LOCUS       sequence-295804-        7093 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 7093)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 7093)
  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-m4133377
FEATURES             Location/Qualifiers
     source          1..7093
                     /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             944..1009
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     polyA_signal    1062..1183
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1073..1092)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1127..1146
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      1298..1344
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1374..1510
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1555..1626
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(1852..2440)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1932..1951)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2940..4149
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     3986..4005
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             4153..4752
                     /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(4153..4172)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4649..4669
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4779..4827
                     /note="synthetic polyadenylation signal"
     CDS             join(5093..5161,5162..5953)
                     /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(5311..5330)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      5966..6102
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    6247..6318
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(6323..6369)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    6387..6435
                     /note="synthetic polyadenylation signal"
     enhancer        6516..6819
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6821..7023
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     6974..6994
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      join(7092..7093,1..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 atggattata aggaccacga
      961 tggagactac aaagatcatg acattgatta caaggacgac gatgacaaat aagctagctt
     1021 gactgactga gatacagcgt accttcagct cacagacatg ataagataca ttgatgagtt
     1081 tggacaaacc acaactagaa tgcagtgaaa aaaatgcttt atttgtgaaa tttgtgatgc
     1141 tattgcttta tttgtaacca ttataagctg caataaacaa gttaacaaca acaattgcat
     1201 tcattttatg tttcaggttc agggggaggt gtgggaggtt ttttaaagca agtaaaacct
     1261 ctacaaatgt ggtattggcc catctctatc ggtatcgtag cataacccct tggggcctct
     1321 aaacgggtct tgaggggttt tttgtgcccc tcgggccgga ttgctatcta ccggcattgg
     1381 cgcagaaaaa aatgcctgat gcgacgctgc gcgtcttata ctcccacata tgccagattc
     1441 agcaacggat acggcttccc caacttgccc acttccatac gtgtcctcct taccagaaat
     1501 ttatccttaa ggtcgtcagc tatcctgcag gcgatctctc gatttcgatc aagacattcc
     1561 tttaatggtc ttttctggac accactaggg gtcagaagta gttcatcaaa ctttcttccc
     1621 tccctaatct cattggttac cttgggctat cgaaacttaa ttaaccagtc aagtcagcta
     1681 cttggcgaga tcgacttgtc tgggtttcga ctacgctcag aattgcgtca gtcaagttcg
     1741 atctggtcct tgctattgca cccgttctcc gattacgagt ttcatttaaa tcatgtgagc
     1801 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag
     1861 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc
     1921 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt
     1981 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
     2041 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
     2101 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
     2161 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat
     2221 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg
     2281 ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa
     2341 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt
     2401 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc
     2461 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt
     2521 atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta
     2581 aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat
     2641 ctcagcgatc tgtctatttc gttcatccat agttgcattt aaatttccga actctccaag
     2701 gccctcgtcg gaaaatcttc aaacctttcg tccgatccat cttgcaggct acctctcgaa
     2761 cgaactatcg caagtctctt ggccggcctt gcgccttggc tattgcttgg cagcgcctat
     2821 cgccaggtat tactccaatc ccgaatatcc gagatcggga tcacccgaga gaagttcaac
     2881 ctacatcctc aatcccgatc tatccgagat ccgaggaata tcgaaatcgg ggcgcgcctg
     2941 gcctccgcgc cgggttttgg cgcctcccgc gggcgccccc ctcgtcacgg cgagcgctgc
     3001 cacgtcagac gaagggcgca ggagcgtcct gatccttccg cccggacgct caggacagcg
     3061 gcccgctgct cataagactc ggccttagaa ccccagtatc agcagaagga cattttagga
     3121 cgggacttgg gtgactctag ggcactggtt ttctttccag agagcggaac aggcgaggaa
