LOCUS       sequence-294994-        9298 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 9298)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 9298)
  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-m4133167
FEATURES             Location/Qualifiers
     source          1..9298
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             2804..2824
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             3173..3190
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     polyA_signal    3267..3388
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3278..3297)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3332..3351
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      3503..3549
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      3579..3715
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    3760..3831
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(4057..4645)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4137..4156)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        5145..6354
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     6191..6210
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             6358..6957
                     /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(6358..6377)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     6854..6874
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    6984..7032
                     /note="synthetic polyadenylation signal"
     CDS             join(7298..7366,7367..8158)
                     /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(7516..7535)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      8171..8307
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    8452..8523
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(8528..8574)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    8592..8640
                     /note="synthetic polyadenylation signal"
     enhancer        8721..9024
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        9026..9228
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     9179..9199
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
ORIGIN
        1 caccatgtct gccgacgcac agtcgttcct gaatagagtt tgtggtgtat ccgcggcccg
       61 gctaactcca tgcggaaccg ggacgagcac agatgtggtg taccgagctt ttgatatcta
      121 taacgacaaa gtcgctggct tcgccaagtt tcttaagacc aactgttgca ggttccaaga
      181 gaaagacgaa gatgataatc tcatagactc atactttgtg gttaagcgtc acacttttag
      241 taattatcag catgaggaaa caatttataa cttgctgaag gactgccctg ccgtagcaaa
      301 acacgacttc tttaaattcc gcatcgatgg cgatatggtc ccgcatatta gcagacagcg
      361 gttaaccaag tacactatgg cagacctggt gtacgccctg cgacacttcg acgagggaaa
      421 ctgtgacacg ttaaaagaaa tcctcgtgac ttataattgt tgcgatgacg attatttcaa
      481 caagaaagat tggtacgatt ttgtcgagaa tcccgatata ctcagagtgt acgctaacct
      541 gggggaaagg gttcgccagg cactattgaa gacagtccaa ttttgcgacg cgatgcggaa
      601 tgccggcatt gtgggtgtcc tgacacttga taaccaggac ctgaatggca actggtatga
      661 ctttggagat ttcattcaga ccacgcccgg gtccggagta cctgtcgttg actcttacta
      721 ctcacttctg atgccaatcc taaccttaac ccgagctctc acagctgaga gtcacgtgga
      781 caccgacctc acaaaaccgt atatcaagtg ggatttgctg aagtacgatt tcacggaaga
      841 gaggttaaag ctctttgatc gttatttcaa atactgggac cagacttatc atcccaactg
      901 tgtgaattgt ctggacgaca gatgcatact tcattgtgcc aatttcaatg tactgtttag
