LOCUS       sequence-295840-        7396 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression plasmid for SARS-CoV-2 2'-O-ribose 
            methyltransferase.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7396)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 7396)
  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-m4133325
FEATURES             Location/Qualifiers
     source          1..7396
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             902..922
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             1271..1288
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     polyA_signal    1365..1486
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1376..1395)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1430..1449
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      1601..1647
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1677..1813
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1858..1929
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(2155..2743)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2235..2254)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        3243..4452
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     4289..4308
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             4456..5055
                     /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(4456..4475)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4952..4972
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    5082..5130
                     /note="synthetic polyadenylation signal"
     CDS             join(5396..5464,5465..6256)
                     /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(5614..5633)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      6269..6405
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    6550..6621
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(6626..6672)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    6690..6738
                     /note="synthetic polyadenylation signal"
     enhancer        6819..7122
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        7124..7326
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     7277..7297
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
ORIGIN
        1 caccatgtcc tcacaagcgt ggcagccagg cgtagcaatg cctaatttat acaagatgca
       61 gcggatgctc cttgaaaaat gcgacctaca gaactatgga gatagtgcta cattgcccaa
      121 ggggataatg atgaatgtgg ccaaatatac ccaactgtgt cagtacctga acactctcac
      181 gctggccgtt ccgtacaaca tgagggtgat tcattttggt gccggcagcg acaagggagt
      241 cgctcccggg accgctgtgc tgcgacagtg gcttcctaca ggcactctgc tagtggattc
      301 tgatttgaat gacttcgtca gcgacgcaga ctcgaccctc atcggagatt gtgcgactgt
      361 tcacacagcc aacaaatggg acctgatcat ttccgatatg tatgatccaa agaccaaaaa
      421 tgtcacgaag gagaacgact caaaggaggg tttctttacg tacatctgcg ggttcatcca
      481 acagaaactt gcactgggcg gaagtgtggc aattaaaata acagaacact cttggaacgc
      541 cgacctatac aagcttatgg gtcattttgc ttggtggacc gcctttgtta ctaatgtaaa
      601 cgcttcaagc tccgaggcct tcctgatagg ttgtaattat ttaggcaagc ctagagagca
      661 gattgatggg tatgtgatgc acgcaaatta catcttctgg cgcaacacaa atccaattca
      721 gctcagttct tattcactgt ttgatatgag caaattcccc cttaaactga gaggcactgc
      781 cgtcatgtcc ttgaaggaag gacaaatcaa cgatatgatc ttgtctctcc ttagcaaggg
      841 gcgtttgatt attagggaga acaatcgggt ggtgatctca tcggacgttc tggtcaacaa
      901 tgaaaacctt tactttcagt cgatgagtga caaaatcatt catctaaccg atgactcctt
      961 tgatacagac gtgttgaagg ccgatggtgc gatactggta gatttctggg ctgaatggtg
     1021 tggcccgtgc aagatgatcg cacccattct ggacgagatc gccgacgagt atcaaggaaa
     1081 attaactgtc gctaagctta atatagatca gaacccaggg acggcaccta aatacggcat
     1141 caggggaata cctaccctcc tgctctttaa gaatggggaa gttgccgcga caaaagtggg
