LOCUS       sequence-295717-        7700 bp    DNA     circular SYN 15-DEC-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 7700)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 7700)
  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-m4134284
FEATURES             Location/Qualifiers
     source          1..7700
                     /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             650..727
                     /codon_start=1
                     /product="calmodulin-binding peptide"
                     /label=CBP
                     /note="derived from skeletal muscle myosin light chain 
                     kinase; binds calmodulin with nanomolar affinity in the 
                     presence of calcium"
                     /translation="KRRWKKNFIAVSAANRFKKISSSGAL"
     CDS             755..775
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     polyA_signal    1240..1361
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1251..1270)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1305..1324
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      1476..1522
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1552..1688
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1733..1804
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(2030..2618)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2110..2129)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        3118..4327
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     4164..4183
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             4331..5359
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     polyA_signal    5386..5434
                     /note="synthetic polyadenylation signal"
     CDS             join(5700..5768,5769..6560)
                     /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(5918..5937)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      6573..6709
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    6854..6925
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(6930..6976)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    6994..7042
                     /note="synthetic polyadenylation signal"
     enhancer        7123..7426
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        7428..7630
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     7581..7601
                     /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 cagtcgggac ggcgaatccc ggggctgata aatccctgga aaaggagatg
      661 gaagaagaac tttattgcag tgtctgcggc taaccggttc aaaaagatca gttcaagcgg
      721 cgccctagat tacgacattc ctacgaccgc ctccgagaat ctctatttcc agggtgaatt
      781 tggcttggca caacatgacg aggccgtcga taacaagttt aataaagaac agcagaacgc
      841 tttctatgag atccttcact taccaaatct gaacgaagag caacgcaacg ctttcattca
      901 gagcctgaaa gacgatccct cgcagtctgc aaatctcctg gccgaagcca agaagctgaa
      961 tgacgcacag gctccaaaag ttgataacaa gtttaacaag gagcagcaga atgctttcta
     1021 cgagatattg caccttccta atctgaatga ggaacaaaga aatgcattta tccagtccct
     1081 aaaagacgat ccatcacaga gtgccaacct gcttgcggaa gccaaaaagc tcaatgatgc
     1141 ccaggccccg aaagtagacg cgaaccatca ataaggatcc gcaggcctct gctagcttga
     1201 ctgactgaga tacagcgtac cttcagctca cagacatgat aagatacatt gatgagtttg
     1261 gacaaaccac aactagaatg cagtgaaaaa aatgctttat ttgtgaaatt tgtgatgcta
     1321 ttgctttatt tgtaaccatt ataagctgca ataaacaagt taacaacaac aattgcattc
     1381 attttatgtt tcaggttcag ggggaggtgt gggaggtttt ttaaagcaag taaaacctct
     1441 acaaatgtgg tattggccca tctctatcgg tatcgtagca taaccccttg gggcctctaa
     1501 acgggtcttg aggggttttt tgtgcccctc gggccggatt gctatctacc ggcattggcg
     1561 cagaaaaaaa tgcctgatgc gacgctgcgc gtcttatact cccacatatg ccagattcag
