LOCUS       sequence-294999-        7531 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression plasmid for SARS-CoV-2 nsp6.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7531)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 7531)
  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-m4133192
FEATURES             Location/Qualifiers
     source          1..7531
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             872..892
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             926..1003
                     /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             1031..1051
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     polyA_signal    1500..1621
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1511..1530)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1565..1584
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      1736..1782
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1812..1948
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1993..2064
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(2290..2878)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2370..2389)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        3378..4587
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     4424..4443
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             4591..5190
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(4591..4610)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5087..5107
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    5217..5265
                     /note="synthetic polyadenylation signal"
     CDS             join(5531..5599,5600..6391)
                     /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(5749..5768)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      6404..6540
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    6685..6756
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(6761..6807)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    6825..6873
                     /note="synthetic polyadenylation signal"
     enhancer        6954..7257
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        7259..7461
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     7412..7432
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      join(7530..7531,1..8)
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
ORIGIN
        1 caccatggca gtgaaaagaa caatcaaggg tacacaccac tggttgttac tcacaatttt
       61 gacttcactt ttagttttag tccagagtac tcaatggtct ttgttctttt ttttgtatga
      121 aaatgccttt ttaccttttg ctatgggtat tattgctatg tctgcttttg caatgatgtt
      181 tgtcaaacat aagcatgcat ttctctgttt gtttttgtta ccttctcttg ccactgtagc
      241 ttattttaat atggtctata tgcctgctag ttgggtgatg cgtattatga catggttgga
      301 tatggttgat actagtttgt ctggttttaa gctaaaagac tgtgttatgt atgcatcagc
      361 tgtagtgtta ctaatcctta tgacagcaag aactgtgtat gatgatggtg ctaggagagt
      421 gtggacactt atgaatgtct tgacactcgt ttataaagtt tattatggta atgctttaga
      481 tcaagccatt tccatgtggg ctcttataat ctctgttact tctaactact caggtgtagt
      541 tacaactgtc atgtttttgg ccagaggtat tgtttttatg tgtgttgagt attgccctat
      601 tttcttcata actggtaata cacttcagtg tataatgcta gtttattgtt tcttaggcta
      661 tttttgtact tgttactttg gcctcttttg tttactcaac cgctacttta gactgactct
      721 tggtgtttat gattacttag tttctacaca ggagtttaga tatatgaatt cacagggact
      781 actcccaccc aagaatagca tagatgcctt caaactcaac attaaattgt tgggtgttgg
      841 tggcaaacct tgtatcaaag tagccactgt agaaaacctt tactttcagt cgggacggcg
      901 aatcccgggg ctgataaatc cctggaaaag gagatggaag aagaacttta ttgcagtgtc
      961 tgcggctaac cggttcaaaa agatcagttc aagcggcgcc ctagattacg acattcctac
     1021 gaccgcctcc gagaatctct atttccaggg tgaatttggc ttggcacaac atgacgaggc
     1081 cgtcgataac aagtttaata aagaacagca gaacgctttc tatgagatcc ttcacttacc
     1141 aaatctgaac gaagagcaac gcaacgcttt cattcagagc ctgaaagacg atccctcgca