     1201 agccctgtct aagggccagc taaaagagtt cttggacgct aacctggcag gttcaggctc
     1261 ggggcacatg caccatcacc atcaccatag cagcgggctc gtgccacgcg gctaagctag
     1321 cttgactgac tgagatacag cgtaccttca gctcacagac atgataagat acattgatga
     1381 gtttggacaa accacaacta gaatgcagtg aaaaaaatgc tttatttgtg aaatttgtga
     1441 tgctattgct ttatttgtaa ccattataag ctgcaataaa caagttaaca acaacaattg
     1501 cattcatttt atgtttcagg ttcaggggga ggtgtgggag gttttttaaa gcaagtaaaa
     1561 cctctacaaa tgtggtattg gcccatctct atcggtatcg tagcataacc ccttggggcc
     1621 tctaaacggg tcttgagggg ttttttgtgc ccctcgggcc ggattgctat ctaccggcat
     1681 tggcgcagaa aaaaatgcct gatgcgacgc tgcgcgtctt atactcccac atatgccaga
     1741 ttcagcaacg gatacggctt ccccaacttg cccacttcca tacgtgtcct ccttaccaga
     1801 aatttatcct taaggtcgtc agctatcctg caggcgatct ctcgatttcg atcaagacat
     1861 tcctttaatg gtcttttctg gacaccacta ggggtcagaa gtagttcatc aaactttctt
     1921 ccctccctaa tctcattggt taccttgggc tatcgaaact taattaacca gtcaagtcag
     1981 ctacttggcg agatcgactt gtctgggttt cgactacgct cagaattgcg tcagtcaagt
     2041 tcgatctggt ccttgctatt gcacccgttc tccgattacg agtttcattt aaatcatgtg
     2101 agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca
     2161 taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa
     2221 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc
     2281 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc
     2341 gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct
     2401 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg
     2461 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag
     2521 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta
     2581 cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg
     2641 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt
     2701 tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt
     2761 ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag
     2821 attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat
     2881 ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc
     2941 tatctcagcg atctgtctat ttcgttcatc catagttgca tttaaatttc cgaactctcc
     3001 aaggccctcg tcggaaaatc ttcaaacctt tcgtccgatc catcttgcag gctacctctc
     3061 gaacgaacta tcgcaagtct cttggccggc cttgcgcctt ggctattgct tggcagcgcc
     3121 tatcgccagg tattactcca atcccgaata tccgagatcg ggatcacccg agagaagttc
     3181 aacctacatc ctcaatcccg atctatccga gatccgagga atatcgaaat cggggcgcgc
     3241 ctggcctccg cgccgggttt tggcgcctcc cgcgggcgcc cccctcgtca cggcgagcgc
     3301 tgccacgtca gacgaagggc gcaggagcgt cctgatcctt ccgcccggac gctcaggaca
     3361 gcggcccgct gctcataaga ctcggcctta gaaccccagt atcagcagaa ggacatttta
     3421 ggacgggact tgggtgactc tagggcactg gttttctttc cagagagcgg aacaggcgag
     3481 gaaaagtagt cccttctcgg cgattctgcg gagggatctc cgtggggcgg tgaacgccga
     3541 tgattatata aggacgcgcc gggtgtggca cagctagttc cgtcgcagcc gggatttggg
     3601 tcgcggttct tgtttgtgga tcgctgtgat cgtcacttgg tgagtagcgg gctgctgggc
     3661 tggccggggc tttcgtggcc gccgggccgc tcggtgggac ggaagcgtgt ggagagaccg
     3721 ccaagggctg tagtctgggt ccgcgagcaa ggttgccctg aactgggggt tggggggagc
     3781 gcagcaaaat ggcggctgtt cccgagtctt gaatggaaga cgcttgtgag gcgggctgtg
     3841 aggtcgttga aacaaggtgg ggggcatggt gggcggcaag aacccaaggt cttgagccct
     3901 tcgctaatgc gggaaagctc ttattcgggt gagatgggct gggcaccatc tggggaccct
     3961 gacgtgaagt ttgtcactga ctggagaact cggtttgtcg tctgttgcgg gggcggcagt
     4021 tatggcggtg ccgttgggca gtgcacccgt acctttggga gcgcgcgccc tcgtcgtgtc
     4081 gtgacgtcac ccgttctgtt ggcttataat gcagggtggg gccacctgcc ggtaggtgtg
     4141 cggtaggctt ttctccgtcg caggacgcag ggttcgggcc tagggtaggc tctcctgaat
     4201 cgacaggcgc cggacctctg gtgaggggag ggataagtga ggcgtcagtt tctttggtcg
     4261 gttttatgta cctatcttct taagtagctg aagctccggt tttgaactat gcgctcgggg