     3181 aagtagtccc ttctcggcga ttctgcggag ggatctccgt ggggcggtga acgccgatga
     3241 ttatataagg acgcgccggg tgtggcacag ctagttccgt cgcagccggg atttgggtcg
     3301 cggttcttgt ttgtggatcg ctgtgatcgt cacttggtga gtagcgggct gctgggctgg
     3361 ccggggcttt cgtggccgcc gggccgctcg gtgggacgga agcgtgtgga gagaccgcca
     3421 agggctgtag tctgggtccg cgagcaaggt tgccctgaac tgggggttgg ggggagcgca
     3481 gcaaaatggc ggctgttccc gagtcttgaa tggaagacgc ttgtgaggcg ggctgtgagg
     3541 tcgttgaaac aaggtggggg gcatggtggg cggcaagaac ccaaggtctt gagcccttcg
     3601 ctaatgcggg aaagctctta ttcgggtgag atgggctggg caccatctgg ggaccctgac
     3661 gtgaagtttg tcactgactg gagaactcgg tttgtcgtct gttgcggggg cggcagttat
     3721 ggcggtgccg ttgggcagtg cacccgtacc tttgggagcg cgcgccctcg tcgtgtcgtg
     3781 acgtcacccg ttctgttggc ttataatgca gggtggggcc acctgccggt aggtgtgcgg
     3841 taggcttttc tccgtcgcag gacgcagggt tcgggcctag ggtaggctct cctgaatcga
     3901 caggcgccgg acctctggtg aggggaggga taagtgaggc gtcagtttct ttggtcggtt
     3961 ttatgtacct atcttcttaa gtagctgaag ctccggtttt gaactatgcg ctcggggttg
     4021 gcgagtgtgt tttgtgaagt tttttaggca ccttttgaaa tgtaatcatt tgggtcaata
     4081 tgtaattttc agtgttagac ttgtaaattg tccgctaaat tctggccgtt tttggctttt
     4141 ttgttagaca acatgaccga gtacaagccc acggtgcgcc tcgccacccg cgacgacgtc
     4201 cccagggccg tacgcaccct cgccgccgcg ttcgccgact accccgccac gcgccacacc
     4261 gtcgatccgg accgccacat cgagcgggtc accgagctgc aagaactctt cctcacgcgc
     4321 gtcgggctcg acatcggcaa ggtgtgggtc gcggacgacg gcgccgcggt ggcggtctgg
     4381 accacgccgg agagcgtcga agcgggggcg gtgttcgccg agatcggccc gcgcatggcc
     4441 gagttgagcg gttcccggct ggccgcgcag caacagatgg agggcctcct ggcgccgcac
     4501 cggcccaagg agcccgcgtg gttcctggcc accgtcggcg tctcgcccga ccaccagggc
     4561 aagggtctgg gcagcgccgt cgtgctcccc ggagtggagg ccgccgagcg cgccggggtg
     4621 cccgccttcc tggagacctc cgcgccccgc aacctcccct tctacgagcg gctcggcttc
     4681 accgtcaccg ccgacgtcga ggtgcccgaa ggaccgcgca cctggtgcat gacccgcaag
     4741 cccggtgcct aggctagtat gtaagcctag tcttagataa taaaatcgct atccatcgaa
     4801 gatggatgtg tgttggtttt ttgtgtgtgt aacgctaggc gcgcctggtg taccgagaac
     4861 gatcctctca gtgcgagtct cgacgatcca tatcgttgct tggcagtcag ccagtcggaa
     4921 tccagcttgg gacccaggaa gtccaatcgt cagatattgt actcaagcct ggtcacggca
     4981 gcgtaccgat ctgtttaaac ctagatattg atagtctgat cggtcaacgt ataatcgagt
     5041 cctagctttt gcaaacatct atcaagagac aggatcagca ggaggctttc gcatgagtat
     5101 tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc
     5161 tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg cgcgagtggg
     5221 ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg
     5281 ctttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga
     5341 cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta
     5401 ttcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc
     5461 tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga ttggaggacc
     5521 gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg
     5581 ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc
     5641 aatggcaaca accttgcgta aactattaac tggcgaacta cttactctag cttcccggca
     5701 acagttgata gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct
     5761 tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat
     5821 cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg
     5881 gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat
     5941 taagcattgg taaccgattc taggtgcatt ggcgcagaaa aaaatgcctg atgcgacgct
     6001 gcgcgtctta tactcccaca tatgccagat tcagcaacgg atacggcttc cccaacttgc
     6061 ccacttccat acgtgtcctc cttaccagaa atttatcctt aagatcgttt aaactcgact
     6121 ctggctctat cgaatctccg tcgtttcgag cttacgcgaa cagccgtggc gctcatttgc
     6181 tcgtcgggca tcgaatctcg tcagctatcg tcagcttacc tttttggcag cgatcgcggc
     6241 tcccgacatc ttggaccatt agctccacag gtatcttctt ccctctagtg gtcataacag
     6301 cagcttcagc tacctctcaa ttcaaaaaac ccctcaagac ccgtttagag gccccaaggg
     6361 gttatgctat caatcgttgc gttacacaca caaaaaacca acacacatcc atcttcgatg
     6421 gatagcgatt ttattatcta actgctgatc gagtgtagcc agatctagta atcaattacg
     6481 gggtcattag ttcatagccc atatatggag ttccgcgtta cataacttac ggtaaatggc
     6541 ccgcctggct gaccgcccaa cgacccccgc ccattgacgt caataatgac gtatgttccc
     6601 atagtaacgc caatagggac tttccattga cgtcaatggg tggagtattt acggtaaact
     6661 gcccacttgg cagtacatca agtgtatcat atgccaagta cgccccctat tgacgtcaat
     6721 gacggtaaat ggcccgcctg gcattatgcc cagtacatga ccttatggga ctttcctact
     6781 tggcagtaca tctacgtatt agtcatcgct attaccatgc tgatgcggtt ttggcagtac
     6841 atcaatgggc gtggatagcg gtttgactca cggggatttc caagtctcca ccccattgac
     6901 gtcaatggga gtttgttttg gcaccaaaat caacgggact ttccaaaatg tcgtaacaac
     6961 tccgccccat tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta tataagcaga
     7021 gctggtttag tgaaccgtca gatcagatct ttgtcgatcc taccatccac tcgacacacc
     7081 cgccagcggc cgc
//
