LOCUS       sequence-300772-        7500 bp    DNA     circular SYN 15-APR-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 7500)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 7500)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: pGBW-m4133256
FEATURES             Location/Qualifiers
     source          1..7500
                     /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             947..964
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     polyA_signal    1033..1154
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1044..1063)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1098..1117
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      1269..1315
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1345..1481
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1526..1597
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(1823..2411)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1903..1922)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2911..4120
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     3957..3976
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             4124..5152
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     polyA_signal    5179..5227
                     /note="synthetic polyadenylation signal"
     CDS             join(5493..5561,5562..6353)
                     /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(5711..5730)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      6366..6502
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    6654..6725
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(6730..6776)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    6794..6842
                     /note="synthetic polyadenylation signal"
     enhancer        6923..7226
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        7228..7430
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     7381..7401
                     /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 cagtcgcacc atcaccatca
      961 ccattaagga tccgcaggcc tctgctagct tgactgactg agatacagcg taccttcagc
     1021 tcacagacat gataagatac attgatgagt ttggacaaac cacaactaga atgcagtgaa
     1081 aaaaatgctt tatttgtgaa atttgtgatg ctattgcttt atttgtaacc attataagct
     1141 gcaataaaca agttaacaac aacaattgca ttcattttat gtttcaggtt cagggggagg
     1201 tgtgggaggt tttttaaagc aagtaaaacc tctacaaatg tggtattggc ccatctctat
     1261 cggtatcgta gcataacccc ttggggcctc taaacgggtc ttgaggggtt ttttgtgccc
     1321 ctcgggccgg attgctatct accggcattg gcgcagaaaa aaatgcctga tgcgacgctg
     1381 cgcgtcttat actcccacat atgccagatt cagcaacgga tacggcttcc ccaacttgcc
     1441 cacttccata cgtgtcctcc ttaccagaaa tttatcctta aggtcgtcag ctatcctgca
     1501 ggcgatctct cgatttcgat caagacattc ctttaatggt cttttctgga caccactagg
     1561 ggtcagaagt agttcatcaa actttcttcc ctccctaatc tcattggtta ccttgggcta
     1621 tcgaaactta attaaccagt caagtcagct acttggcgag atcgacttgt ctgggtttcg
     1681 actacgctca gaattgcgtc agtcaagttc gatctggtcc ttgctattgc acccgttctc
     1741 cgattacgag tttcatttaa atcatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     1801 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     1861 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     1921 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     1981 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     2041 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     2101 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     2161 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     2221 cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct
     2281 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     2341 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     2401 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa
     2461 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt
     2521 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca
     2581 gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca
     2641 tagttgcatt taaatttccg aactctccaa ggccctcgtc ggaaaatctt caaacctttc
     2701 gtccgatcca tcttgcaggc tacctctcga acgaactatc gcaagtctct tggccggcct
     2761 tgcgccttgg ctattgcttg gcagcgccta tcgccaggta ttactccaat cccgaatatc
     2821 cgagatcggg atcacccgag agaagttcaa cctacatcct caatcccgat ctatccgaga
     2881 tccgaggaat atcgaaatcg gggcgcgcct ggcctccgcg ccgggttttg gcgcctcccg
     2941 cgggcgcccc cctcgtcacg gcgagcgctg ccacgtcaga cgaagggcgc aggagcgtcc
     3001 tgatccttcc gcccggacgc tcaggacagc ggcccgctgc tcataagact cggccttaga
     3061 accccagtat cagcagaagg acattttagg acgggacttg ggtgactcta gggcactggt
     3121 tttctttcca gagagcggaa caggcgagga aaagtagtcc cttctcggcg attctgcgga
     3181 gggatctccg tggggcggtg aacgccgatg attatataag gacgcgccgg gtgtggcaca
     3241 gctagttccg tcgcagccgg gatttgggtc gcggttcttg tttgtggatc gctgtgatcg
     3301 tcacttggtg agtagcgggc tgctgggctg gccggggctt tcgtggccgc cgggccgctc
     3361 ggtgggacgg aagcgtgtgg agagaccgcc aagggctgta gtctgggtcc gcgagcaagg
     3421 ttgccctgaa ctgggggttg gggggagcgc agcaaaatgg cggctgttcc cgagtcttga
