LOCUS       sequence-295829-        7102 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 7102)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 7102)
  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-m4133922
FEATURES             Location/Qualifiers
     source          1..7102
                     /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    1071..1192
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1082..1101)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1136..1155
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      1307..1353
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1383..1519
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1564..1635
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(1861..2449)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1941..1960)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2949..4158
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     3995..4014
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             4162..4761
                     /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(4162..4181)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4658..4678
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4788..4836
                     /note="synthetic polyadenylation signal"
     CDS             join(5102..5170,5171..5962)
                     /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(5320..5339)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      5975..6111
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    6256..6327
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(6332..6378)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    6396..6444
                     /note="synthetic polyadenylation signal"
     enhancer        6525..6828
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6830..7032
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     6983..7003
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      join(7101..7102,1..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 agctagcttg actgactgag atacagcgta ccttcagctc acagacatga taagatacat
     1081 tgatgagttt ggacaaacca caactagaat gcagtgaaaa aaatgcttta tttgtgaaat
     1141 ttgtgatgct attgctttat ttgtaaccat tataagctgc aataaacaag ttaacaacaa
     1201 caattgcatt cattttatgt ttcaggttca gggggaggtg tgggaggttt tttaaagcaa
     1261 gtaaaacctc tacaaatgtg gtattggccc atctctatcg gtatcgtagc ataacccctt
     1321 ggggcctcta aacgggtctt gaggggtttt ttgtgcccct cgggccggat tgctatctac
     1381 cggcattggc gcagaaaaaa atgcctgatg cgacgctgcg cgtcttatac tcccacatat
     1441 gccagattca gcaacggata cggcttcccc aacttgccca cttccatacg tgtcctcctt
     1501 accagaaatt tatccttaag gtcgtcagct atcctgcagg cgatctctcg atttcgatca
     1561 agacattcct ttaatggtct tttctggaca ccactagggg tcagaagtag ttcatcaaac
     1621 tttcttccct ccctaatctc attggttacc ttgggctatc gaaacttaat taaccagtca
     1681 agtcagctac ttggcgagat cgacttgtct gggtttcgac tacgctcaga attgcgtcag
     1741 tcaagttcga tctggtcctt gctattgcac ccgttctccg attacgagtt tcatttaaat
     1801 catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt
     1861 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
     1921 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg
     1981 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
     2041 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
     2101 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
     2161 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
     2221 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc
     2281 taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac
     2341 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg
     2401 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt
     2461 gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt
     2521 catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa
     2581 atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga
     2641 ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcattta aatttccgaa
     2701 ctctccaagg ccctcgtcgg aaaatcttca aacctttcgt ccgatccatc ttgcaggcta
     2761 cctctcgaac gaactatcgc aagtctcttg gccggccttg cgccttggct attgcttggc
     2821 agcgcctatc gccaggtatt actccaatcc cgaatatccg agatcgggat cacccgagag
     2881 aagttcaacc tacatcctca atcccgatct atccgagatc cgaggaatat cgaaatcggg
     2941 gcgcgcctgg cctccgcgcc gggttttggc gcctcccgcg ggcgcccccc tcgtcacggc
     3001 gagcgctgcc acgtcagacg aagggcgcag gagcgtcctg atccttccgc ccggacgctc
     3061 aggacagcgg cccgctgctc ataagactcg gccttagaac cccagtatca gcagaaggac
     3121 attttaggac gggacttggg tgactctagg gcactggttt tctttccaga gagcggaaca
     3181 ggcgaggaaa agtagtccct tctcggcgat tctgcggagg gatctccgtg gggcggtgaa
