LOCUS       sequence-295523-        7223 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 7223)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 7223)
  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-m4133991
FEATURES             Location/Qualifiers
     source          1..7223
                     /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..694
                     /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"
     polyA_signal    763..884
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(774..793)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     828..847
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      999..1045
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1075..1211
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1256..1327
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(1553..2141)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1633..1652)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2641..3850
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     3687..3706
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             3854..4882
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     polyA_signal    4909..4957
                     /note="synthetic polyadenylation signal"
     CDS             join(5223..5291,5292..6083)
                     /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(5441..5460)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      6096..6232
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    6377..6448
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(6453..6499)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    6517..6565
                     /note="synthetic polyadenylation signal"
     enhancer        6646..6949
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6951..7153
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     7104..7124
                     /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 cagtcgaaga ggcgatggaa aaagaatttt atcgcggtga gtgcagccaa
      661 cagattcaaa aagatatcct cgtctggtgc cctctaagga tccgcaggcc tctgctagct
      721 tgactgactg agatacagcg taccttcagc tcacagacat gataagatac attgatgagt
      781 ttggacaaac cacaactaga atgcagtgaa aaaaatgctt tatttgtgaa atttgtgatg
      841 ctattgcttt atttgtaacc attataagct gcaataaaca agttaacaac aacaattgca
      901 ttcattttat gtttcaggtt cagggggagg tgtgggaggt tttttaaagc aagtaaaacc
      961 tctacaaatg tggtattggc ccatctctat cggtatcgta gcataacccc ttggggcctc
     1021 taaacgggtc ttgaggggtt ttttgtgccc ctcgggccgg attgctatct accggcattg
     1081 gcgcagaaaa aaatgcctga tgcgacgctg cgcgtcttat actcccacat atgccagatt
     1141 cagcaacgga tacggcttcc ccaacttgcc cacttccata cgtgtcctcc ttaccagaaa
     1201 tttatcctta aggtcgtcag ctatcctgca ggcgatctct cgatttcgat caagacattc
     1261 ctttaatggt cttttctgga caccactagg ggtcagaagt agttcatcaa actttcttcc
     1321 ctccctaatc tcattggtta ccttgggcta tcgaaactta attaaccagt caagtcagct
     1381 acttggcgag atcgacttgt ctgggtttcg actacgctca gaattgcgtc agtcaagttc
     1441 gatctggtcc ttgctattgc acccgttctc cgattacgag tttcatttaa atcatgtgag
     1501 caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata
     1561 ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc
     1621 cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg
     1681 ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc
     1741 tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg
     1801 gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc
     1861 ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga
     1921 ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg
     1981 gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa
     2041 aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg
     2101 tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt
     2161 ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat
     2221 tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct
     2281 aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta
     2341 tctcagcgat ctgtctattt cgttcatcca tagttgcatt taaatttccg aactctccaa
     2401 ggccctcgtc ggaaaatctt caaacctttc gtccgatcca tcttgcaggc tacctctcga
     2461 acgaactatc gcaagtctct tggccggcct tgcgccttgg ctattgcttg gcagcgccta
     2521 tcgccaggta ttactccaat cccgaatatc cgagatcggg atcacccgag agaagttcaa
     2581 cctacatcct caatcccgat ctatccgaga tccgaggaat atcgaaatcg gggcgcgcct
     2641 ggcctccgcg ccgggttttg gcgcctcccg cgggcgcccc cctcgtcacg gcgagcgctg
     2701 ccacgtcaga cgaagggcgc aggagcgtcc tgatccttcc gcccggacgc tcaggacagc
     2761 ggcccgctgc tcataagact cggccttaga accccagtat cagcagaagg acattttagg
     2821 acgggacttg ggtgactcta gggcactggt tttctttcca gagagcggaa caggcgagga
     2881 aaagtagtcc cttctcggcg attctgcgga gggatctccg tggggcggtg aacgccgatg
     2941 attatataag gacgcgccgg gtgtggcaca gctagttccg tcgcagccgg gatttgggtc
     3001 gcggttcttg tttgtggatc gctgtgatcg tcacttggtg agtagcgggc tgctgggctg
     3061 gccggggctt tcgtggccgc cgggccgctc ggtgggacgg aagcgtgtgg agagaccgcc
     3121 aagggctgta gtctgggtcc gcgagcaagg ttgccctgaa ctgggggttg gggggagcgc
