LOCUS       sequence-295775-        7520 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 7520)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 7520)
  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-m4133575
FEATURES             Location/Qualifiers
     source          1..7520
                     /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             926..991
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     polyA_signal    1060..1181
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1071..1090)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1125..1144
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      1296..1342
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1372..1508
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1553..1624
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(1850..2438)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1930..1949)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2938..4147
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     3984..4003
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             4151..5179
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     polyA_signal    5206..5254
                     /note="synthetic polyadenylation signal"
     CDS             join(5520..5588,5589..6380)
                     /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(5738..5757)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      6393..6529
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    6674..6745
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(6750..6796)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    6814..6862
                     /note="synthetic polyadenylation signal"
     enhancer        6943..7246
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        7248..7450
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     7401..7421
                     /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 cgatggatta taaggaccac gatggagact acaaagatca
      961 tgacattgat tacaaggacg acgatgacaa ataaggatcc gcaggcctct gctagcttga
     1021 ctgactgaga tacagcgtac cttcagctca cagacatgat aagatacatt gatgagtttg
     1081 gacaaaccac aactagaatg cagtgaaaaa aatgctttat ttgtgaaatt tgtgatgcta
     1141 ttgctttatt tgtaaccatt ataagctgca ataaacaagt taacaacaac aattgcattc
     1201 attttatgtt tcaggttcag ggggaggtgt gggaggtttt ttaaagcaag taaaacctct
     1261 acaaatgtgg tattggccca tctctatcgg tatcgtagca taaccccttg gggcctctaa
     1321 acgggtcttg aggggttttt tgtgcccctc gggccggatt gctatctacc ggcattggcg
     1381 cagaaaaaaa tgcctgatgc gacgctgcgc gtcttatact cccacatatg ccagattcag
     1441 caacggatac ggcttcccca acttgcccac ttccatacgt gtcctcctta ccagaaattt
     1501 atccttaagg tcgtcagcta tcctgcaggc gatctctcga tttcgatcaa gacattcctt
     1561 taatggtctt ttctggacac cactaggggt cagaagtagt tcatcaaact ttcttccctc
     1621 cctaatctca ttggttacct tgggctatcg aaacttaatt aaccagtcaa gtcagctact
     1681 tggcgagatc gacttgtctg ggtttcgact acgctcagaa ttgcgtcagt caagttcgat
     1741 ctggtccttg ctattgcacc cgttctccga ttacgagttt catttaaatc atgtgagcaa
     1801 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
     1861 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
     1921 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
     1981 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
     2041 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
     2101 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
     2161 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
     2221 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
     2281 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
     2341 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
     2401 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
     2461 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
     2521 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
     2581 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
     2641 cagcgatctg tctatttcgt tcatccatag ttgcatttaa atttccgaac tctccaaggc
     2701 cctcgtcgga aaatcttcaa acctttcgtc cgatccatct tgcaggctac ctctcgaacg
     2761 aactatcgca agtctcttgg ccggccttgc gccttggcta ttgcttggca gcgcctatcg
     2821 ccaggtatta ctccaatccc gaatatccga gatcgggatc acccgagaga agttcaacct
     2881 acatcctcaa tcccgatcta tccgagatcc gaggaatatc gaaatcgggg cgcgcctggc
     2941 ctccgcgccg ggttttggcg cctcccgcgg gcgcccccct cgtcacggcg agcgctgcca
     3001 cgtcagacga agggcgcagg agcgtcctga tccttccgcc cggacgctca ggacagcggc
     3061 ccgctgctca taagactcgg ccttagaacc ccagtatcag cagaaggaca ttttaggacg
     3121 ggacttgggt gactctaggg cactggtttt ctttccagag agcggaacag gcgaggaaaa
     3181 gtagtccctt ctcggcgatt ctgcggaggg atctccgtgg ggcggtgaac gccgatgatt
     3241 atataaggac gcgccgggtg tggcacagct agttccgtcg cagccgggat ttgggtcgcg
     3301 gttcttgttt gtggatcgct gtgatcgtca cttggtgagt agcgggctgc tgggctggcc
     3361 ggggctttcg tggccgccgg gccgctcggt gggacggaag cgtgtggaga gaccgccaag
     3421 ggctgtagtc tgggtccgcg agcaaggttg ccctgaactg ggggttgggg ggagcgcagc