     3481 atggaagacg cttgtgaggc gggctgtgag gtcgttgaaa caaggtgggg ggcatggtgg
     3541 gcggcaagaa cccaaggtct tgagcccttc gctaatgcgg gaaagctctt attcgggtga
     3601 gatgggctgg gcaccatctg gggaccctga cgtgaagttt gtcactgact ggagaactcg
     3661 gtttgtcgtc tgttgcgggg gcggcagtta tggcggtgcc gttgggcagt gcacccgtac
     3721 ctttgggagc gcgcgccctc gtcgtgtcgt gacgtcaccc gttctgttgg cttataatgc
     3781 agggtggggc cacctgccgg taggtgtgcg gtaggctttt ctccgtcgca ggacgcaggg
     3841 ttcgggccta gggtaggctc tcctgaatcg acaggcgccg gacctctggt gaggggaggg
     3901 ataagtgagg cgtcagtttc tttggtcggt tttatgtacc tatcttctta agtagctgaa
     3961 gctccggttt tgaactatgc gctcggggtt ggcgagtgtg ttttgtgaag ttttttaggc
     4021 accttttgaa atgtaatcat ttgggtcaat atgtaatttt cagtgttaga cttgtaaatt
     4081 gtccgctaaa ttctggccgt ttttggcttt tttgttagac aacatgggta aaaagcctga
     4141 actcaccgcg acgtctgtcg agaagtttct gatcgaaaag ttcgacagcg tctccgacct
     4201 gatgcagctc tcggagggcg aagaatctcg tgctttcagc ttcgatgtag gagggcgtgg
     4261 atatgtcctg cgggtaaata gctgcgccga tggtttctac aaagatcgtt atgtttatcg
     4321 gcactttgca tccgccgcgc tcccgattcc ggaagtgctt gacattgggg agttcagcga
     4381 gagcctgacc tattgcatct cccgccgtgc acagggtgtc acgttgcaag acctgcctga
     4441 aaccgaactg cccgctgttc tgcagccggt cgcggaggca atggatgcca tcgctgccgc
     4501 cgatcttagc cagacgagcg ggttcggccc attcggaccg caaggaatcg gtcaatacac
     4561 tacatggcgt gatttcatat gcgcgattgc tgatccccat gtgtatcact ggcaaactgt
     4621 gatggacgac accgtcagtg cgtccgtcgc gcaggctctc gatgagctga tgctttgggc
     4681 cgaggactgc cccgaagtcc ggcacctcgt gcacgcggat ttcggctcca acaatgtcct
     4741 gacggacaat ggccgcataa cagcggtcat tgactggagc gaggcgatgt tcggggattc
     4801 ccaatacgag gtcgccaaca tcttcttctg gaggccgtgg ttggcttgta tggagcagca
     4861 gacgcgctac ttcgagcgga ggcatccgga gcttgcagga tcgccccggc tccgggcgta
     4921 tatgctccgc attggtcttg accaactcta tcagagcttg gttgacggca atttcgatga
     4981 tgcagcttgg gcgcagggtc gatgcgacgc aatcgtccga tccggagccg ggactgtcgg
     5041 gcgtacacaa atcgcccgca gaagcgccgc cgtctggacc gatggctgtg tagaagtact
     5101 cgccgatagt ggaaaccgac gccccagcac tcgtccgagg gcaaaggaat aagctagtat
     5161 gtaagcctag tcttagataa taaaatcgct atccatcgaa gatggatgtg tgttggtttt
     5221 ttgtgtgtgt aacgctaggc gcgcctggtg taccgagaac gatcctctca gtgcgagtct
     5281 cgacgatcca tatcgttgct tggcagtcag ccagtcggaa tccagcttgg gacccaggaa
     5341 gtccaatcgt cagatattgt actcaagcct ggtcacggca gcgtaccgat ctgtttaaac
     5401 ctagatattg atagtctgat cggtcaacgt ataatcgagt cctagctttt gcaaacatct
     5461 atcaagagac aggatcagca ggaggctttc gcatgagtat tcaacatttc cgtgtcgccc
     5521 ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga
     5581 aagtaaaaga tgctgaagat cagttgggtg cgcgagtggg ttacatcgaa ctggatctca
     5641 acagcggtaa gatccttgag agttttcgcc ccgaagaacg ctttccaatg atgagcactt
     5701 ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg
     5761 gtcgccgcat acactattct cagaatgact tggttgagta ttcaccagtc acagaaaagc
     5821 atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata
     5881 acactgcggc caacttactt ctgacaacga ttggaggacc gaaggagcta accgcttttt
     5941 tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag
     6001 ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca accttgcgta
     6061 aactattaac tggcgaacta cttactctag cttcccggca acagttgata gactggatgg
     6121 aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg
     6181 ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag
     6241 atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg
     6301 aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg taaccgattc
     6361 taggtgcatt ggcgcagaaa aaaatgcctg atgcgacgct gcgcgtctta tactcccaca
     6421 tatgccagat tcagcaacgg atacggcttc cccaacttgc ccacttccat acgtgtcctc
     6481 cttaccagaa atttatcctt aagatcccga atcgtttaaa ctcgactctg gctctatcga
     6541 atctccgtcg tttcgagctt acgcgaacag ccgtggcgct catttgctcg tcgggcatcg
     6601 aatctcgtca gctatcgtca gcttaccttt ttggcagcga tcgcggctcc cgacatcttg
     6661 gaccattagc tccacaggta tcttcttccc tctagtggtc ataacagcag cttcagctac
     6721 ctctcaattc aaaaaacccc tcaagacccg tttagaggcc ccaaggggtt atgctatcaa
     6781 tcgttgcgtt acacacacaa aaaaccaaca cacatccatc ttcgatggat agcgatttta
     6841 ttatctaact gctgatcgag tgtagccaga tctagtaatc aattacgggg tcattagttc
     6901 atagcccata tatggagttc cgcgttacat aacttacggt aaatggcccg cctggctgac
     6961 cgcccaacga cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa
     7021 tagggacttt ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag
     7081 tacatcaagt gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc
     7141 ccgcctggca ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct
     7201 acgtattagt catcgctatt accatgctga tgcggttttg gcagtacatc aatgggcgtg
     7261 gatagcggtt tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt
     7321 tgttttggca ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga
     7381 cgcaaatggg cggtaggcgt gtacggtggg aggtctatat aagcagagct ggtttagtga
     7441 accgtcagat cagatctttg tcgatcctac catccactcg acacacccgc cagcggccgc
//