      961 cacagtgttc ccaccaactt cgtttggtcc tctagtgcgc aaaatctttg tggatggggt
     1021 gcccttcgtt gtttccacag gctatcactt tcgtgagctc ggcgtagttc acaaccaaga
     1081 tgtcaacctg cattcttcca ggttaagttt taaggaactt ctggtgtatg ccgcagaccc
     1141 tgcgatgcac gctgcaagcg gtaatttgtt gctggataag cggaccacgt gcttcagcgt
     1201 ggcggcttta acaaataacg ttgccttcca aaccgtcaag cctggcaact tcaataaaga
     1261 tttctacgat ttcgccgtca gtaaagggtt ttttaaggag ggatcaagcg tagaattgaa
     1321 acattttttc tttgctcagg acggtaatgc cgctatctcg gactacgact actaccggta
     1381 taacctacca actatgtgcg acattcggca gctgttgttc gtggtggagg tcgttgataa
     1441 atactttgat tgctacgatg gcggatgcat taacgccaat caagtcatcg tgaacaatct
     1501 ggacaagagc gccggcttcc cctttaacaa atggggaaag gcaaggctat attatgactc
     1561 tatgtcctat gaggatcagg acgctctctt cgcgtatacc aagcgcaacg ttatccccac
     1621 aattacccag atgaacctca aatatgcaat cagcgcaaag aacagagcga ggacagttgc
     1681 tggggtgagt atatgttcga caatgactaa caggcagttt caccagaagc tgttgaaatc
     1741 tattgccgct acacgagggg ctaccgtcgt gattggcacc tccaaattct acggagggtg
     1801 gcataatatg ttaaaaactg tgtactccga cgtggaaaac cctcacctga tgggttggga
     1861 ttatcctaag tgcgaccggg cgatgccaaa catgctgaga attatggcgt cattagtcct
     1921 ggcgcgaaag cacaccacgt gctgtagtct gtcacatcgc ttttacagat tggctaatga
     1981 atgcgcccaa gtgcttagcg agatggtgat gtgtgggggt tcactttacg tgaagcccgg
     2041 aggaacgtcc tcaggcgacg ccacaactgc ctacgcaaat tcagtattca acatatgtca
     2101 ggccgttaca gcaaacgtga atgctctgct gtctactgat gggaacaaga tcgccgataa
     2161 gtacgtccgg aacctgcaac acaggctgta cgagtgtctg tatcgtaatc gagatgtaga
     2221 caccgacttc gtgaacgaat tttacgcata cttacggaaa catttctcca tgatgatcct
     2281 ctccgacgat gcagttgtgt gctttaattc aacttacgcc tctcagggct tggtggccag
     2341 tatcaaaaat tttaagtcgg ttctttatta tcagaataac gtgtttatga gcgaggctaa
     2401 gtgttggacc gagactgatc tgaccaaagg cccgcacgag ttctgttccc agcacaccat
     2461 gctggtcaag cagggtgatg actatgtcta tttaccatac ccagatccga gtaggatctt
     2521 gggagccggt tgttttgtgg atgacattgt gaagactgac ggtacactca tgattgagcg
     2581 cttcgtcagt ttggcaatcg acgcttaccc ccttaccaaa catccaaatc aggaatatgc
     2641 ggatgtgttt cacctgtacc ttcaatacat aaggaaactt catgatgaac taactggaca
     2701 tatgctcgac atgtattctg taatgcttac aaatgacaat acctcacggt actgggaacc
     2761 ggagttttat gaagccatgt acaccccaca tactgtcctg caagaaaacc tttactttca
     2821 gtcgatgagt gacaaaatca ttcatctaac cgatgactcc tttgatacag acgtgttgaa
     2881 ggccgatggt gcgatactgg tagatttctg ggctgaatgg tgtggcccgt gcaagatgat
     2941 cgcacccatt ctggacgaga tcgccgacga gtatcaagga aaattaactg tcgctaagct
     3001 taatatagat cagaacccag ggacggcacc taaatacggc atcaggggaa tacctaccct
     3061 cctgctcttt aagaatgggg aagttgccgc gacaaaagtg ggagccctgt ctaagggcca
     3121 gctaaaagag ttcttggacg ctaacctggc aggttcaggc tcggggcaca tgcaccatca
     3181 ccatcaccat agcagcgggc tcgtgccacg cggctaagct agcttgactg actgagatac
     3241 agcgtacctt cagctcacag acatgataag atacattgat gagtttggac aaaccacaac
     3301 tagaatgcag tgaaaaaaat gctttatttg tgaaatttgt gatgctattg ctttatttgt
     3361 aaccattata agctgcaata aacaagttaa caacaacaat tgcattcatt ttatgtttca
     3421 ggttcagggg gaggtgtggg aggtttttta aagcaagtaa aacctctaca aatgtggtat
     3481 tggcccatct ctatcggtat cgtagcataa ccccttgggg cctctaaacg ggtcttgagg
     3541 ggttttttgt gcccctcggg ccggattgct atctaccggc attggcgcag aaaaaaatgc
     3601 ctgatgcgac gctgcgcgtc ttatactccc acatatgcca gattcagcaa cggatacggc
     3661 ttccccaact tgcccacttc catacgtgtc ctccttacca gaaatttatc cttaaggtcg