     1201 gtctgcaaat ctcctggccg aagccaagaa gctgaatgac gcacaggctc caaaagttga
     1261 taacaagttt aacaaggagc agcagaatgc tttctacgag atattgcacc ttcctaatct
     1321 gaatgaggaa caaagaaatg catttatcca gtccctaaaa gacgatccat cacagagtgc
     1381 caacctgctt gcggaagcca aaaagctcaa tgatgcccag gccccgaaag tagacgcgaa
     1441 ccatcaataa gctagcttga ctgactgaga tacagcgtac cttcagctca cagacatgat
     1501 aagatacatt gatgagtttg gacaaaccac aactagaatg cagtgaaaaa aatgctttat
     1561 ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt ataagctgca ataaacaagt
     1621 taacaacaac aattgcattc attttatgtt tcaggttcag ggggaggtgt gggaggtttt
     1681 ttaaagcaag taaaacctct acaaatgtgg tattggccca tctctatcgg tatcgtagca
     1741 taaccccttg gggcctctaa acgggtcttg aggggttttt tgtgcccctc gggccggatt
     1801 gctatctacc ggcattggcg cagaaaaaaa tgcctgatgc gacgctgcgc gtcttatact
     1861 cccacatatg ccagattcag caacggatac ggcttcccca acttgcccac ttccatacgt
     1921 gtcctcctta ccagaaattt atccttaagg tcgtcagcta tcctgcaggc gatctctcga
     1981 tttcgatcaa gacattcctt taatggtctt ttctggacac cactaggggt cagaagtagt
     2041 tcatcaaact ttcttccctc cctaatctca ttggttacct tgggctatcg aaacttaatt
     2101 aaccagtcaa gtcagctact tggcgagatc gacttgtctg ggtttcgact acgctcagaa
     2161 ttgcgtcagt caagttcgat ctggtccttg ctattgcacc cgttctccga ttacgagttt
     2221 catttaaatc atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt
     2281 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag
     2341 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc
     2401 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc
     2461 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt
     2521 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt
     2581 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc
     2641 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa
     2701 gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa
     2761 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg
     2821 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga
     2881 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg
     2941 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg
     3001 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt
     3061 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcatttaa
     3121 atttccgaac tctccaaggc cctcgtcgga aaatcttcaa acctttcgtc cgatccatct
     3181 tgcaggctac ctctcgaacg aactatcgca agtctcttgg ccggccttgc gccttggcta
     3241 ttgcttggca gcgcctatcg ccaggtatta ctccaatccc gaatatccga gatcgggatc
     3301 acccgagaga agttcaacct acatcctcaa tcccgatcta tccgagatcc gaggaatatc
     3361 gaaatcgggg cgcgcctggc ctccgcgccg ggttttggcg cctcccgcgg gcgcccccct
     3421 cgtcacggcg agcgctgcca cgtcagacga agggcgcagg agcgtcctga tccttccgcc
     3481 cggacgctca ggacagcggc ccgctgctca taagactcgg ccttagaacc ccagtatcag
     3541 cagaaggaca ttttaggacg ggacttgggt gactctaggg cactggtttt ctttccagag
     3601 agcggaacag gcgaggaaaa gtagtccctt ctcggcgatt ctgcggaggg atctccgtgg
     3661 ggcggtgaac gccgatgatt atataaggac gcgccgggtg tggcacagct agttccgtcg
     3721 cagccgggat ttgggtcgcg gttcttgttt gtggatcgct gtgatcgtca cttggtgagt
     3781 agcgggctgc tgggctggcc ggggctttcg tggccgccgg gccgctcggt gggacggaag
     3841 cgtgtggaga gaccgccaag ggctgtagtc tgggtccgcg agcaaggttg ccctgaactg
     3901 ggggttgggg ggagcgcagc aaaatggcgg ctgttcccga gtcttgaatg gaagacgctt
     3961 gtgaggcggg ctgtgaggtc gttgaaacaa ggtggggggc atggtgggcg gcaagaaccc
     4021 aaggtcttga gcccttcgct aatgcgggaa agctcttatt cgggtgagat gggctgggca
     4081 ccatctgggg accctgacgt gaagtttgtc actgactgga gaactcggtt tgtcgtctgt
     4141 tgcgggggcg gcagttatgg cggtgccgtt gggcagtgca cccgtacctt tgggagcgcg
     4201 cgccctcgtc gtgtcgtgac gtcacccgtt ctgttggctt ataatgcagg gtggggccac
     4261 ctgccggtag gtgtgcggta ggcttttctc cgtcgcagga cgcagggttc gggcctaggg
     4321 taggctctcc tgaatcgaca ggcgccggac ctctggtgag gggagggata agtgaggcgt