     3241 cgccgatgat tatataagga cgcgccgggt gtggcacagc tagttccgtc gcagccggga
     3301 tttgggtcgc ggttcttgtt tgtggatcgc tgtgatcgtc acttggtgag tagcgggctg
     3361 ctgggctggc cggggctttc gtggccgccg ggccgctcgg tgggacggaa gcgtgtggag
     3421 agaccgccaa gggctgtagt ctgggtccgc gagcaaggtt gccctgaact gggggttggg
     3481 gggagcgcag caaaatggcg gctgttcccg agtcttgaat ggaagacgct tgtgaggcgg
     3541 gctgtgaggt cgttgaaaca aggtgggggg catggtgggc ggcaagaacc caaggtcttg
     3601 agcccttcgc taatgcggga aagctcttat tcgggtgaga tgggctgggc accatctggg
     3661 gaccctgacg tgaagtttgt cactgactgg agaactcggt ttgtcgtctg ttgcgggggc
     3721 ggcagttatg gcggtgccgt tgggcagtgc acccgtacct ttgggagcgc gcgccctcgt
     3781 cgtgtcgtga cgtcacccgt tctgttggct tataatgcag ggtggggcca cctgccggta
     3841 ggtgtgcggt aggcttttct ccgtcgcagg acgcagggtt cgggcctagg gtaggctctc
     3901 ctgaatcgac aggcgccgga cctctggtga ggggagggat aagtgaggcg tcagtttctt
     3961 tggtcggttt tatgtaccta tcttcttaag tagctgaagc tccggttttg aactatgcgc
     4021 tcggggttgg cgagtgtgtt ttgtgaagtt ttttaggcac cttttgaaat gtaatcattt
     4081 gggtcaatat gtaattttca gtgttagact tgtaaattgt ccgctaaatt ctggccgttt
     4141 ttggcttttt tgttagacaa catgaccgag tacaagccca cggtgcgcct cgccacccgc
     4201 gacgacgtcc ccagggccgt acgcaccctc gccgccgcgt tcgccgacta ccccgccacg
     4261 cgccacaccg tcgatccgga ccgccacatc gagcgggtca ccgagctgca agaactcttc
     4321 ctcacgcgcg tcgggctcga catcggcaag gtgtgggtcg cggacgacgg cgccgcggtg
     4381 gcggtctgga ccacgccgga gagcgtcgaa gcgggggcgg tgttcgccga gatcggcccg
     4441 cgcatggccg agttgagcgg ttcccggctg gccgcgcagc aacagatgga gggcctcctg
     4501 gcgccgcacc ggcccaagga gcccgcgtgg ttcctggcca ccgtcggcgt ctcgcccgac
     4561 caccagggca agggtctggg cagcgccgtc gtgctccccg gagtggaggc cgccgagcgc
     4621 gccggggtgc ccgccttcct ggagacctcc gcgccccgca acctcccctt ctacgagcgg
     4681 ctcggcttca ccgtcaccgc cgacgtcgag gtgcccgaag gaccgcgcac ctggtgcatg
     4741 acccgcaagc ccggtgccta ggctagtatg taagcctagt cttagataat aaaatcgcta
     4801 tccatcgaag atggatgtgt gttggttttt tgtgtgtgta acgctaggcg cgcctggtgt
     4861 accgagaacg atcctctcag tgcgagtctc gacgatccat atcgttgctt ggcagtcagc
     4921 cagtcggaat ccagcttggg acccaggaag tccaatcgtc agatattgta ctcaagcctg
     4981 gtcacggcag cgtaccgatc tgtttaaacc tagatattga tagtctgatc ggtcaacgta
     5041 taatcgagtc ctagcttttg caaacatcta tcaagagaca ggatcagcag gaggctttcg
     5101 catgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc
     5161 tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc
     5221 gcgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc
     5281 cgaagaacgc tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc
     5341 ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt
     5401 ggttgagtat tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt
     5461 atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat
     5521 tggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct
     5581 tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat
     5641 gcctgtagca atggcaacaa ccttgcgtaa actattaact ggcgaactac ttactctagc
     5701 ttcccggcaa cagttgatag actggatgga ggcggataaa gttgcaggac cacttctgcg
     5761 ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc
     5821 tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta
     5881 cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc
     5941 ctcactgatt aagcattggt aaccgattct aggtgcattg gcgcagaaaa aaatgcctga
     6001 tgcgacgctg cgcgtcttat actcccacat atgccagatt cagcaacgga tacggcttcc
     6061 ccaacttgcc cacttccata cgtgtcctcc ttaccagaaa tttatcctta agatcgttta
     6121 aactcgactc tggctctatc gaatctccgt cgtttcgagc ttacgcgaac agccgtggcg
     6181 ctcatttgct cgtcgggcat cgaatctcgt cagctatcgt cagcttacct ttttggcagc
     6241 gatcgcggct cccgacatct tggaccatta gctccacagg tatcttcttc cctctagtgg
     6301 tcataacagc agcttcagct acctctcaat tcaaaaaacc cctcaagacc cgtttagagg
     6361 ccccaagggg ttatgctatc aatcgttgcg ttacacacac aaaaaaccaa cacacatcca
     6421 tcttcgatgg atagcgattt tattatctaa ctgctgatcg agtgtagcca gatctagtaa
     6481 tcaattacgg ggtcattagt tcatagccca tatatggagt tccgcgttac ataacttacg
     6541 gtaaatggcc cgcctggctg accgcccaac gacccccgcc cattgacgtc aataatgacg
     6601 tatgttccca tagtaacgcc aatagggact ttccattgac gtcaatgggt ggagtattta
     6661 cggtaaactg cccacttggc agtacatcaa gtgtatcata tgccaagtac gccccctatt
     6721 gacgtcaatg acggtaaatg gcccgcctgg cattatgccc agtacatgac cttatgggac
     6781 tttcctactt ggcagtacat ctacgtatta gtcatcgcta ttaccatgct gatgcggttt
     6841 tggcagtaca tcaatgggcg tggatagcgg tttgactcac ggggatttcc aagtctccac
     6901 cccattgacg tcaatgggag tttgttttgg caccaaaatc aacgggactt tccaaaatgt
     6961 cgtaacaact ccgccccatt gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat
     7021 ataagcagag ctggtttagt gaaccgtcag atcagatctt tgtcgatcct accatccact
     7081 cgacacaccc gccagcggcc gc
//
