LOCUS       sequence-295622-        7414 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression plasmid for SARS-CoV-2 3C-like proteinase.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7414)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 7414)
  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-m4133282
FEATURES             Location/Qualifiers
     source          1..7414
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             920..940
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             1289..1306
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     polyA_signal    1383..1504
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1394..1413)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1448..1467
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      1619..1665
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1695..1831
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1876..1947
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(2173..2761)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2253..2272)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        3261..4470
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     4307..4326
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             4474..5073
                     /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(4474..4493)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4970..4990
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    5100..5148
                     /note="synthetic polyadenylation signal"
     CDS             join(5414..5482,5483..6274)
                     /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(5632..5651)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      6287..6423
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    6568..6639
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(6644..6690)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    6708..6756
                     /note="synthetic polyadenylation signal"
     enhancer        6837..7140
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        7142..7344
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     7295..7315
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      join(7413..7414,1..8)
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
ORIGIN
        1 caccatgggt tttagaaaaa tggcattccc atctggtaaa gttgagggtt gtatggtaca
       61 agtaacttgt ggtacaacta cacttaacgg tctttggctt gatgacgtag tttactgtcc
      121 aagacatgtg atctgcacct ctgaagacat gcttaaccct aattatgaag atttactcat
      181 tcgtaagtct aatcataatt tcttggtaca ggctggtaat gttcaactca gggttattgg
      241 acattctatg caaaattgtg tacttaagct taaggttgat acagccaatc ctaagacacc
      301 taagtataag tttgttcgca ttcaaccagg acagactttt tcagtgttag cttgttacaa
      361 tggttcacca tctggtgttt accaatgtgc tatgaggccc aatttcacta ttaagggttc
      421 attccttaat ggttcatgtg gtagtgttgg ttttaacata gattatgact gtgtctcttt
      481 ttgttacatg caccatatgg aattaccaac tggagttcat gctggcacag acttagaagg
      541 taacttttat ggaccttttg ttgacaggca aacagcacaa gcagctggta cggacacaac
      601 tattacagtt aatgttttag cttggttgta cgctgctgtt ataaatggag acaggtggtt
      661 tctcaatcga tttaccacaa ctcttaatga ctttaacctt gtggctatga agtacaatta
      721 tgaacctcta acacaagacc atgttgacat actaggacct ctttctgctc aaactggaat
      781 tgccgtttta gatatgtgtg cttcattaaa agaattactg caaaatggta tgaatggacg
      841 taccatattg ggtagtgctt tattagaaga tgaatttaca ccttttgatg ttgttagaca
      901 atgctcaggt gttactttcg aaaaccttta ctttcagtcg atgagtgaca aaatcattca
      961 tctaaccgat gactcctttg atacagacgt gttgaaggcc gatggtgcga tactggtaga
     1021 tttctgggct gaatggtgtg gcccgtgcaa gatgatcgca cccattctgg acgagatcgc
     1081 cgacgagtat caaggaaaat taactgtcgc taagcttaat atagatcaga acccagggac
