LOCUS       sequence-295859-        7547 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 7547)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 7547)
  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-m4133507
FEATURES             Location/Qualifiers
     source          1..7547
                     /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             941..1018
                     /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    1087..1208
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1098..1117)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1152..1171
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      1323..1369
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1399..1535
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1580..1651
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(1877..2465)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1957..1976)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2965..4174
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     4011..4030
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             4178..5206
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     polyA_signal    5233..5281
                     /note="synthetic polyadenylation signal"
     CDS             join(5547..5615,5616..6407)
                     /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(5765..5784)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      6420..6556
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    6701..6772
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(6777..6823)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    6841..6889
                     /note="synthetic polyadenylation signal"
     enhancer        6970..7273
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        7275..7477
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     7428..7448
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      join(7546..7547,1..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 aagaggcgat ggaaaaagaa
      961 ttttatcgcg gtgagtgcag ccaacagatt caaaaagata tcctcgtctg gtgccctcta
     1021 aggatccgca ggcctctgct agcttgactg actgagatac agcgtacctt cagctcacag
     1081 acatgataag atacattgat gagtttggac aaaccacaac tagaatgcag tgaaaaaaat
     1141 gctttatttg tgaaatttgt gatgctattg ctttatttgt aaccattata agctgcaata
     1201 aacaagttaa caacaacaat tgcattcatt ttatgtttca ggttcagggg gaggtgtggg
     1261 aggtttttta aagcaagtaa aacctctaca aatgtggtat tggcccatct ctatcggtat
     1321 cgtagcataa ccccttgggg cctctaaacg ggtcttgagg ggttttttgt gcccctcggg
     1381 ccggattgct atctaccggc attggcgcag aaaaaaatgc ctgatgcgac gctgcgcgtc
     1441 ttatactccc acatatgcca gattcagcaa cggatacggc ttccccaact tgcccacttc
     1501 catacgtgtc ctccttacca gaaatttatc cttaaggtcg tcagctatcc tgcaggcgat
     1561 ctctcgattt cgatcaagac attcctttaa tggtcttttc tggacaccac taggggtcag
     1621 aagtagttca tcaaactttc ttccctccct aatctcattg gttaccttgg gctatcgaaa
     1681 cttaattaac cagtcaagtc agctacttgg cgagatcgac ttgtctgggt ttcgactacg
     1741 ctcagaattg cgtcagtcaa gttcgatctg gtccttgcta ttgcacccgt tctccgatta
     1801 cgagtttcat ttaaatcatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
     1861 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
     1921 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
     1981 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
     2041 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
     2101 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
     2161 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
     2221 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
     2281 tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc
     2341 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
     2401 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
     2461 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
     2521 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt
     2581 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc
     2641 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
     2701 catttaaatt tccgaactct ccaaggccct cgtcggaaaa tcttcaaacc tttcgtccga
     2761 tccatcttgc aggctacctc tcgaacgaac tatcgcaagt ctcttggccg gccttgcgcc
     2821 ttggctattg cttggcagcg cctatcgcca ggtattactc caatcccgaa tatccgagat
     2881 cgggatcacc cgagagaagt tcaacctaca tcctcaatcc cgatctatcc gagatccgag
     2941 gaatatcgaa atcggggcgc gcctggcctc cgcgccgggt tttggcgcct cccgcgggcg
     3001 cccccctcgt cacggcgagc gctgccacgt cagacgaagg gcgcaggagc gtcctgatcc
     3061 ttccgcccgg acgctcagga cagcggcccg ctgctcataa gactcggcct tagaacccca
     3121 gtatcagcag aaggacattt taggacggga cttgggtgac tctagggcac tggttttctt
     3181 tccagagagc ggaacaggcg aggaaaagta gtcccttctc ggcgattctg cggagggatc
     3241 tccgtggggc ggtgaacgcc gatgattata taaggacgcg ccgggtgtgg cacagctagt
     3301 tccgtcgcag ccgggatttg ggtcgcggtt cttgtttgtg gatcgctgtg atcgtcactt
     3361 ggtgagtagc gggctgctgg gctggccggg gctttcgtgg ccgccgggcc gctcggtggg
     3421 acggaagcgt gtggagagac cgccaagggc tgtagtctgg gtccgcgagc aaggttgccc