     4381 cagtttcttt ggtcggtttt atgtacctat cttcttaagt agctgaagct ccggttttga
     4441 actatgcgct cggggttggc gagtgtgttt tgtgaagttt tttaggcacc ttttgaaatg
     4501 taatcatttg ggtcaatatg taattttcag tgttagactt gtaaattgtc cgctaaattc
     4561 tggccgtttt tggctttttt gttagacaac atgaccgagt acaagcccac ggtgcgcctc
     4621 gccacccgcg acgacgtccc cagggccgta cgcaccctcg ccgccgcgtt cgccgactac
     4681 cccgccacgc gccacaccgt cgatccggac cgccacatcg agcgggtcac cgagctgcaa
     4741 gaactcttcc tcacgcgcgt cgggctcgac atcggcaagg tgtgggtcgc ggacgacggc
     4801 gccgcggtgg cggtctggac cacgccggag agcgtcgaag cgggggcggt gttcgccgag
     4861 atcggcccgc gcatggccga gttgagcggt tcccggctgg ccgcgcagca acagatggag
     4921 ggcctcctgg cgccgcaccg gcccaaggag cccgcgtggt tcctggccac cgtcggcgtc
     4981 tcgcccgacc accagggcaa gggtctgggc agcgccgtcg tgctccccgg agtggaggcc
     5041 gccgagcgcg ccggggtgcc cgccttcctg gagacctccg cgccccgcaa cctccccttc
     5101 tacgagcggc tcggcttcac cgtcaccgcc gacgtcgagg tgcccgaagg accgcgcacc
     5161 tggtgcatga cccgcaagcc cggtgcctag gctagtatgt aagcctagtc ttagataata
     5221 aaatcgctat ccatcgaaga tggatgtgtg ttggtttttt gtgtgtgtaa cgctaggcgc
     5281 gcctggtgta ccgagaacga tcctctcagt gcgagtctcg acgatccata tcgttgcttg
     5341 gcagtcagcc agtcggaatc cagcttggga cccaggaagt ccaatcgtca gatattgtac
     5401 tcaagcctgg tcacggcagc gtaccgatct gtttaaacct agatattgat agtctgatcg
     5461 gtcaacgtat aatcgagtcc tagcttttgc aaacatctat caagagacag gatcagcagg
     5521 aggctttcgc atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt
     5581 ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca
     5641 gttgggtgcg cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag
     5701 ttttcgcccc gaagaacgct ttccaatgat gagcactttt aaagttctgc tatgtggcgc
     5761 ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac actattctca
     5821 gaatgacttg gttgagtatt caccagtcac agaaaagcat cttacggatg gcatgacagt
     5881 aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct
     5941 gacaacgatt ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt
     6001 aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga
     6061 caccacgatg cctgtagcaa tggcaacaac cttgcgtaaa ctattaactg gcgaactact
     6121 tactctagct tcccggcaac agttgataga ctggatggag gcggataaag ttgcaggacc
     6181 acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga
     6241 gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt
     6301 agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga
     6361 gataggtgcc tcactgatta agcattggta accgattcta ggtgcattgg cgcagaaaaa
     6421 aatgcctgat gcgacgctgc gcgtcttata ctcccacata tgccagattc agcaacggat
     6481 acggcttccc caacttgccc acttccatac gtgtcctcct taccagaaat ttatccttaa
     6541 gatcgtttaa actcgactct ggctctatcg aatctccgtc gtttcgagct tacgcgaaca
     6601 gccgtggcgc tcatttgctc gtcgggcatc gaatctcgtc agctatcgtc agcttacctt
     6661 tttggcagcg atcgcggctc ccgacatctt ggaccattag ctccacaggt atcttcttcc
     6721 ctctagtggt cataacagca gcttcagcta cctctcaatt caaaaaaccc ctcaagaccc
     6781 gtttagaggc cccaaggggt tatgctatca atcgttgcgt tacacacaca aaaaaccaac
     6841 acacatccat cttcgatgga tagcgatttt attatctaac tgctgatcga gtgtagccag
     6901 atctagtaat caattacggg gtcattagtt catagcccat atatggagtt ccgcgttaca
     6961 taacttacgg taaatggccc gcctggctga ccgcccaacg acccccgccc attgacgtca
     7021 ataatgacgt atgttcccat agtaacgcca atagggactt tccattgacg tcaatgggtg
     7081 gagtatttac ggtaaactgc ccacttggca gtacatcaag tgtatcatat gccaagtacg
     7141 ccccctattg acgtcaatga cggtaaatgg cccgcctggc attatgccca gtacatgacc
     7201 ttatgggact ttcctacttg gcagtacatc tacgtattag tcatcgctat taccatgctg
     7261 atgcggtttt ggcagtacat caatgggcgt ggatagcggt ttgactcacg gggatttcca
     7321 agtctccacc ccattgacgt caatgggagt ttgttttggc accaaaatca acgggacttt
     7381 ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg gcggtaggcg tgtacggtgg
     7441 gaggtctata taagcagagc tggtttagtg aaccgtcaga tcagatcttt gtcgatccta
     7501 ccatccactc gacacacccg ccagcggccg c
//