     1141 ggcacctaaa tacggcatca ggggaatacc taccctcctg ctctttaaga atggggaagt
     1201 tgccgcgaca aaagtgggag ccctgtctaa gggccagcta aaagagttct tggacgctaa
     1261 cctggcaggt tcaggctcgg ggcacatgca ccatcaccat caccatagca gcgggctcgt
     1321 gccacgcggc taagctagct tgactgactg agatacagcg taccttcagc tcacagacat
     1381 gataagatac attgatgagt ttggacaaac cacaactaga atgcagtgaa aaaaatgctt
     1441 tatttgtgaa atttgtgatg ctattgcttt atttgtaacc attataagct gcaataaaca
     1501 agttaacaac aacaattgca ttcattttat gtttcaggtt cagggggagg tgtgggaggt
     1561 tttttaaagc aagtaaaacc tctacaaatg tggtattggc ccatctctat cggtatcgta
     1621 gcataacccc ttggggcctc taaacgggtc ttgaggggtt ttttgtgccc ctcgggccgg
     1681 attgctatct accggcattg gcgcagaaaa aaatgcctga tgcgacgctg cgcgtcttat
     1741 actcccacat atgccagatt cagcaacgga tacggcttcc ccaacttgcc cacttccata
     1801 cgtgtcctcc ttaccagaaa tttatcctta aggtcgtcag ctatcctgca ggcgatctct
     1861 cgatttcgat caagacattc ctttaatggt cttttctgga caccactagg ggtcagaagt
     1921 agttcatcaa actttcttcc ctccctaatc tcattggtta ccttgggcta tcgaaactta
     1981 attaaccagt caagtcagct acttggcgag atcgacttgt ctgggtttcg actacgctca
     2041 gaattgcgtc agtcaagttc gatctggtcc ttgctattgc acccgttctc cgattacgag
     2101 tttcatttaa atcatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
     2161 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
     2221 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
     2281 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     2341 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
     2401 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     2461 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     2521 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     2581 gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct
     2641 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     2701 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca
     2761 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
     2821 agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa
     2881 atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg
     2941 cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcatt
     3001 taaatttccg aactctccaa ggccctcgtc ggaaaatctt caaacctttc gtccgatcca
     3061 tcttgcaggc tacctctcga acgaactatc gcaagtctct tggccggcct tgcgccttgg
     3121 ctattgcttg gcagcgccta tcgccaggta ttactccaat cccgaatatc cgagatcggg
     3181 atcacccgag agaagttcaa cctacatcct caatcccgat ctatccgaga tccgaggaat
     3241 atcgaaatcg gggcgcgcct ggcctccgcg ccgggttttg gcgcctcccg cgggcgcccc
     3301 cctcgtcacg gcgagcgctg ccacgtcaga cgaagggcgc aggagcgtcc tgatccttcc
     3361 gcccggacgc tcaggacagc ggcccgctgc tcataagact cggccttaga accccagtat
     3421 cagcagaagg acattttagg acgggacttg ggtgactcta gggcactggt tttctttcca
     3481 gagagcggaa caggcgagga aaagtagtcc cttctcggcg attctgcgga gggatctccg
     3541 tggggcggtg aacgccgatg attatataag gacgcgccgg gtgtggcaca gctagttccg
     3601 tcgcagccgg gatttgggtc gcggttcttg tttgtggatc gctgtgatcg tcacttggtg
     3661 agtagcgggc tgctgggctg gccggggctt tcgtggccgc cgggccgctc ggtgggacgg
     3721 aagcgtgtgg agagaccgcc aagggctgta gtctgggtcc gcgagcaagg ttgccctgaa
     3781 ctgggggttg gggggagcgc agcaaaatgg cggctgttcc cgagtcttga atggaagacg
     3841 cttgtgaggc gggctgtgag gtcgttgaaa caaggtgggg ggcatggtgg gcggcaagaa
     3901 cccaaggtct tgagcccttc gctaatgcgg gaaagctctt attcgggtga gatgggctgg
     3961 gcaccatctg gggaccctga cgtgaagttt gtcactgact ggagaactcg gtttgtcgtc
     4021 tgttgcgggg gcggcagtta tggcggtgcc gttgggcagt gcacccgtac ctttgggagc
     4081 gcgcgccctc gtcgtgtcgt gacgtcaccc gttctgttgg cttataatgc agggtggggc
     4141 cacctgccgg taggtgtgcg gtaggctttt ctccgtcgca ggacgcaggg ttcgggccta
     4201 gggtaggctc tcctgaatcg acaggcgccg gacctctggt gaggggaggg ataagtgagg
     4261 cgtcagtttc tttggtcggt tttatgtacc tatcttctta agtagctgaa gctccggttt
