LOCUS       sequence-295343-        8947 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression plasmid for SARS-CoV-2 RNA-dependent RNA 
            polymerase.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8947)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 8947)
  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-m4134209
FEATURES             Location/Qualifiers
     source          1..8947
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             2798..2818
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             2819..2863
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     polyA_signal    2916..3037
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2927..2946)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2981..3000
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      3152..3198
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      3228..3364
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    3409..3480
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(3706..4294)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3786..3805)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        4794..6003
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     5840..5859
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             6007..6606
                     /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(6007..6026)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     6503..6523
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    6633..6681
                     /note="synthetic polyadenylation signal"
     CDS             join(6947..7015,7016..7807)
                     /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(7165..7184)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      7820..7956
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    8101..8172
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(8177..8223)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    8241..8289
                     /note="synthetic polyadenylation signal"
     enhancer        8370..8673
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        8675..8877
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     8828..8848
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      join(8946..8947,1..8)
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
ORIGIN
        1 caccatggct gatgcacaat cgtttttaaa ccgggtttgc ggtgtaagtg cagcccgtct
       61 tacaccgtgc ggcacaggca ctagtactga tgtcgtatac agggcttttg acatctacaa
      121 tgataaagta gctggttttg ctaaattcct aaaaactaat tgttgtcgct tccaagaaaa
      181 ggacgaagat gacaatttaa ttgattctta ctttgtagtt aagagacaca ctttctctaa
      241 ctaccaacat gaagaaacaa tttataattt acttaaggat tgtccagctg ttgctaaaca
      301 tgacttcttt aagtttagaa tagacggtga catggtacca catatatcac gtcaacgtct
      361 tactaaatac acaatggcag acctcgtcta tgctttaagg cattttgatg aaggtaattg
      421 tgacacatta aaagaaatac ttgtcacata caattgttgt gatgatgatt atttcaataa
      481 aaaggactgg tatgattttg tagaaaaccc agatatatta cgcgtatacg ccaacttagg
      541 tgaacgtgta cgccaagctt tgttaaaaac agtacaattc tgtgatgcca tgcgaaatgc
      601 tggtattgtt ggtgtactga cattagataa tcaagatctc aatggtaact ggtatgattt
      661 cggtgatttc atacaaacca cgccaggtag tggagttcct gttgtagatt cttattattc
      721 attgttaatg cctatattaa ccttgaccag ggctttaact gcagagtcac atgttgacac
      781 tgacttaaca aagccttaca ttaagtggga tttgttaaaa tatgacttca cggaagagag
      841 gttaaaactc tttgaccgtt attttaaata ttgggatcag acataccacc caaattgtgt
      901 taactgtttg gatgacagat gcattctgca ttgtgcaaac tttaatgttt tattctctac
      961 agtgttccca cctacaagtt ttggaccact agtgagaaaa atatttgttg atggtgttcc
     1021 atttgtagtt tcaactggat accacttcag agagctaggt gttgtacata atcaggatgt
     1081 aaacttacat agctctagac ttagttttaa ggaattactt gtgtatgctg ctgaccctgc
     1141 tatgcacgct gcttctggta atctattact agataaacgc actacgtgct tttcagtagc
     1201 tgcacttact aacaatgttg cttttcaaac tgtcaaaccc ggtaatttta acaaagactt
     1261 ctatgacttt gctgtgtcta agggtttctt taaggaagga agttctgttg aattaaaaca
     1321 cttcttcttt gctcaggatg gtaatgctgc tatcagcgat tatgactact atcgttataa
     1381 tctaccaaca atgtgtgata tcagacaact actatttgta gttgaagttg ttgataagta
     1441 ctttgattgt tacgatggtg gctgtattaa tgctaaccaa gtcatcgtca acaacctaga
     1501 caaatcagct ggttttccat ttaataaatg gggtaaggct agactttatt atgattcaat
     1561 gagttatgag gatcaagatg cacttttcgc atatacaaaa cgtaatgtca tccctactat
     1621 aactcaaatg aatcttaagt atgccattag tgcaaagaat agagctcgca ccgtagctgg
