LOCUS       sequence-295418-        7069 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 7069)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 7069)
  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-m4133187
FEATURES             Location/Qualifiers
     source          1..7069
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             896..916
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             920..985
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     polyA_signal    1038..1159
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1049..1068)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1103..1122
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      1274..1320
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1350..1486
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1531..1602
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(1828..2416)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1908..1927)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2916..4125
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     3962..3981
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             4129..4728
                     /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(4129..4148)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4625..4645
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4755..4803
                     /note="synthetic polyadenylation signal"
     CDS             join(5069..5137,5138..5929)
                     /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(5287..5306)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      5942..6078
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    6223..6294
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(6299..6345)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    6363..6411
                     /note="synthetic polyadenylation signal"
     enhancer        6492..6795
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6797..6999
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     6950..6970
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
ORIGIN
        1 caccatgagt caagcgtggc aaccgggtgt tgctatgcct aatctttaca aaatgcaaag
       61 aatgctatta gaaaagtgtg accttcaaaa ttatggtgat agtgcaacat tacctaaagg
      121 cataatgatg aatgtcgcaa aatatactca actgtgtcaa tatttaaaca cattaacatt
      181 agctgtaccc tataatatga gagttataca ttttggtgct ggttctgata aaggagttgc
      241 accaggtaca gctgttttaa gacagtggtt gcctacgggt acgctgcttg tcgattcaga
      301 tcttaatgac tttgtctctg atgcagattc aactttgatt ggtgattgtg caactgtaca
      361 tacagctaat aaatgggatc tcattattag tgatatgtac gaccctaaga ctaaaaatgt
      421 tacaaaagaa aatgactcta aagagggttt tttcacttac atttgtgggt ttatacaaca
      481 aaagctagct cttggaggtt ccgtggctat aaagataaca gaacattctt ggaatgctga
      541 tctttataag ctcatgggac acttcgcatg gtggacagcc tttgttacta atgtgaatgc
      601 gtcatcatct gaagcatttt taattggatg taattatctt ggcaaaccac gcgaacaaat
      661 agatggttat gtcatgcatg caaattacat attttggagg aatacaaatc caattcagtt
      721 gtcttcctat tctttatttg acatgagtaa atttcccctt aaattaaggg gtactgctgt
      781 tatgtcttta aaagaaggtc aaatcaatga tatgatttta tctcttctta gtaaaggtag
      841 acttataatt agagaaaaca acagagttgt tatttctagt gatgttcttg ttaacgaaaa
      901 cctttacttt cagtcgatgg attataagga ccacgatgga gactacaaag atcatgacat
      961 tgattacaag gacgacgatg acaaataagc tagcttgact gactgagata cagcgtacct
     1021 tcagctcaca gacatgataa gatacattga tgagtttgga caaaccacaa ctagaatgca
     1081 gtgaaaaaaa tgctttattt gtgaaatttg tgatgctatt gctttatttg taaccattat