     1621 caacggatac ggcttcccca acttgcccac ttccatacgt gtcctcctta ccagaaattt
     1681 atccttaagg tcgtcagcta tcctgcaggc gatctctcga tttcgatcaa gacattcctt
     1741 taatggtctt ttctggacac cactaggggt cagaagtagt tcatcaaact ttcttccctc
     1801 cctaatctca ttggttacct tgggctatcg aaacttaatt aaccagtcaa gtcagctact
     1861 tggcgagatc gacttgtctg ggtttcgact acgctcagaa ttgcgtcagt caagttcgat
     1921 ctggtccttg ctattgcacc cgttctccga ttacgagttt catttaaatc atgtgagcaa
     1981 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
     2041 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
     2101 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
     2161 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
     2221 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
     2281 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
     2341 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
     2401 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
     2461 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
     2521 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
     2581 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
     2641 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
     2701 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
     2761 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
     2821 cagcgatctg tctatttcgt tcatccatag ttgcatttaa atttccgaac tctccaaggc
     2881 cctcgtcgga aaatcttcaa acctttcgtc cgatccatct tgcaggctac ctctcgaacg
     2941 aactatcgca agtctcttgg ccggccttgc gccttggcta ttgcttggca gcgcctatcg
     3001 ccaggtatta ctccaatccc gaatatccga gatcgggatc acccgagaga agttcaacct
     3061 acatcctcaa tcccgatcta tccgagatcc gaggaatatc gaaatcgggg cgcgcctggc
     3121 ctccgcgccg ggttttggcg cctcccgcgg gcgcccccct cgtcacggcg agcgctgcca
     3181 cgtcagacga agggcgcagg agcgtcctga tccttccgcc cggacgctca ggacagcggc
     3241 ccgctgctca taagactcgg ccttagaacc ccagtatcag cagaaggaca ttttaggacg
     3301 ggacttgggt gactctaggg cactggtttt ctttccagag agcggaacag gcgaggaaaa
     3361 gtagtccctt ctcggcgatt ctgcggaggg atctccgtgg ggcggtgaac gccgatgatt
     3421 atataaggac gcgccgggtg tggcacagct agttccgtcg cagccgggat ttgggtcgcg
     3481 gttcttgttt gtggatcgct gtgatcgtca cttggtgagt agcgggctgc tgggctggcc
     3541 ggggctttcg tggccgccgg gccgctcggt gggacggaag cgtgtggaga gaccgccaag
     3601 ggctgtagtc tgggtccgcg agcaaggttg ccctgaactg ggggttgggg ggagcgcagc
     3661 aaaatggcgg ctgttcccga gtcttgaatg gaagacgctt gtgaggcggg ctgtgaggtc
     3721 gttgaaacaa ggtggggggc atggtgggcg gcaagaaccc aaggtcttga gcccttcgct
     3781 aatgcgggaa agctcttatt cgggtgagat gggctgggca ccatctgggg accctgacgt
     3841 gaagtttgtc actgactgga gaactcggtt tgtcgtctgt tgcgggggcg gcagttatgg
     3901 cggtgccgtt gggcagtgca cccgtacctt tgggagcgcg cgccctcgtc gtgtcgtgac
     3961 gtcacccgtt ctgttggctt ataatgcagg gtggggccac ctgccggtag gtgtgcggta
     4021 ggcttttctc cgtcgcagga cgcagggttc gggcctaggg taggctctcc tgaatcgaca
     4081 ggcgccggac ctctggtgag gggagggata agtgaggcgt cagtttcttt ggtcggtttt
     4141 atgtacctat cttcttaagt agctgaagct ccggttttga actatgcgct cggggttggc
     4201 gagtgtgttt tgtgaagttt tttaggcacc ttttgaaatg taatcatttg ggtcaatatg
     4261 taattttcag tgttagactt gtaaattgtc cgctaaattc tggccgtttt tggctttttt
     4321 gttagacaac atgggtaaaa agcctgaact caccgcgacg tctgtcgaga agtttctgat
     4381 cgaaaagttc gacagcgtct ccgacctgat gcagctctcg gagggcgaag aatctcgtgc
     4441 tttcagcttc gatgtaggag ggcgtggata tgtcctgcgg gtaaatagct gcgccgatgg
     4501 tttctacaaa gatcgttatg tttatcggca ctttgcatcc gccgcgctcc cgattccgga
     4561 agtgcttgac attggggagt tcagcgagag cctgacctat tgcatctccc gccgtgcaca
     4621 gggtgtcacg ttgcaagacc tgcctgaaac cgaactgccc gctgttctgc agccggtcgc