     1681 tgtctctatc tgtagtacta tgaccaatag acagtttcat caaaaattat tgaaatcaat
     1741 agccgccact agaggagcta ctgtagtaat tggaacaagc aaattctatg gtggttggca
     1801 caacatgtta aaaactgttt atagtgatgt agaaaaccct caccttatgg gttgggatta
     1861 tcctaaatgt gatagagcca tgcctaacat gcttagaatt atggcctcac ttgttcttgc
     1921 tcgcaaacat acaacgtgtt gtagcttgtc acaccgtttc tatagattag ctaatgagtg
     1981 tgctcaagta ttgagtgaaa tggtcatgtg tggcggttca ctatatgtta aaccaggtgg
     2041 aacctcatca ggagatgcca caactgctta tgctaatagt gtttttaaca tttgtcaagc
     2101 tgtcacggcc aatgttaatg cacttttatc tactgatggt aacaaaattg ccgataagta
     2161 tgtccgcaat ttacaacaca gactttatga gtgtctctat agaaatagag atgttgacac
     2221 agactttgtg aatgagtttt acgcatattt gcgtaaacat ttctcaatga tgatactctc
     2281 tgacgatgct gttgtgtgtt tcaatagcac ttatgcatct caaggtctag tggctagcat
     2341 aaagaacttt aagtcagttc tttattatca aaacaatgtt tttatgtctg aagcaaaatg
     2401 ttggactgag actgacctta ctaaaggacc tcatgaattt tgctctcaac atacaatgct
     2461 agttaaacag ggtgatgatt atgtgtacct tccttaccca gatccatcaa gaatcctagg
     2521 ggccggctgt tttgtagatg atatcgtaaa aacagatggt acacttatga ttgaacggtt
     2581 cgtgtcttta gctatagatg cttacccact tactaaacat cctaatcagg agtatgctga
     2641 tgtctttcat ttgtacttac aatacataag aaagctacat gatgagttaa caggacacat
     2701 gttagacatg tattctgtta tgcttactaa tgataacact tcaaggtatt gggaacctga
     2761 gttttatgag gctatgtaca caccgcatac agtcttagaa aacctttact ttcagtcgaa
     2821 agaaaccgcg gcagctaagt tcgagaggca gcacatggac agttaagcta gcttgactga
     2881 ctgagataca gcgtaccttc agctcacaga catgataaga tacattgatg agtttggaca
     2941 aaccacaact agaatgcagt gaaaaaaatg ctttatttgt gaaatttgtg atgctattgc
     3001 tttatttgta accattataa gctgcaataa acaagttaac aacaacaatt gcattcattt
     3061 tatgtttcag gttcaggggg aggtgtggga ggttttttaa agcaagtaaa acctctacaa
     3121 atgtggtatt ggcccatctc tatcggtatc gtagcataac cccttggggc ctctaaacgg
     3181 gtcttgaggg gttttttgtg cccctcgggc cggattgcta tctaccggca ttggcgcaga
     3241 aaaaaatgcc tgatgcgacg ctgcgcgtct tatactccca catatgccag attcagcaac
     3301 ggatacggct tccccaactt gcccacttcc atacgtgtcc tccttaccag aaatttatcc
     3361 ttaaggtcgt cagctatcct gcaggcgatc tctcgatttc gatcaagaca ttcctttaat
     3421 ggtcttttct ggacaccact aggggtcaga agtagttcat caaactttct tccctcccta
     3481 atctcattgg ttaccttggg ctatcgaaac ttaattaacc agtcaagtca gctacttggc
     3541 gagatcgact tgtctgggtt tcgactacgc tcagaattgc gtcagtcaag ttcgatctgg
     3601 tccttgctat tgcacccgtt ctccgattac gagtttcatt taaatcatgt gagcaaaagg
     3661 ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg
     3721 cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg
     3781 actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac
     3841 cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca
     3901 tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt
     3961 gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc
     4021 caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag
     4081 agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac
     4141 tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt
     4201 tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa
     4261 gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg
     4321 gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa
     4381 aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat
     4441 atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc
     4501 gatctgtcta tttcgttcat ccatagttgc atttaaattt ccgaactctc caaggccctc
     4561 gtcggaaaat cttcaaacct ttcgtccgat ccatcttgca ggctacctct cgaacgaact
     4621 atcgcaagtc tcttggccgg ccttgcgcct tggctattgc ttggcagcgc ctatcgccag
     4681 gtattactcc aatcccgaat atccgagatc gggatcaccc gagagaagtt caacctacat
     4741 cctcaatccc gatctatccg agatccgagg aatatcgaaa tcggggcgcg cctggcctcc
     4801 gcgccgggtt ttggcgcctc ccgcgggcgc ccccctcgtc acggcgagcg ctgccacgtc
     4861 agacgaaggg cgcaggagcg tcctgatcct tccgcccgga cgctcaggac agcggcccgc
     4921 tgctcataag actcggcctt agaaccccag tatcagcaga aggacatttt aggacgggac
     4981 ttgggtgact ctagggcact ggttttcttt ccagagagcg gaacaggcga ggaaaagtag
     5041 tcccttctcg gcgattctgc ggagggatct ccgtggggcg gtgaacgccg atgattatat
     5101 aaggacgcgc cgggtgtggc acagctagtt ccgtcgcagc cgggatttgg gtcgcggttc
     5161 ttgtttgtgg atcgctgtga tcgtcacttg gtgagtagcg ggctgctggg ctggccgggg
     5221 ctttcgtggc cgccgggccg ctcggtggga cggaagcgtg tggagagacc gccaagggct
     5281 gtagtctggg tccgcgagca aggttgccct gaactggggg ttggggggag cgcagcaaaa