     1141 aagctgcaat aaacaagtta acaacaacaa ttgcattcat tttatgtttc aggttcaggg
     1201 ggaggtgtgg gaggtttttt aaagcaagta aaacctctac aaatgtggta ttggcccatc
     1261 tctatcggta tcgtagcata accccttggg gcctctaaac gggtcttgag gggttttttg
     1321 tgcccctcgg gccggattgc tatctaccgg cattggcgca gaaaaaaatg cctgatgcga
     1381 cgctgcgcgt cttatactcc cacatatgcc agattcagca acggatacgg cttccccaac
     1441 ttgcccactt ccatacgtgt cctccttacc agaaatttat ccttaaggtc gtcagctatc
     1501 ctgcaggcga tctctcgatt tcgatcaaga cattccttta atggtctttt ctggacacca
     1561 ctaggggtca gaagtagttc atcaaacttt cttccctccc taatctcatt ggttaccttg
     1621 ggctatcgaa acttaattaa ccagtcaagt cagctacttg gcgagatcga cttgtctggg
     1681 tttcgactac gctcagaatt gcgtcagtca agttcgatct ggtccttgct attgcacccg
     1741 ttctccgatt acgagtttca tttaaatcat gtgagcaaaa ggccagcaaa aggccaggaa
     1801 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca
     1861 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc
     1921 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
     1981 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta
     2041 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
     2101 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
     2161 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
     2221 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg
     2281 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
     2341 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
     2401 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     2461 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
     2521 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
     2581 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     2641 atccatagtt gcatttaaat ttccgaactc tccaaggccc tcgtcggaaa atcttcaaac
     2701 ctttcgtccg atccatcttg caggctacct ctcgaacgaa ctatcgcaag tctcttggcc
     2761 ggccttgcgc cttggctatt gcttggcagc gcctatcgcc aggtattact ccaatcccga
     2821 atatccgaga tcgggatcac ccgagagaag ttcaacctac atcctcaatc ccgatctatc
     2881 cgagatccga ggaatatcga aatcggggcg cgcctggcct ccgcgccggg ttttggcgcc
     2941 tcccgcgggc gcccccctcg tcacggcgag cgctgccacg tcagacgaag ggcgcaggag
     3001 cgtcctgatc cttccgcccg gacgctcagg acagcggccc gctgctcata agactcggcc
     3061 ttagaacccc agtatcagca gaaggacatt ttaggacggg acttgggtga ctctagggca
     3121 ctggttttct ttccagagag cggaacaggc gaggaaaagt agtcccttct cggcgattct
     3181 gcggagggat ctccgtgggg cggtgaacgc cgatgattat ataaggacgc gccgggtgtg
     3241 gcacagctag ttccgtcgca gccgggattt gggtcgcggt tcttgtttgt ggatcgctgt
     3301 gatcgtcact tggtgagtag cgggctgctg ggctggccgg ggctttcgtg gccgccgggc
     3361 cgctcggtgg gacggaagcg tgtggagaga ccgccaaggg ctgtagtctg ggtccgcgag
     3421 caaggttgcc ctgaactggg ggttgggggg agcgcagcaa aatggcggct gttcccgagt
     3481 cttgaatgga agacgcttgt gaggcgggct gtgaggtcgt tgaaacaagg tggggggcat
     3541 ggtgggcggc aagaacccaa ggtcttgagc ccttcgctaa tgcgggaaag ctcttattcg
     3601 ggtgagatgg gctgggcacc atctggggac cctgacgtga agtttgtcac tgactggaga
     3661 actcggtttg tcgtctgttg cgggggcggc agttatggcg gtgccgttgg gcagtgcacc
     3721 cgtacctttg ggagcgcgcg ccctcgtcgt gtcgtgacgt cacccgttct gttggcttat
     3781 aatgcagggt ggggccacct gccggtaggt gtgcggtagg cttttctccg tcgcaggacg
     3841 cagggttcgg gcctagggta ggctctcctg aatcgacagg cgccggacct ctggtgaggg
     3901 gagggataag tgaggcgtca gtttctttgg tcggttttat gtacctatct tcttaagtag
     3961 ctgaagctcc ggttttgaac tatgcgctcg gggttggcga gtgtgttttg tgaagttttt
     4021 taggcacctt ttgaaatgta atcatttggg tcaatatgta attttcagtg ttagacttgt
     4081 aaattgtccg ctaaattctg gccgtttttg gcttttttgt tagacaacat gaccgagtac