     4321 ttggcgagtg tgttttgtga agttttttag gcaccttttg aaatgtaatc atttgggtca
     4381 atatgtaatt ttcagtgtta gacttgtaaa ttgtccgcta aattctggcc gtttttggct
     4441 tttttgttag acaacatgac cgagtacaag cccacggtgc gcctcgccac ccgcgacgac
     4501 gtccccaggg ccgtacgcac cctcgccgcc gcgttcgccg actaccccgc cacgcgccac
     4561 accgtcgatc cggaccgcca catcgagcgg gtcaccgagc tgcaagaact cttcctcacg
     4621 cgcgtcgggc tcgacatcgg caaggtgtgg gtcgcggacg acggcgccgc ggtggcggtc
     4681 tggaccacgc cggagagcgt cgaagcgggg gcggtgttcg ccgagatcgg cccgcgcatg
     4741 gccgagttga gcggttcccg gctggccgcg cagcaacaga tggagggcct cctggcgccg
     4801 caccggccca aggagcccgc gtggttcctg gccaccgtcg gcgtctcgcc cgaccaccag
     4861 ggcaagggtc tgggcagcgc cgtcgtgctc cccggagtgg aggccgccga gcgcgccggg
     4921 gtgcccgcct tcctggagac ctccgcgccc cgcaacctcc ccttctacga gcggctcggc
     4981 ttcaccgtca ccgccgacgt cgaggtgccc gaaggaccgc gcacctggtg catgacccgc
     5041 aagcccggtg cctaggctag tatgtaagcc tagtcttaga taataaaatc gctatccatc
     5101 gaagatggat gtgtgttggt tttttgtgtg tgtaacgcta ggcgcgcctg gtgtaccgag
     5161 aacgatcctc tcagtgcgag tctcgacgat ccatatcgtt gcttggcagt cagccagtcg
     5221 gaatccagct tgggacccag gaagtccaat cgtcagatat tgtactcaag cctggtcacg
     5281 gcagcgtacc gatctgttta aacctagata ttgatagtct gatcggtcaa cgtataatcg
     5341 agtcctagct tttgcaaaca tctatcaaga gacaggatca gcaggaggct ttcgcatgag
     5401 tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt
     5461 tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcgcgagt
     5521 gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga
     5581 acgctttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat
     5641 tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga
     5701 gtattcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag
     5761 tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa cgattggagg
     5821 accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg
     5881 ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt
     5941 agcaatggca acaaccttgc gtaaactatt aactggcgaa ctacttactc tagcttcccg
     6001 gcaacagttg atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc
     6061 ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg
     6121 tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac
     6181 ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact
     6241 gattaagcat tggtaaccga ttctaggtgc attggcgcag aaaaaaatgc ctgatgcgac
     6301 gctgcgcgtc ttatactccc acatatgcca gattcagcaa cggatacggc ttccccaact
     6361 tgcccacttc catacgtgtc ctccttacca gaaatttatc cttaagatcg tttaaactcg
     6421 actctggctc tatcgaatct ccgtcgtttc gagcttacgc gaacagccgt ggcgctcatt
     6481 tgctcgtcgg gcatcgaatc tcgtcagcta tcgtcagctt acctttttgg cagcgatcgc
     6541 ggctcccgac atcttggacc attagctcca caggtatctt cttccctcta gtggtcataa
     6601 cagcagcttc agctacctct caattcaaaa aacccctcaa gacccgttta gaggccccaa
     6661 ggggttatgc tatcaatcgt tgcgttacac acacaaaaaa ccaacacaca tccatcttcg
     6721 atggatagcg attttattat ctaactgctg atcgagtgta gccagatcta gtaatcaatt
     6781 acggggtcat tagttcatag cccatatatg gagttccgcg ttacataact tacggtaaat
     6841 ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt
     6901 cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa
     6961 actgcccact tggcagtaca tcaagtgtat catatgccaa gtacgccccc tattgacgtc
     7021 aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttatg ggactttcct
     7081 acttggcagt acatctacgt attagtcatc gctattacca tgctgatgcg gttttggcag
     7141 tacatcaatg ggcgtggata gcggtttgac tcacggggat ttccaagtct ccaccccatt
     7201 gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg actttccaaa atgtcgtaac
     7261 aactccgccc cattgacgca aatgggcggt aggcgtgtac ggtgggaggt ctatataagc
     7321 agagctggtt tagtgaaccg tcagatcaga tctttgtcga tcctaccatc cactcgacac
     7381 acccgccagc ggccgc
//