     3481 tgaactgggg gttgggggga gcgcagcaaa atggcggctg ttcccgagtc ttgaatggaa
     3541 gacgcttgtg aggcgggctg tgaggtcgtt gaaacaaggt ggggggcatg gtgggcggca
     3601 agaacccaag gtcttgagcc cttcgctaat gcgggaaagc tcttattcgg gtgagatggg
     3661 ctgggcacca tctggggacc ctgacgtgaa gtttgtcact gactggagaa ctcggtttgt
     3721 cgtctgttgc gggggcggca gttatggcgg tgccgttggg cagtgcaccc gtacctttgg
     3781 gagcgcgcgc cctcgtcgtg tcgtgacgtc acccgttctg ttggcttata atgcagggtg
     3841 gggccacctg ccggtaggtg tgcggtaggc ttttctccgt cgcaggacgc agggttcggg
     3901 cctagggtag gctctcctga atcgacaggc gccggacctc tggtgagggg agggataagt
     3961 gaggcgtcag tttctttggt cggttttatg tacctatctt cttaagtagc tgaagctccg
     4021 gttttgaact atgcgctcgg ggttggcgag tgtgttttgt gaagtttttt aggcaccttt
     4081 tgaaatgtaa tcatttgggt caatatgtaa ttttcagtgt tagacttgta aattgtccgc
     4141 taaattctgg ccgtttttgg cttttttgtt agacaacatg ggtaaaaagc ctgaactcac
     4201 cgcgacgtct gtcgagaagt ttctgatcga aaagttcgac agcgtctccg acctgatgca
     4261 gctctcggag ggcgaagaat ctcgtgcttt cagcttcgat gtaggagggc gtggatatgt
     4321 cctgcgggta aatagctgcg ccgatggttt ctacaaagat cgttatgttt atcggcactt
     4381 tgcatccgcc gcgctcccga ttccggaagt gcttgacatt ggggagttca gcgagagcct
     4441 gacctattgc atctcccgcc gtgcacaggg tgtcacgttg caagacctgc ctgaaaccga
     4501 actgcccgct gttctgcagc cggtcgcgga ggcaatggat gccatcgctg ccgccgatct
     4561 tagccagacg agcgggttcg gcccattcgg accgcaagga atcggtcaat acactacatg
     4621 gcgtgatttc atatgcgcga ttgctgatcc ccatgtgtat cactggcaaa ctgtgatgga
     4681 cgacaccgtc agtgcgtccg tcgcgcaggc tctcgatgag ctgatgcttt gggccgagga
     4741 ctgccccgaa gtccggcacc tcgtgcacgc ggatttcggc tccaacaatg tcctgacgga
     4801 caatggccgc ataacagcgg tcattgactg gagcgaggcg atgttcgggg attcccaata
     4861 cgaggtcgcc aacatcttct tctggaggcc gtggttggct tgtatggagc agcagacgcg
     4921 ctacttcgag cggaggcatc cggagcttgc aggatcgccc cggctccggg cgtatatgct
     4981 ccgcattggt cttgaccaac tctatcagag cttggttgac ggcaatttcg atgatgcagc
     5041 ttgggcgcag ggtcgatgcg acgcaatcgt ccgatccgga gccgggactg tcgggcgtac
     5101 acaaatcgcc cgcagaagcg ccgccgtctg gaccgatggc tgtgtagaag tactcgccga
     5161 tagtggaaac cgacgcccca gcactcgtcc gagggcaaag gaataagcta gtatgtaagc
     5221 ctagtcttag ataataaaat cgctatccat cgaagatgga tgtgtgttgg ttttttgtgt
     5281 gtgtaacgct aggcgcgcct ggtgtaccga gaacgatcct ctcagtgcga gtctcgacga
     5341 tccatatcgt tgcttggcag tcagccagtc ggaatccagc ttgggaccca ggaagtccaa
     5401 tcgtcagata ttgtactcaa gcctggtcac ggcagcgtac cgatctgttt aaacctagat
     5461 attgatagtc tgatcggtca acgtataatc gagtcctagc ttttgcaaac atctatcaag
     5521 agacaggatc agcaggaggc tttcgcatga gtattcaaca tttccgtgtc gcccttattc
     5581 ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa
     5641 aagatgctga agatcagttg ggtgcgcgag tgggttacat cgaactggat ctcaacagcg
     5701 gtaagatcct tgagagtttt cgccccgaag aacgctttcc aatgatgagc acttttaaag
     5761 ttctgctatg tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc
     5821 gcatacacta ttctcagaat gacttggttg agtattcacc agtcacagaa aagcatctta
     5881 cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg
     5941 cggccaactt acttctgaca acgattggag gaccgaagga gctaaccgct tttttgcaca
     6001 acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac
     6061 caaacgacga gcgtgacacc acgatgcctg tagcaatggc aacaaccttg cgtaaactat
     6121 taactggcga actacttact ctagcttccc ggcaacagtt gatagactgg atggaggcgg
     6181 ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata
     6241 aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta
     6301 agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa
     6361 atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaaccg attctaggtg
     6421 cattggcgca gaaaaaaatg cctgatgcga cgctgcgcgt cttatactcc cacatatgcc
     6481 agattcagca acggatacgg cttccccaac ttgcccactt ccatacgtgt cctccttacc
     6541 agaaatttat ccttaagatc gtttaaactc gactctggct ctatcgaatc tccgtcgttt
     6601 cgagcttacg cgaacagccg tggcgctcat ttgctcgtcg ggcatcgaat ctcgtcagct
     6661 atcgtcagct tacctttttg gcagcgatcg cggctcccga catcttggac cattagctcc
     6721 acaggtatct tcttccctct agtggtcata acagcagctt cagctacctc tcaattcaaa
     6781 aaacccctca agacccgttt agaggcccca aggggttatg ctatcaatcg ttgcgttaca
     6841 cacacaaaaa accaacacac atccatcttc gatggatagc gattttatta tctaactgct
     6901 gatcgagtgt agccagatct agtaatcaat tacggggtca ttagttcata gcccatatat
     6961 ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc
     7021 ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag ggactttcca
     7081 ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac atcaagtgta
     7141 tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg cctggcatta
     7201 tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg tattagtcat
     7261 cgctattacc atgctgatgc ggttttggca gtacatcaat gggcgtggat agcggtttga
     7321 ctcacgggga tttccaagtc tccaccccat tgacgtcaat gggagtttgt tttggcacca
     7381 aaatcaacgg gactttccaa aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg
     7441 taggcgtgta cggtgggagg tctatataag cagagctggt ttagtgaacc gtcagatcag
     7501 atctttgtcg atcctaccat ccactcgaca cacccgccag cggccgc
//