     3181 agcaaaatgg cggctgttcc cgagtcttga atggaagacg cttgtgaggc gggctgtgag
     3241 gtcgttgaaa caaggtgggg ggcatggtgg gcggcaagaa cccaaggtct tgagcccttc
     3301 gctaatgcgg gaaagctctt attcgggtga gatgggctgg gcaccatctg gggaccctga
     3361 cgtgaagttt gtcactgact ggagaactcg gtttgtcgtc tgttgcgggg gcggcagtta
     3421 tggcggtgcc gttgggcagt gcacccgtac ctttgggagc gcgcgccctc gtcgtgtcgt
     3481 gacgtcaccc gttctgttgg cttataatgc agggtggggc cacctgccgg taggtgtgcg
     3541 gtaggctttt ctccgtcgca ggacgcaggg ttcgggccta gggtaggctc tcctgaatcg
     3601 acaggcgccg gacctctggt gaggggaggg ataagtgagg cgtcagtttc tttggtcggt
     3661 tttatgtacc tatcttctta agtagctgaa gctccggttt tgaactatgc gctcggggtt
     3721 ggcgagtgtg ttttgtgaag ttttttaggc accttttgaa atgtaatcat ttgggtcaat
     3781 atgtaatttt cagtgttaga cttgtaaatt gtccgctaaa ttctggccgt ttttggcttt
     3841 tttgttagac aacatgggta aaaagcctga actcaccgcg acgtctgtcg agaagtttct
     3901 gatcgaaaag ttcgacagcg tctccgacct gatgcagctc tcggagggcg aagaatctcg
     3961 tgctttcagc ttcgatgtag gagggcgtgg atatgtcctg cgggtaaata gctgcgccga
     4021 tggtttctac aaagatcgtt atgtttatcg gcactttgca tccgccgcgc tcccgattcc
     4081 ggaagtgctt gacattgggg agttcagcga gagcctgacc tattgcatct cccgccgtgc
     4141 acagggtgtc acgttgcaag acctgcctga aaccgaactg cccgctgttc tgcagccggt
     4201 cgcggaggca atggatgcca tcgctgccgc cgatcttagc cagacgagcg ggttcggccc
     4261 attcggaccg caaggaatcg gtcaatacac tacatggcgt gatttcatat gcgcgattgc
     4321 tgatccccat gtgtatcact ggcaaactgt gatggacgac accgtcagtg cgtccgtcgc
     4381 gcaggctctc gatgagctga tgctttgggc cgaggactgc cccgaagtcc ggcacctcgt
     4441 gcacgcggat ttcggctcca acaatgtcct gacggacaat ggccgcataa cagcggtcat
     4501 tgactggagc gaggcgatgt tcggggattc ccaatacgag gtcgccaaca tcttcttctg
     4561 gaggccgtgg ttggcttgta tggagcagca gacgcgctac ttcgagcgga ggcatccgga
     4621 gcttgcagga tcgccccggc tccgggcgta tatgctccgc attggtcttg accaactcta
     4681 tcagagcttg gttgacggca atttcgatga tgcagcttgg gcgcagggtc gatgcgacgc
     4741 aatcgtccga tccggagccg ggactgtcgg gcgtacacaa atcgcccgca gaagcgccgc
     4801 cgtctggacc gatggctgtg tagaagtact cgccgatagt ggaaaccgac gccccagcac
     4861 tcgtccgagg gcaaaggaat aagctagtat gtaagcctag tcttagataa taaaatcgct
     4921 atccatcgaa gatggatgtg tgttggtttt ttgtgtgtgt aacgctaggc gcgcctggtg
     4981 taccgagaac gatcctctca gtgcgagtct cgacgatcca tatcgttgct tggcagtcag
     5041 ccagtcggaa tccagcttgg gacccaggaa gtccaatcgt cagatattgt actcaagcct
     5101 ggtcacggca gcgtaccgat ctgtttaaac ctagatattg atagtctgat cggtcaacgt
     5161 ataatcgagt cctagctttt gcaaacatct atcaagagac aggatcagca ggaggctttc
     5221 gcatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc
     5281 ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg
     5341 cgcgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc
     5401 ccgaagaacg ctttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat
     5461 cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact
     5521 tggttgagta ttcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat
     5581 tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga
     5641 ttggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc
     5701 ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga
     5761 tgcctgtagc aatggcaaca accttgcgta aactattaac tggcgaacta cttactctag
     5821 cttcccggca acagttgata gactggatgg aggcggataa agttgcagga ccacttctgc
     5881 gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt
     5941 ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct
     6001 acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg
     6061 cctcactgat taagcattgg taaccgattc taggtgcatt ggcgcagaaa aaaatgcctg
     6121 atgcgacgct gcgcgtctta tactcccaca tatgccagat tcagcaacgg atacggcttc
     6181 cccaacttgc ccacttccat acgtgtcctc cttaccagaa atttatcctt aagatcgttt
     6241 aaactcgact ctggctctat cgaatctccg tcgtttcgag cttacgcgaa cagccgtggc
     6301 gctcatttgc tcgtcgggca tcgaatctcg tcagctatcg tcagcttacc tttttggcag
     6361 cgatcgcggc tcccgacatc ttggaccatt agctccacag gtatcttctt ccctctagtg
     6421 gtcataacag cagcttcagc tacctctcaa ttcaaaaaac ccctcaagac ccgtttagag
     6481 gccccaaggg gttatgctat caatcgttgc gttacacaca caaaaaacca acacacatcc
     6541 atcttcgatg gatagcgatt ttattatcta actgctgatc gagtgtagcc agatctagta
     6601 atcaattacg gggtcattag ttcatagccc atatatggag ttccgcgtta cataacttac
     6661 ggtaaatggc ccgcctggct gaccgcccaa cgacccccgc ccattgacgt caataatgac
     6721 gtatgttccc atagtaacgc caatagggac tttccattga cgtcaatggg tggagtattt
     6781 acggtaaact gcccacttgg cagtacatca agtgtatcat atgccaagta cgccccctat
     6841 tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc cagtacatga ccttatggga
     6901 ctttcctact tggcagtaca tctacgtatt agtcatcgct attaccatgc tgatgcggtt
     6961 ttggcagtac atcaatgggc gtggatagcg gtttgactca cggggatttc caagtctcca
     7021 ccccattgac gtcaatggga gtttgttttg gcaccaaaat caacgggact ttccaaaatg
     7081 tcgtaacaac tccgccccat tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta
     7141 tataagcaga gctggtttag tgaaccgtca gatcagatct ttgtcgatcc taccatccac
     7201 tcgacacacc cgccagcggc cgc
//