     3481 aaaatggcgg ctgttcccga gtcttgaatg gaagacgctt gtgaggcggg ctgtgaggtc
     3541 gttgaaacaa ggtggggggc atggtgggcg gcaagaaccc aaggtcttga gcccttcgct
     3601 aatgcgggaa agctcttatt cgggtgagat gggctgggca ccatctgggg accctgacgt
     3661 gaagtttgtc actgactgga gaactcggtt tgtcgtctgt tgcgggggcg gcagttatgg
     3721 cggtgccgtt gggcagtgca cccgtacctt tgggagcgcg cgccctcgtc gtgtcgtgac
     3781 gtcacccgtt ctgttggctt ataatgcagg gtggggccac ctgccggtag gtgtgcggta
     3841 ggcttttctc cgtcgcagga cgcagggttc gggcctaggg taggctctcc tgaatcgaca
     3901 ggcgccggac ctctggtgag gggagggata agtgaggcgt cagtttcttt ggtcggtttt
     3961 atgtacctat cttcttaagt agctgaagct ccggttttga actatgcgct cggggttggc
     4021 gagtgtgttt tgtgaagttt tttaggcacc ttttgaaatg taatcatttg ggtcaatatg
     4081 taattttcag tgttagactt gtaaattgtc cgctaaattc tggccgtttt tggctttttt
     4141 gttagacaac atgggtaaaa agcctgaact caccgcgacg tctgtcgaga agtttctgat
     4201 cgaaaagttc gacagcgtct ccgacctgat gcagctctcg gagggcgaag aatctcgtgc
     4261 tttcagcttc gatgtaggag ggcgtggata tgtcctgcgg gtaaatagct gcgccgatgg
     4321 tttctacaaa gatcgttatg tttatcggca ctttgcatcc gccgcgctcc cgattccgga
     4381 agtgcttgac attggggagt tcagcgagag cctgacctat tgcatctccc gccgtgcaca
     4441 gggtgtcacg ttgcaagacc tgcctgaaac cgaactgccc gctgttctgc agccggtcgc
     4501 ggaggcaatg gatgccatcg ctgccgccga tcttagccag acgagcgggt tcggcccatt
     4561 cggaccgcaa ggaatcggtc aatacactac atggcgtgat ttcatatgcg cgattgctga
     4621 tccccatgtg tatcactggc aaactgtgat ggacgacacc gtcagtgcgt ccgtcgcgca
     4681 ggctctcgat gagctgatgc tttgggccga ggactgcccc gaagtccggc acctcgtgca
     4741 cgcggatttc ggctccaaca atgtcctgac ggacaatggc cgcataacag cggtcattga
     4801 ctggagcgag gcgatgttcg gggattccca atacgaggtc gccaacatct tcttctggag
     4861 gccgtggttg gcttgtatgg agcagcagac gcgctacttc gagcggaggc atccggagct
     4921 tgcaggatcg ccccggctcc gggcgtatat gctccgcatt ggtcttgacc aactctatca
     4981 gagcttggtt gacggcaatt tcgatgatgc agcttgggcg cagggtcgat gcgacgcaat
     5041 cgtccgatcc ggagccggga ctgtcgggcg tacacaaatc gcccgcagaa gcgccgccgt
     5101 ctggaccgat ggctgtgtag aagtactcgc cgatagtgga aaccgacgcc ccagcactcg
     5161 tccgagggca aaggaataag ctagtatgta agcctagtct tagataataa aatcgctatc
     5221 catcgaagat ggatgtgtgt tggttttttg tgtgtgtaac gctaggcgcg cctggtgtac
     5281 cgagaacgat cctctcagtg cgagtctcga cgatccatat cgttgcttgg cagtcagcca
     5341 gtcggaatcc agcttgggac ccaggaagtc caatcgtcag atattgtact caagcctggt
     5401 cacggcagcg taccgatctg tttaaaccta gatattgata gtctgatcgg tcaacgtata
     5461 atcgagtcct agcttttgca aacatctatc aagagacagg atcagcagga ggctttcgca
     5521 tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg
     5581 tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcgc
     5641 gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg
     5701 aagaacgctt tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc
     5761 gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg
     5821 ttgagtattc accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat
     5881 gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgattg
     5941 gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg
     6001 atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc
     6061 ctgtagcaat ggcaacaacc ttgcgtaaac tattaactgg cgaactactt actctagctt
     6121 cccggcaaca gttgatagac tggatggagg cggataaagt tgcaggacca cttctgcgct
     6181 cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc
     6241 gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca
     6301 cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct
     6361 cactgattaa gcattggtaa ccgattctag gtgcattggc gcagaaaaaa atgcctgatg
     6421 cgacgctgcg cgtcttatac tcccacatat gccagattca gcaacggata cggcttcccc
     6481 aacttgccca cttccatacg tgtcctcctt accagaaatt tatccttaag atcgtttaaa
     6541 ctcgactctg gctctatcga atctccgtcg tttcgagctt acgcgaacag ccgtggcgct
     6601 catttgctcg tcgggcatcg aatctcgtca gctatcgtca gcttaccttt ttggcagcga
     6661 tcgcggctcc cgacatcttg gaccattagc tccacaggta tcttcttccc tctagtggtc
     6721 ataacagcag cttcagctac ctctcaattc aaaaaacccc tcaagacccg tttagaggcc
     6781 ccaaggggtt atgctatcaa tcgttgcgtt acacacacaa aaaaccaaca cacatccatc
     6841 ttcgatggat agcgatttta ttatctaact gctgatcgag tgtagccaga tctagtaatc
     6901 aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat aacttacggt
     6961 aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta
     7021 tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg
     7081 gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga
     7141 cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt
     7201 tcctacttgg cagtacatct acgtattagt catcgctatt accatgctga tgcggttttg
     7261 gcagtacatc aatgggcgtg gatagcggtt tgactcacgg ggatttccaa gtctccaccc
     7321 cattgacgtc aatgggagtt tgttttggca ccaaaatcaa cgggactttc caaaatgtcg
     7381 taacaactcc gccccattga cgcaaatggg cggtaggcgt gtacggtggg aggtctatat
     7441 aagcagagct ggtttagtga accgtcagat cagatctttg tcgatcctac catccactcg
     7501 acacacccgc cagcggccgc
//
