LOCUS       sequence-295332-        7420 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 7420)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 7420)
  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-m4133623
FEATURES             Location/Qualifiers
     source          1..7420
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             926..946
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             1295..1312
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     polyA_signal    1389..1510
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1400..1419)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1454..1473
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      1625..1671
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1701..1837
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1882..1953
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(2179..2767)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2259..2278)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        3267..4476
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     4313..4332
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             4480..5079
                     /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(4480..4499)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4976..4996
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    5106..5154
                     /note="synthetic polyadenylation signal"
     CDS             join(5420..5488,5489..6280)
                     /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(5638..5657)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      6293..6429
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    6574..6645
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(6650..6696)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    6714..6762
                     /note="synthetic polyadenylation signal"
     enhancer        6843..7146
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        7148..7350
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     7301..7321
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
ORIGIN
        1 caccatgagt gggttccgga aaatggcgtt tccctccggt aaggtcgaag gatgcatggt
       61 tcaggtgacc tgtggcacaa ctacgttaaa cggcctatgg ctggacgatg tggtatactg
      121 tcctcgacac gtcatctgca catcagagga catgctgaat ccaaactatg aggatttgct
      181 cataagaaag agcaatcata actttctggt gcaagctgga aatgtacagc ttagggttat
      241 tgggcactct atgcagaact gcgtgttaaa actcaaggtg gataccgcca atccgaaaac
      301 acctaagtac aaattcgtgc gcatccagcc aggccaaacc ttctctgtcc tggcatgtta
      361 taacggcagc ccctcaggtg tctaccagtg cgccatgcgc cctaatttta ctattaaggg
      421 ttccttcttg aacgggtcgt gcggatcggt cggctttaat atcgactatg actgtgtgag
      481 tttttgctac atgcatcaca tggaacttcc gactggggtt cacgctggaa cggatctgga
      541 gggtaacttc tatggaccct tcgtggatag acagaccgcc caggcagccg gcactgacac
      601 taccattacg gtgaatgttc tagcttggct ctacgcagct gtaataaacg gcgatcgttg
      661 gttcttgaac aggtttacaa caaccctgaa cgactttaat ctagtcgcca tgaaatataa
      721 ttatgaacca ctgacccaag atcatgtaga cattctcggg ccactgagcg cacagacagg
      781 gatcgctgtg ctggacatgt gtgccagttt aaaggagctg cttcagaatg gtatgaacgg
      841 ccgaacaatc ctagggtctg cgctcttaga ggatgaattt acccccttcg atgtagtgcg
      901 gcaatgtagc ggcgttactt ttcaagaaaa cctttacttt cagtcgatga gtgacaaaat
      961 cattcatcta accgatgact cctttgatac agacgtgttg aaggccgatg gtgcgatact
     1021 ggtagatttc tgggctgaat ggtgtggccc gtgcaagatg atcgcaccca ttctggacga
     1081 gatcgccgac gagtatcaag gaaaattaac tgtcgctaag cttaatatag atcagaaccc
     1141 agggacggca cctaaatacg gcatcagggg aatacctacc ctcctgctct ttaagaatgg
     1201 ggaagttgcc gcgacaaaag tgggagccct gtctaagggc cagctaaaag agttcttgga