     4321 tgaactatgc gctcggggtt ggcgagtgtg ttttgtgaag ttttttaggc accttttgaa
     4381 atgtaatcat ttgggtcaat atgtaatttt cagtgttaga cttgtaaatt gtccgctaaa
     4441 ttctggccgt ttttggcttt tttgttagac aacatgaccg agtacaagcc cacggtgcgc
     4501 ctcgccaccc gcgacgacgt ccccagggcc gtacgcaccc tcgccgccgc gttcgccgac
     4561 taccccgcca cgcgccacac cgtcgatccg gaccgccaca tcgagcgggt caccgagctg
     4621 caagaactct tcctcacgcg cgtcgggctc gacatcggca aggtgtgggt cgcggacgac
     4681 ggcgccgcgg tggcggtctg gaccacgccg gagagcgtcg aagcgggggc ggtgttcgcc
     4741 gagatcggcc cgcgcatggc cgagttgagc ggttcccggc tggccgcgca gcaacagatg
     4801 gagggcctcc tggcgccgca ccggcccaag gagcccgcgt ggttcctggc caccgtcggc
     4861 gtctcgcccg accaccaggg caagggtctg ggcagcgccg tcgtgctccc cggagtggag
     4921 gccgccgagc gcgccggggt gcccgccttc ctggagacct ccgcgccccg caacctcccc
     4981 ttctacgagc ggctcggctt caccgtcacc gccgacgtcg aggtgcccga aggaccgcgc
     5041 acctggtgca tgacccgcaa gcccggtgcc taggctagta tgtaagccta gtcttagata
     5101 ataaaatcgc tatccatcga agatggatgt gtgttggttt tttgtgtgtg taacgctagg
     5161 cgcgcctggt gtaccgagaa cgatcctctc agtgcgagtc tcgacgatcc atatcgttgc
     5221 ttggcagtca gccagtcgga atccagcttg ggacccagga agtccaatcg tcagatattg
     5281 tactcaagcc tggtcacggc agcgtaccga tctgtttaaa cctagatatt gatagtctga
     5341 tcggtcaacg tataatcgag tcctagcttt tgcaaacatc tatcaagaga caggatcagc
     5401 aggaggcttt cgcatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc
     5461 attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga
     5521 tcagttgggt gcgcgagtgg gttacatcga actggatctc aacagcggta agatccttga
     5581 gagttttcgc cccgaagaac gctttccaat gatgagcact tttaaagttc tgctatgtgg
     5641 cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc
     5701 tcagaatgac ttggttgagt attcaccagt cacagaaaag catcttacgg atggcatgac
     5761 agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact
     5821 tctgacaacg attggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca
     5881 tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg
     5941 tgacaccacg atgcctgtag caatggcaac aaccttgcgt aaactattaa ctggcgaact
     6001 acttactcta gcttcccggc aacagttgat agactggatg gaggcggata aagttgcagg
     6061 accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg
     6121 tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat
     6181 cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc
     6241 tgagataggt gcctcactga ttaagcattg gtaaccgatt ctaggtgcat tggcgcagaa
     6301 aaaaatgcct gatgcgacgc tgcgcgtctt atactcccac atatgccaga ttcagcaacg
     6361 gatacggctt ccccaacttg cccacttcca tacgtgtcct ccttaccaga aatttatcct
     6421 taagatcgtt taaactcgac tctggctcta tcgaatctcc gtcgtttcga gcttacgcga
     6481 acagccgtgg cgctcatttg ctcgtcgggc atcgaatctc gtcagctatc gtcagcttac
     6541 ctttttggca gcgatcgcgg ctcccgacat cttggaccat tagctccaca ggtatcttct
     6601 tccctctagt ggtcataaca gcagcttcag ctacctctca attcaaaaaa cccctcaaga
     6661 cccgtttaga ggccccaagg ggttatgcta tcaatcgttg cgttacacac acaaaaaacc
     6721 aacacacatc catcttcgat ggatagcgat tttattatct aactgctgat cgagtgtagc
     6781 cagatctagt aatcaattac ggggtcatta gttcatagcc catatatgga gttccgcgtt
     6841 acataactta cggtaaatgg cccgcctggc tgaccgccca acgacccccg cccattgacg
     6901 tcaataatga cgtatgttcc catagtaacg ccaataggga ctttccattg acgtcaatgg
     6961 gtggagtatt tacggtaaac tgcccacttg gcagtacatc aagtgtatca tatgccaagt
     7021 acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc ccagtacatg
     7081 accttatggg actttcctac ttggcagtac atctacgtat tagtcatcgc tattaccatg
     7141 ctgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt
     7201 ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac
     7261 tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg
     7321 tgggaggtct atataagcag agctggttta gtgaaccgtc agatcagatc tttgtcgatc
     7381 ctaccatcca ctcgacacac ccgccagcgg ccgc
//