     3721 tcagctatcc tgcaggcgat ctctcgattt cgatcaagac attcctttaa tggtcttttc
     3781 tggacaccac taggggtcag aagtagttca tcaaactttc ttccctccct aatctcattg
     3841 gttaccttgg gctatcgaaa cttaattaac cagtcaagtc agctacttgg cgagatcgac
     3901 ttgtctgggt ttcgactacg ctcagaattg cgtcagtcaa gttcgatctg gtccttgcta
     3961 ttgcacccgt tctccgatta cgagtttcat ttaaatcatg tgagcaaaag gccagcaaaa
     4021 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     4081 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     4141 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     4201 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     4261 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     4321 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     4381 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     4441 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac
     4501 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     4561 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     4621 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     4681 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
     4741 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
     4801 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     4861 atttcgttca tccatagttg catttaaatt tccgaactct ccaaggccct cgtcggaaaa
     4921 tcttcaaacc tttcgtccga tccatcttgc aggctacctc tcgaacgaac tatcgcaagt
     4981 ctcttggccg gccttgcgcc ttggctattg cttggcagcg cctatcgcca ggtattactc
     5041 caatcccgaa tatccgagat cgggatcacc cgagagaagt tcaacctaca tcctcaatcc
     5101 cgatctatcc gagatccgag gaatatcgaa atcggggcgc gcctggcctc cgcgccgggt
     5161 tttggcgcct cccgcgggcg cccccctcgt cacggcgagc gctgccacgt cagacgaagg
     5221 gcgcaggagc gtcctgatcc ttccgcccgg acgctcagga cagcggcccg ctgctcataa
     5281 gactcggcct tagaacccca gtatcagcag aaggacattt taggacggga cttgggtgac
     5341 tctagggcac tggttttctt tccagagagc ggaacaggcg aggaaaagta gtcccttctc
     5401 ggcgattctg cggagggatc tccgtggggc ggtgaacgcc gatgattata taaggacgcg
     5461 ccgggtgtgg cacagctagt tccgtcgcag ccgggatttg ggtcgcggtt cttgtttgtg
     5521 gatcgctgtg atcgtcactt ggtgagtagc gggctgctgg gctggccggg gctttcgtgg
     5581 ccgccgggcc gctcggtggg acggaagcgt gtggagagac cgccaagggc tgtagtctgg
     5641 gtccgcgagc aaggttgccc tgaactgggg gttgggggga gcgcagcaaa atggcggctg
     5701 ttcccgagtc ttgaatggaa gacgcttgtg aggcgggctg tgaggtcgtt gaaacaaggt
     5761 ggggggcatg gtgggcggca agaacccaag gtcttgagcc cttcgctaat gcgggaaagc
     5821 tcttattcgg gtgagatggg ctgggcacca tctggggacc ctgacgtgaa gtttgtcact
     5881 gactggagaa ctcggtttgt cgtctgttgc gggggcggca gttatggcgg tgccgttggg
     5941 cagtgcaccc gtacctttgg gagcgcgcgc cctcgtcgtg tcgtgacgtc acccgttctg
     6001 ttggcttata atgcagggtg gggccacctg ccggtaggtg tgcggtaggc ttttctccgt
     6061 cgcaggacgc agggttcggg cctagggtag gctctcctga atcgacaggc gccggacctc
     6121 tggtgagggg agggataagt gaggcgtcag tttctttggt cggttttatg tacctatctt
     6181 cttaagtagc tgaagctccg gttttgaact atgcgctcgg ggttggcgag tgtgttttgt
     6241 gaagtttttt aggcaccttt tgaaatgtaa tcatttgggt caatatgtaa ttttcagtgt
     6301 tagacttgta aattgtccgc taaattctgg ccgtttttgg cttttttgtt agacaacatg
     6361 accgagtaca agcccacggt gcgcctcgcc acccgcgacg acgtccccag ggccgtacgc
     6421 accctcgccg ccgcgttcgc cgactacccc gccacgcgcc acaccgtcga tccggaccgc
     6481 cacatcgagc gggtcaccga gctgcaagaa ctcttcctca cgcgcgtcgg gctcgacatc
     6541 ggcaaggtgt gggtcgcgga cgacggcgcc gcggtggcgg tctggaccac gccggagagc