     1261 cgctaacctg gcaggttcag gctcggggca catgcaccat caccatcacc atagcagcgg
     1321 gctcgtgcca cgcggctaag ctagcttgac tgactgagat acagcgtacc ttcagctcac
     1381 agacatgata agatacattg atgagtttgg acaaaccaca actagaatgc agtgaaaaaa
     1441 atgctttatt tgtgaaattt gtgatgctat tgctttattt gtaaccatta taagctgcaa
     1501 taaacaagtt aacaacaaca attgcattca ttttatgttt caggttcagg gggaggtgtg
     1561 ggaggttttt taaagcaagt aaaacctcta caaatgtggt attggcccat ctctatcggt
     1621 atcgtagcat aaccccttgg ggcctctaaa cgggtcttga ggggtttttt gtgcccctcg
     1681 ggccggattg ctatctaccg gcattggcgc agaaaaaaat gcctgatgcg acgctgcgcg
     1741 tcttatactc ccacatatgc cagattcagc aacggatacg gcttccccaa cttgcccact
     1801 tccatacgtg tcctccttac cagaaattta tccttaaggt cgtcagctat cctgcaggcg
     1861 atctctcgat ttcgatcaag acattccttt aatggtcttt tctggacacc actaggggtc
     1921 agaagtagtt catcaaactt tcttccctcc ctaatctcat tggttacctt gggctatcga
     1981 aacttaatta accagtcaag tcagctactt ggcgagatcg acttgtctgg gtttcgacta
     2041 cgctcagaat tgcgtcagtc aagttcgatc tggtccttgc tattgcaccc gttctccgat
     2101 tacgagtttc atttaaatca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     2161 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     2221 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     2281 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     2341 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
     2401 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     2461 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     2521 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     2581 gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc
     2641 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     2701 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     2761 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     2821 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
     2881 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta
     2941 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     3001 tgcatttaaa tttccgaact ctccaaggcc ctcgtcggaa aatcttcaaa cctttcgtcc
     3061 gatccatctt gcaggctacc tctcgaacga actatcgcaa gtctcttggc cggccttgcg
     3121 ccttggctat tgcttggcag cgcctatcgc caggtattac tccaatcccg aatatccgag
     3181 atcgggatca cccgagagaa gttcaaccta catcctcaat cccgatctat ccgagatccg
     3241 aggaatatcg aaatcggggc gcgcctggcc tccgcgccgg gttttggcgc ctcccgcggg
     3301 cgcccccctc gtcacggcga gcgctgccac gtcagacgaa gggcgcagga gcgtcctgat
     3361 ccttccgccc ggacgctcag gacagcggcc cgctgctcat aagactcggc cttagaaccc
     3421 cagtatcagc agaaggacat tttaggacgg gacttgggtg actctagggc actggttttc
     3481 tttccagaga gcggaacagg cgaggaaaag tagtcccttc tcggcgattc tgcggaggga
     3541 tctccgtggg gcggtgaacg ccgatgatta tataaggacg cgccgggtgt ggcacagcta
     3601 gttccgtcgc agccgggatt tgggtcgcgg ttcttgtttg tggatcgctg tgatcgtcac
     3661 ttggtgagta gcgggctgct gggctggccg gggctttcgt ggccgccggg ccgctcggtg
     3721 ggacggaagc gtgtggagag accgccaagg gctgtagtct gggtccgcga gcaaggttgc
     3781 cctgaactgg gggttggggg gagcgcagca aaatggcggc tgttcccgag tcttgaatgg
     3841 aagacgcttg tgaggcgggc tgtgaggtcg ttgaaacaag gtggggggca tggtgggcgg
     3901 caagaaccca aggtcttgag cccttcgcta atgcgggaaa gctcttattc gggtgagatg
     3961 ggctgggcac catctgggga ccctgacgtg aagtttgtca ctgactggag aactcggttt
     4021 gtcgtctgtt gcgggggcgg cagttatggc ggtgccgttg ggcagtgcac ccgtaccttt
     4081 gggagcgcgc gccctcgtcg tgtcgtgacg tcacccgttc tgttggctta taatgcaggg
     4141 tggggccacc tgccggtagg tgtgcggtag gcttttctcc gtcgcaggac gcagggttcg
     4201 ggcctagggt aggctctcct gaatcgacag gcgccggacc tctggtgagg ggagggataa
     4261 gtgaggcgtc agtttctttg gtcggtttta tgtacctatc ttcttaagta gctgaagctc
     4321 cggttttgaa ctatgcgctc ggggttggcg agtgtgtttt gtgaagtttt ttaggcacct
