LOCUS       sequence-295092-        6994 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 6994)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 6994)
  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-m4134399
FEATURES             Location/Qualifiers
     source          1..6994
                     /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             893..910
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     polyA_signal    963..1084
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(974..993)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1028..1047
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      1199..1245
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1275..1411
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1456..1527
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(1753..2341)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1833..1852)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2841..4050
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     3887..3906
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             4054..4653
                     /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(4054..4073)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4550..4570
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4680..4728
                     /note="synthetic polyadenylation signal"
     CDS             join(4994..5062,5063..5854)
                     /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(5212..5231)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      5867..6003
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    6148..6219
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(6224..6270)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    6288..6336
                     /note="synthetic polyadenylation signal"
     enhancer        6417..6720
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6722..6924
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     6875..6895
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      join(6993..6994,1..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 cgcaccatca
      901 ccatcaccat taagctagct tgactgactg agatacagcg taccttcagc tcacagacat
      961 gataagatac attgatgagt ttggacaaac cacaactaga atgcagtgaa aaaaatgctt
     1021 tatttgtgaa atttgtgatg ctattgcttt atttgtaacc attataagct gcaataaaca
     1081 agttaacaac aacaattgca ttcattttat gtttcaggtt cagggggagg tgtgggaggt
     1141 tttttaaagc aagtaaaacc tctacaaatg tggtattggc ccatctctat cggtatcgta
     1201 gcataacccc ttggggcctc taaacgggtc ttgaggggtt ttttgtgccc ctcgggccgg
     1261 attgctatct accggcattg gcgcagaaaa aaatgcctga tgcgacgctg cgcgtcttat
     1321 actcccacat atgccagatt cagcaacgga tacggcttcc ccaacttgcc cacttccata
     1381 cgtgtcctcc ttaccagaaa tttatcctta aggtcgtcag ctatcctgca ggcgatctct
     1441 cgatttcgat caagacattc ctttaatggt cttttctgga caccactagg ggtcagaagt
     1501 agttcatcaa actttcttcc ctccctaatc tcattggtta ccttgggcta tcgaaactta
     1561 attaaccagt caagtcagct acttggcgag atcgacttgt ctgggtttcg actacgctca
     1621 gaattgcgtc agtcaagttc gatctggtcc ttgctattgc acccgttctc cgattacgag
     1681 tttcatttaa atcatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
     1741 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
     1801 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
     1861 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     1921 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
     1981 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     2041 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     2101 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     2161 gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct
     2221 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     2281 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca
     2341 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
     2401 agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa
     2461 atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg
     2521 cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcatt
     2581 taaatttccg aactctccaa ggccctcgtc ggaaaatctt caaacctttc gtccgatcca
     2641 tcttgcaggc tacctctcga acgaactatc gcaagtctct tggccggcct tgcgccttgg
     2701 ctattgcttg gcagcgccta tcgccaggta ttactccaat cccgaatatc cgagatcggg
     2761 atcacccgag agaagttcaa cctacatcct caatcccgat ctatccgaga tccgaggaat
     2821 atcgaaatcg gggcgcgcct ggcctccgcg ccgggttttg gcgcctcccg cgggcgcccc
     2881 cctcgtcacg gcgagcgctg ccacgtcaga cgaagggcgc aggagcgtcc tgatccttcc
     2941 gcccggacgc tcaggacagc ggcccgctgc tcataagact cggccttaga accccagtat
     3001 cagcagaagg acattttagg acgggacttg ggtgactcta gggcactggt tttctttcca
     3061 gagagcggaa caggcgagga aaagtagtcc cttctcggcg attctgcgga gggatctccg