     6601 gtcgaagcgg gggcggtgtt cgccgagatc ggcccgcgca tggccgagtt gagcggttcc
     6661 cggctggccg cgcagcaaca gatggagggc ctcctggcgc cgcaccggcc caaggagccc
     6721 gcgtggttcc tggccaccgt cggcgtctcg cccgaccacc agggcaaggg tctgggcagc
     6781 gccgtcgtgc tccccggagt ggaggccgcc gagcgcgccg gggtgcccgc cttcctggag
     6841 acctccgcgc cccgcaacct ccccttctac gagcggctcg gcttcaccgt caccgccgac
     6901 gtcgaggtgc ccgaaggacc gcgcacctgg tgcatgaccc gcaagcccgg tgcctaggct
     6961 agtatgtaag cctagtctta gataataaaa tcgctatcca tcgaagatgg atgtgtgttg
     7021 gttttttgtg tgtgtaacgc taggcgcgcc tggtgtaccg agaacgatcc tctcagtgcg
     7081 agtctcgacg atccatatcg ttgcttggca gtcagccagt cggaatccag cttgggaccc
     7141 aggaagtcca atcgtcagat attgtactca agcctggtca cggcagcgta ccgatctgtt
     7201 taaacctaga tattgatagt ctgatcggtc aacgtataat cgagtcctag cttttgcaaa
     7261 catctatcaa gagacaggat cagcaggagg ctttcgcatg agtattcaac atttccgtgt
     7321 cgcccttatt cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct
     7381 ggtgaaagta aaagatgctg aagatcagtt gggtgcgcga gtgggttaca tcgaactgga
     7441 tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa gaacgctttc caatgatgag
     7501 cacttttaaa gttctgctat gtggcgcggt attatcccgt attgacgccg ggcaagagca
     7561 actcggtcgc cgcatacact attctcagaa tgacttggtt gagtattcac cagtcacaga
     7621 aaagcatctt acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag
     7681 tgataacact gcggccaact tacttctgac aacgattgga ggaccgaagg agctaaccgc
     7741 ttttttgcac aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa
     7801 tgaagccata ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg caacaacctt
     7861 gcgtaaacta ttaactggcg aactacttac tctagcttcc cggcaacagt tgatagactg
     7921 gatggaggcg gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt
     7981 tattgctgat aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg
     8041 gccagatggt aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat
     8101 ggatgaacga aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaacc
     8161 gattctaggt gcattggcgc agaaaaaaat gcctgatgcg acgctgcgcg tcttatactc
     8221 ccacatatgc cagattcagc aacggatacg gcttccccaa cttgcccact tccatacgtg
     8281 tcctccttac cagaaattta tccttaagat cgtttaaact cgactctggc tctatcgaat
     8341 ctccgtcgtt tcgagcttac gcgaacagcc gtggcgctca tttgctcgtc gggcatcgaa
     8401 tctcgtcagc tatcgtcagc ttaccttttt ggcagcgatc gcggctcccg acatcttgga
     8461 ccattagctc cacaggtatc ttcttccctc tagtggtcat aacagcagct tcagctacct
     8521 ctcaattcaa aaaacccctc aagacccgtt tagaggcccc aaggggttat gctatcaatc
     8581 gttgcgttac acacacaaaa aaccaacaca catccatctt cgatggatag cgattttatt
     8641 atctaactgc tgatcgagtg tagccagatc tagtaatcaa ttacggggtc attagttcat
     8701 agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg
     8761 cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata
     8821 gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta
     8881 catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc
     8941 gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca gtacatctac
     9001 gtattagtca tcgctattac catgctgatg cggttttggc agtacatcaa tgggcgtgga
     9061 tagcggtttg actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg
     9121 ttttggcacc aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg
     9181 caaatgggcg gtaggcgtgt acggtgggag gtctatataa gcagagctgg tttagtgaac
     9241 cgtcagatca gatctttgtc gatcctacca tccactcgac acacccgcca gcggccgc
//
