgtacgg tgggaggtct atataagcag agctggttta gtgaaccgtc agatcagatc
     7321 tttgtcgatc ctaccatcca ctcgacacac ccgccagcgg ccgc
//