     3121 tggggcggtg aacgccgatg attatataag gacgcgccgg gtgtggcaca gctagttccg
     3181 tcgcagccgg gatttgggtc gcggttcttg tttgtggatc gctgtgatcg tcacttggtg
     3241 agtagcgggc tgctgggctg gccggggctt tcgtggccgc cgggccgctc ggtgggacgg
     3301 aagcgtgtgg agagaccgcc aagggctgta gtctgggtcc gcgagcaagg ttgccctgaa
     3361 ctgggggttg gggggagcgc agcaaaatgg cggctgttcc cgagtcttga atggaagacg
     3421 cttgtgaggc gggctgtgag gtcgttgaaa caaggtgggg ggcatggtgg gcggcaagaa
     3481 cccaaggtct tgagcccttc gctaatgcgg gaaagctctt attcgggtga gatgggctgg
     3541 gcaccatctg gggaccctga cgtgaagttt gtcactgact ggagaactcg gtttgtcgtc
     3601 tgttgcgggg gcggcagtta tggcggtgcc gttgggcagt gcacccgtac ctttgggagc
     3661 gcgcgccctc gtcgtgtcgt gacgtcaccc gttctgttgg cttataatgc agggtggggc
     3721 cacctgccgg taggtgtgcg gtaggctttt ctccgtcgca ggacgcaggg ttcgggccta
     3781 gggtaggctc tcctgaatcg acaggcgccg gacctctggt gaggggaggg ataagtgagg
     3841 cgtcagtttc tttggtcggt tttatgtacc tatcttctta agtagctgaa gctccggttt
     3901 tgaactatgc gctcggggtt ggcgagtgtg ttttgtgaag ttttttaggc accttttgaa
     3961 atgtaatcat ttgggtcaat atgtaatttt cagtgttaga cttgtaaatt gtccgctaaa
     4021 ttctggccgt ttttggcttt tttgttagac aacatgaccg agtacaagcc cacggtgcgc
     4081 ctcgccaccc gcgacgacgt ccccagggcc gtacgcaccc tcgccgccgc gttcgccgac
     4141 taccccgcca cgcgccacac cgtcgatccg gaccgccaca tcgagcgggt caccgagctg
     4201 caagaactct tcctcacgcg cgtcgggctc gacatcggca aggtgtgggt cgcggacgac
     4261 ggcgccgcgg tggcggtctg gaccacgccg gagagcgtcg aagcgggggc ggtgttcgcc
     4321 gagatcggcc cgcgcatggc cgagttgagc ggttcccggc tggccgcgca gcaacagatg
     4381 gagggcctcc tggcgccgca ccggcccaag gagcccgcgt ggttcctggc caccgtcggc
     4441 gtctcgcccg accaccaggg caagggtctg ggcagcgccg tcgtgctccc cggagtggag
     4501 gccgccgagc gcgccggggt gcccgccttc ctggagacct ccgcgccccg caacctcccc
     4561 ttctacgagc ggctcggctt caccgtcacc gccgacgtcg aggtgcccga aggaccgcgc
     4621 acctggtgca tgacccgcaa gcccggtgcc taggctagta tgtaagccta gtcttagata
     4681 ataaaatcgc tatccatcga agatggatgt gtgttggttt tttgtgtgtg taacgctagg
     4741 cgcgcctggt gtaccgagaa cgatcctctc agtgcgagtc tcgacgatcc atatcgttgc
     4801 ttggcagtca gccagtcgga atccagcttg ggacccagga agtccaatcg tcagatattg
     4861 tactcaagcc tggtcacggc agcgtaccga tctgtttaaa cctagatatt gatagtctga
     4921 tcggtcaacg tataatcgag tcctagcttt tgcaaacatc tatcaagaga caggatcagc
     4981 aggaggcttt cgcatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc
     5041 attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga
     5101 tcagttgggt gcgcgagtgg gttacatcga actggatctc aacagcggta agatccttga
     5161 gagttttcgc cccgaagaac gctttccaat gatgagcact tttaaagttc tgctatgtgg
     5221 cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc
     5281 tcagaatgac ttggttgagt attcaccagt cacagaaaag catcttacgg atggcatgac
     5341 agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact
     5401 tctgacaacg attggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca
     5461 tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg
     5521 tgacaccacg atgcctgtag caatggcaac aaccttgcgt aaactattaa ctggcgaact
     5581 acttactcta gcttcccggc aacagttgat agactggatg gaggcggata aagttgcagg
     5641 accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg
     5701 tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat
     5761 cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc
     5821 tgagataggt gcctcactga ttaagcattg gtaaccgatt ctaggtgcat tggcgcagaa
     5881 aaaaatgcct gatgcgacgc tgcgcgtctt atactcccac atatgccaga ttcagcaacg
     5941 gatacggctt ccccaacttg cccacttcca tacgtgtcct ccttaccaga aatttatcct
     6001 taagatcgtt taaactcgac tctggctcta tcgaatctcc gtcgtttcga gcttacgcga
     6061 acagccgtgg cgctcatttg ctcgtcgggc atcgaatctc gtcagctatc gtcagcttac
     6121 ctttttggca gcgatcgcgg ctcccgacat cttggaccat tagctccaca ggtatcttct
     6181 tccctctagt ggtcataaca gcagcttcag ctacctctca attcaaaaaa cccctcaaga
     6241 cccgtttaga ggccccaagg ggttatgcta tcaatcgttg cgttacacac acaaaaaacc
     6301 aacacacatc catcttcgat ggatagcgat tttattatct aactgctgat cgagtgtagc
     6361 cagatctagt aatcaattac ggggtcatta gttcatagcc catatatgga gttccgcgtt
     6421 acataactta cggtaaatgg cccgcctggc tgaccgccca acgacccccg cccattgacg
     6481 tcaataatga cgtatgttcc catagtaacg ccaataggga ctttccattg acgtcaatgg
     6541 gtggagtatt tacggtaaac tgcccacttg gcagtacatc aagtgtatca tatgccaagt
     6601 acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc ccagtacatg
     6661 accttatggg actttcctac ttggcagtac atctacgtat tagtcatcgc tattaccatg
     6721 ctgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt
     6781 ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac
     6841 tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg
     6901 tgggaggtct atataagcag agctggttta gtgaaccgtc agatcagatc tttgtcgatc
     6961 ctaccatcca ctcgacacac ccgccagcgg ccgc
//