     4381 tttgaaatgt aatcatttgg gtcaatatgt aattttcagt gttagacttg taaattgtcc
     4441 gctaaattct ggccgttttt ggcttttttg ttagacaaca tgaccgagta caagcccacg
     4501 gtgcgcctcg ccacccgcga cgacgtcccc agggccgtac gcaccctcgc cgccgcgttc
     4561 gccgactacc ccgccacgcg ccacaccgtc gatccggacc gccacatcga gcgggtcacc
     4621 gagctgcaag aactcttcct cacgcgcgtc gggctcgaca tcggcaaggt gtgggtcgcg
     4681 gacgacggcg ccgcggtggc ggtctggacc acgccggaga gcgtcgaagc gggggcggtg
     4741 ttcgccgaga tcggcccgcg catggccgag ttgagcggtt cccggctggc cgcgcagcaa
     4801 cagatggagg gcctcctggc gccgcaccgg cccaaggagc ccgcgtggtt cctggccacc
     4861 gtcggcgtct cgcccgacca ccagggcaag ggtctgggca gcgccgtcgt gctccccgga
     4921 gtggaggccg ccgagcgcgc cggggtgccc gccttcctgg agacctccgc gccccgcaac
     4981 ctccccttct acgagcggct cggcttcacc gtcaccgccg acgtcgaggt gcccgaagga
     5041 ccgcgcacct ggtgcatgac ccgcaagccc ggtgcctagg ctagtatgta agcctagtct
     5101 tagataataa aatcgctatc catcgaagat ggatgtgtgt tggttttttg tgtgtgtaac
     5161 gctaggcgcg cctggtgtac cgagaacgat cctctcagtg cgagtctcga cgatccatat
     5221 cgttgcttgg cagtcagcca gtcggaatcc agcttgggac ccaggaagtc caatcgtcag
     5281 atattgtact caagcctggt cacggcagcg taccgatctg tttaaaccta gatattgata
     5341 gtctgatcgg tcaacgtata atcgagtcct agcttttgca aacatctatc aagagacagg
     5401 atcagcagga ggctttcgca tgagtattca acatttccgt gtcgccctta ttcccttttt
     5461 tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc
     5521 tgaagatcag ttgggtgcgc gagtgggtta catcgaactg gatctcaaca gcggtaagat
     5581 ccttgagagt tttcgccccg aagaacgctt tccaatgatg agcactttta aagttctgct
     5641 atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca
     5701 ctattctcag aatgacttgg ttgagtattc accagtcaca gaaaagcatc ttacggatgg
     5761 catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa
     5821 cttacttctg acaacgattg gaggaccgaa ggagctaacc gcttttttgc acaacatggg
     5881 ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga
     5941 cgagcgtgac accacgatgc ctgtagcaat ggcaacaacc ttgcgtaaac tattaactgg
     6001 cgaactactt actctagctt cccggcaaca gttgatagac tggatggagg cggataaagt
     6061 tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg
     6121 agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc
     6181 ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca
     6241 gatcgctgag ataggtgcct cactgattaa gcattggtaa ccgattctag gtgcattggc
     6301 gcagaaaaaa atgcctgatg cgacgctgcg cgtcttatac tcccacatat gccagattca
     6361 gcaacggata cggcttcccc aacttgccca cttccatacg tgtcctcctt accagaaatt
     6421 tatccttaag atcgtttaaa ctcgactctg gctctatcga atctccgtcg tttcgagctt
     6481 acgcgaacag ccgtggcgct catttgctcg tcgggcatcg aatctcgtca gctatcgtca
     6541 gcttaccttt ttggcagcga tcgcggctcc cgacatcttg gaccattagc tccacaggta
     6601 tcttcttccc tctagtggtc ataacagcag cttcagctac ctctcaattc aaaaaacccc
     6661 tcaagacccg tttagaggcc ccaaggggtt atgctatcaa tcgttgcgtt acacacacaa
     6721 aaaaccaaca cacatccatc ttcgatggat agcgatttta ttatctaact gctgatcgag
     6781 tgtagccaga tctagtaatc aattacgggg tcattagttc atagcccata tatggagttc
     6841 cgcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga cccccgccca
     6901 ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt ccattgacgt
     6961 caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt gtatcatatg
     7021 ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag
     7081 tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt catcgctatt
     7141 accatgctga tgcggttttg gcagtacatc aatgggcgtg gatagcggtt tgactcacgg
     7201 ggatttccaa gtctccaccc cattgacgtc aatgggagtt tgttttggca ccaaaatcaa
     7261 cgggactttc caaaatgtcg taacaactcc gccccattga cgcaaatggg cggtaggcgt
     7321 gtacggtggg aggtctatat aagcagagct ggtttagtga accgtcagat cagatctttg
     7381 tcgatcctac catccactcg acacacccgc cagcggccgc
//