     4141 aagcccacgg tgcgcctcgc cacccgcgac gacgtcccca gggccgtacg caccctcgcc
     4201 gccgcgttcg ccgactaccc cgccacgcgc cacaccgtcg atccggaccg ccacatcgag
     4261 cgggtcaccg agctgcaaga actcttcctc acgcgcgtcg ggctcgacat cggcaaggtg
     4321 tgggtcgcgg acgacggcgc cgcggtggcg gtctggacca cgccggagag cgtcgaagcg
     4381 ggggcggtgt tcgccgagat cggcccgcgc atggccgagt tgagcggttc ccggctggcc
     4441 gcgcagcaac agatggaggg cctcctggcg ccgcaccggc ccaaggagcc cgcgtggttc
     4501 ctggccaccg tcggcgtctc gcccgaccac cagggcaagg gtctgggcag cgccgtcgtg
     4561 ctccccggag tggaggccgc cgagcgcgcc ggggtgcccg ccttcctgga gacctccgcg
     4621 ccccgcaacc tccccttcta cgagcggctc ggcttcaccg tcaccgccga cgtcgaggtg
     4681 cccgaaggac cgcgcacctg gtgcatgacc cgcaagcccg gtgcctaggc tagtatgtaa
     4741 gcctagtctt agataataaa atcgctatcc atcgaagatg gatgtgtgtt ggttttttgt
     4801 gtgtgtaacg ctaggcgcgc ctggtgtacc gagaacgatc ctctcagtgc gagtctcgac
     4861 gatccatatc gttgcttggc agtcagccag tcggaatcca gcttgggacc caggaagtcc
     4921 aatcgtcaga tattgtactc aagcctggtc acggcagcgt accgatctgt ttaaacctag
     4981 atattgatag tctgatcggt caacgtataa tcgagtccta gcttttgcaa acatctatca
     5041 agagacagga tcagcaggag gctttcgcat gagtattcaa catttccgtg tcgcccttat
     5101 tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt
     5161 aaaagatgct gaagatcagt tgggtgcgcg agtgggttac atcgaactgg atctcaacag
     5221 cggtaagatc cttgagagtt ttcgccccga agaacgcttt ccaatgatga gcacttttaa
     5281 agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg
     5341 ccgcatacac tattctcaga atgacttggt tgagtattca ccagtcacag aaaagcatct
     5401 tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac
     5461 tgcggccaac ttacttctga caacgattgg aggaccgaag gagctaaccg cttttttgca
     5521 caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat
     5581 accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacct tgcgtaaact
     5641 attaactggc gaactactta ctctagcttc ccggcaacag ttgatagact ggatggaggc
     5701 ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga
     5761 taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg
     5821 taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg
     5881 aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac cgattctagg
     5941 tgcattggcg cagaaaaaaa tgcctgatgc gacgctgcgc gtcttatact cccacatatg
     6001 ccagattcag caacggatac ggcttcccca acttgcccac ttccatacgt gtcctcctta
     6061 ccagaaattt atccttaaga tcgtttaaac tcgactctgg ctctatcgaa tctccgtcgt
     6121 ttcgagctta cgcgaacagc cgtggcgctc atttgctcgt cgggcatcga atctcgtcag
     6181 ctatcgtcag cttacctttt tggcagcgat cgcggctccc gacatcttgg accattagct
     6241 ccacaggtat cttcttccct ctagtggtca taacagcagc ttcagctacc tctcaattca
     6301 aaaaacccct caagacccgt ttagaggccc caaggggtta tgctatcaat cgttgcgtta
     6361 cacacacaaa aaaccaacac acatccatct tcgatggata gcgattttat tatctaactg
     6421 ctgatcgagt gtagccagat ctagtaatca attacggggt cattagttca tagcccatat
     6481 atggagttcc gcgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac
     6541 ccccgcccat tgacgtcaat aatgacgtat gttcccatag taacgccaat agggactttc
     6601 cattgacgtc aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg
     6661 tatcatatgc caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat
     6721 tatgcccagt acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc
     6781 atcgctatta ccatgctgat gcggttttgg cagtacatca atgggcgtgg atagcggttt
     6841 gactcacggg gatttccaag tctccacccc attgacgtca atgggagttt gttttggcac
     6901 caaaatcaac gggactttcc aaaatgtcgt aacaactccg ccccattgac gcaaatgggc
     6961 ggtaggcgtg tacggtggga ggtctatata agcagagctg gtttagtgaa ccgtcagatc
     7021 agatctttgt cgatcctacc atccactcga cacacccgcc agcggccgc
//