     4681 ggaggcaatg gatgccatcg ctgccgccga tcttagccag acgagcgggt tcggcccatt
     4741 cggaccgcaa ggaatcggtc aatacactac atggcgtgat ttcatatgcg cgattgctga
     4801 tccccatgtg tatcactggc aaactgtgat ggacgacacc gtcagtgcgt ccgtcgcgca
     4861 ggctctcgat gagctgatgc tttgggccga ggactgcccc gaagtccggc acctcgtgca
     4921 cgcggatttc ggctccaaca atgtcctgac ggacaatggc cgcataacag cggtcattga
     4981 ctggagcgag gcgatgttcg gggattccca atacgaggtc gccaacatct tcttctggag
     5041 gccgtggttg gcttgtatgg agcagcagac gcgctacttc gagcggaggc atccggagct
     5101 tgcaggatcg ccccggctcc gggcgtatat gctccgcatt ggtcttgacc aactctatca
     5161 gagcttggtt gacggcaatt tcgatgatgc agcttgggcg cagggtcgat gcgacgcaat
     5221 cgtccgatcc ggagccggga ctgtcgggcg tacacaaatc gcccgcagaa gcgccgccgt
     5281 ctggaccgat ggctgtgtag aagtactcgc cgatagtgga aaccgacgcc ccagcactcg
     5341 tccgagggca aaggaataag ctagtatgta agcctagtct tagataataa aatcgctatc
     5401 catcgaagat ggatgtgtgt tggttttttg tgtgtgtaac gctaggcgcg cctggtgtac
     5461 cgagaacgat cctctcagtg cgagtctcga cgatccatat cgttgcttgg cagtcagcca
     5521 gtcggaatcc agcttgggac ccaggaagtc caatcgtcag atattgtact caagcctggt
     5581 cacggcagcg taccgatctg tttaaaccta gatattgata gtctgatcgg tcaacgtata
     5641 atcgagtcct agcttttgca aacatctatc aagagacagg atcagcagga ggctttcgca
     5701 tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg
     5761 tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcgc
     5821 gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg
     5881 aagaacgctt tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc
     5941 gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg
     6001 ttgagtattc accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat
     6061 gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgattg
     6121 gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg
     6181 atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc
     6241 ctgtagcaat ggcaacaacc ttgcgtaaac tattaactgg cgaactactt actctagctt
     6301 cccggcaaca gttgatagac tggatggagg cggataaagt tgcaggacca cttctgcgct
     6361 cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc
     6421 gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca
     6481 cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct
     6541 cactgattaa gcattggtaa ccgattctag gtgcattggc gcagaaaaaa atgcctgatg
     6601 cgacgctgcg cgtcttatac tcccacatat gccagattca gcaacggata cggcttcccc
     6661 aacttgccca cttccatacg tgtcctcctt accagaaatt tatccttaag atcgtttaaa
     6721 ctcgactctg gctctatcga atctccgtcg tttcgagctt acgcgaacag ccgtggcgct
     6781 catttgctcg tcgggcatcg aatctcgtca gctatcgtca gcttaccttt ttggcagcga
     6841 tcgcggctcc cgacatcttg gaccattagc tccacaggta tcttcttccc tctagtggtc
     6901 ataacagcag cttcagctac ctctcaattc aaaaaacccc tcaagacccg tttagaggcc
     6961 ccaaggggtt atgctatcaa tcgttgcgtt acacacacaa aaaaccaaca cacatccatc
     7021 ttcgatggat agcgatttta ttatctaact gctgatcgag tgtagccaga tctagtaatc
     7081 aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat aacttacggt
     7141 aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta
     7201 tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg
     7261 gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga
     7321 cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt
     7381 tcctacttgg cagtacatct acgtattagt catcgctatt accatgctga tgcggttttg
     7441 gcagtacatc aatgggcgtg gatagcggtt tgactcacgg ggatttccaa gtctccaccc
     7501 cattgacgtc aatgggagtt tgttttggca ccaaaatcaa cgggactttc caaaatgtcg
     7561 taacaactcc gccccattga cgcaaatggg cggtaggcgt gtacggtggg aggtctatat
     7621 aagcagagct ggtttagtga accgtcagat cagatctttg tcgatcctac catccactcg
     7681 acacacccgc cagcggccgc
//