     5341 tggcggctgt tcccgagtct tgaatggaag acgcttgtga ggcgggctgt gaggtcgttg
     5401 aaacaaggtg gggggcatgg tgggcggcaa gaacccaagg tcttgagccc ttcgctaatg
     5461 cgggaaagct cttattcggg tgagatgggc tgggcaccat ctggggaccc tgacgtgaag
     5521 tttgtcactg actggagaac tcggtttgtc gtctgttgcg ggggcggcag ttatggcggt
     5581 gccgttgggc agtgcacccg tacctttggg agcgcgcgcc ctcgtcgtgt cgtgacgtca
     5641 cccgttctgt tggcttataa tgcagggtgg ggccacctgc cggtaggtgt gcggtaggct
     5701 tttctccgtc gcaggacgca gggttcgggc ctagggtagg ctctcctgaa tcgacaggcg
     5761 ccggacctct ggtgagggga gggataagtg aggcgtcagt ttctttggtc ggttttatgt
     5821 acctatcttc ttaagtagct gaagctccgg ttttgaacta tgcgctcggg gttggcgagt
     5881 gtgttttgtg aagtttttta ggcacctttt gaaatgtaat catttgggtc aatatgtaat
     5941 tttcagtgtt agacttgtaa attgtccgct aaattctggc cgtttttggc ttttttgtta
     6001 gacaacatga ccgagtacaa gcccacggtg cgcctcgcca cccgcgacga cgtccccagg
     6061 gccgtacgca ccctcgccgc cgcgttcgcc gactaccccg ccacgcgcca caccgtcgat
     6121 ccggaccgcc acatcgagcg ggtcaccgag ctgcaagaac tcttcctcac gcgcgtcggg
     6181 ctcgacatcg gcaaggtgtg ggtcgcggac gacggcgccg cggtggcggt ctggaccacg
     6241 ccggagagcg tcgaagcggg ggcggtgttc gccgagatcg gcccgcgcat ggccgagttg
     6301 agcggttccc ggctggccgc gcagcaacag atggagggcc tcctggcgcc gcaccggccc
     6361 aaggagcccg cgtggttcct ggccaccgtc ggcgtctcgc ccgaccacca gggcaagggt
     6421 ctgggcagcg ccgtcgtgct ccccggagtg gaggccgccg agcgcgccgg ggtgcccgcc
     6481 ttcctggaga cctccgcgcc ccgcaacctc cccttctacg agcggctcgg cttcaccgtc
     6541 accgccgacg tcgaggtgcc cgaaggaccg cgcacctggt gcatgacccg caagcccggt
     6601 gcctaggcta gtatgtaagc ctagtcttag ataataaaat cgctatccat cgaagatgga
     6661 tgtgtgttgg ttttttgtgt gtgtaacgct aggcgcgcct ggtgtaccga gaacgatcct
     6721 ctcagtgcga gtctcgacga tccatatcgt tgcttggcag tcagccagtc ggaatccagc
     6781 ttgggaccca ggaagtccaa tcgtcagata ttgtactcaa gcctggtcac ggcagcgtac
     6841 cgatctgttt aaacctagat attgatagtc tgatcggtca acgtataatc gagtcctagc
     6901 ttttgcaaac atctatcaag agacaggatc agcaggaggc tttcgcatga gtattcaaca
     6961 tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt ttgctcaccc
     7021 agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcgcgag tgggttacat
     7081 cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag aacgctttcc
     7141 aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta ttgacgccgg
     7201 gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg agtattcacc
     7261 agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca gtgctgccat
     7321 aaccatgagt gataacactg cggccaactt acttctgaca acgattggag gaccgaagga
     7381 gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc gttgggaacc
     7441 ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg tagcaatggc
     7501 aacaaccttg cgtaaactat taactggcga actacttact ctagcttccc ggcaacagtt
     7561 gatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg cccttccggc
     7621 tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg gtatcattgc
     7681 agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga cggggagtca
     7741 ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac tgattaagca
     7801 ttggtaaccg attctaggtg cattggcgca gaaaaaaatg cctgatgcga cgctgcgcgt
     7861 cttatactcc cacatatgcc agattcagca acggatacgg cttccccaac ttgcccactt
     7921 ccatacgtgt cctccttacc agaaatttat ccttaagatc gtttaaactc gactctggct
     7981 ctatcgaatc tccgtcgttt cgagcttacg cgaacagccg tggcgctcat ttgctcgtcg
     8041 ggcatcgaat ctcgtcagct atcgtcagct tacctttttg gcagcgatcg cggctcccga
     8101 catcttggac cattagctcc acaggtatct tcttccctct agtggtcata acagcagctt
     8161 cagctacctc tcaattcaaa aaacccctca agacccgttt agaggcccca aggggttatg
     8221 ctatcaatcg ttgcgttaca cacacaaaaa accaacacac atccatcttc gatggatagc
     8281 gattttatta tctaactgct gatcgagtgt agccagatct agtaatcaat tacggggtca
     8341 ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct
     8401 ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta
     8461 acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac
     8521 ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt
     8581 aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag
     8641 tacatctacg tattagtcat cgctattacc atgctgatgc ggttttggca gtacatcaat
     8701 gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat tgacgtcaat
     8761 gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa caactccgcc
     8821 ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag cagagctggt
     8881 ttagtgaacc gtcagatcag atctttgtcg atcctaccat ccactcgaca cacccgccag
     8941 cggccgc
//
