LOCUS       sequence-295743-        7018 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 7018)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 7018)
  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-m4133419
FEATURES             Location/Qualifiers
     source          1..7018
                     /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             917..934
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     polyA_signal    987..1108
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(998..1017)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1052..1071
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      1223..1269
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1299..1435
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1480..1551
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(1777..2365)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1857..1876)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2865..4074
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     3911..3930
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             4078..4677
                     /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(4078..4097)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4574..4594
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4704..4752
                     /note="synthetic polyadenylation signal"
     CDS             join(5018..5086,5087..5878)
                     /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(5236..5255)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      5891..6027
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    6172..6243
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(6248..6294)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    6312..6360
                     /note="synthetic polyadenylation signal"
     enhancer        6441..6744
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6746..6948
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     6899..6919
                     /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 cagtcgcacc atcaccatca ccattaagct agcttgactg actgagatac
      961 agcgtacctt cagctcacag acatgataag atacattgat gagtttggac aaaccacaac
     1021 tagaatgcag tgaaaaaaat gctttatttg tgaaatttgt gatgctattg ctttatttgt
     1081 aaccattata agctgcaata aacaagttaa caacaacaat tgcattcatt ttatgtttca
     1141 ggttcagggg gaggtgtggg aggtttttta aagcaagtaa aacctctaca aatgtggtat
     1201 tggcccatct ctatcggtat cgtagcataa ccccttgggg cctctaaacg ggtcttgagg
     1261 ggttttttgt gcccctcggg ccggattgct atctaccggc attggcgcag aaaaaaatgc
     1321 ctgatgcgac gctgcgcgtc ttatactccc acatatgcca gattcagcaa cggatacggc
     1381 ttccccaact tgcccacttc catacgtgtc ctccttacca gaaatttatc cttaaggtcg
     1441 tcagctatcc tgcaggcgat ctctcgattt cgatcaagac attcctttaa tggtcttttc
     1501 tggacaccac taggggtcag aagtagttca tcaaactttc ttccctccct aatctcattg
     1561 gttaccttgg gctatcgaaa cttaattaac cagtcaagtc agctacttgg cgagatcgac
     1621 ttgtctgggt ttcgactacg ctcagaattg cgtcagtcaa gttcgatctg gtccttgcta
     1681 ttgcacccgt tctccgatta cgagtttcat ttaaatcatg tgagcaaaag gccagcaaaa
     1741 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     1801 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     1861 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     1921 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     1981 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     2041 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     2101 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     2161 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac
     2221 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     2281 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     2341 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     2401 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
     2461 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
     2521 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     2581 atttcgttca tccatagttg catttaaatt tccgaactct ccaaggccct cgtcggaaaa
     2641 tcttcaaacc tttcgtccga tccatcttgc aggctacctc tcgaacgaac tatcgcaagt
     2701 ctcttggccg gccttgcgcc ttggctattg cttggcagcg cctatcgcca ggtattactc
     2761 caatcccgaa tatccgagat cgggatcacc cgagagaagt tcaacctaca tcctcaatcc
     2821 cgatctatcc gagatccgag gaatatcgaa atcggggcgc gcctggcctc cgcgccgggt
     2881 tttggcgcct cccgcgggcg cccccctcgt cacggcgagc gctgccacgt cagacgaagg
     2941 gcgcaggagc gtcctgatcc ttccgcccgg acgctcagga cagcggcccg ctgctcataa
     3001 gactcggcct tagaacccca gtatcagcag aaggacattt taggacggga cttgggtgac
     3061 tctagggcac tggttttctt tccagagagc ggaacaggcg aggaaaagta gtcccttctc
     3121 ggcgattctg cggagggatc tccgtggggc ggtgaacgcc gatgattata taaggacgcg
     3181 ccgggtgtgg cacagctagt tccgtcgcag ccgggatttg ggtcgcggtt cttgtttgtg
     3241 gatcgctgtg atcgtcactt ggtgagtagc gggctgctgg gctggccggg gctttcgtgg
     3301 ccgccgggcc gctcggtggg acggaagcgt gtggagagac cgccaagggc tgtagtctgg
     3361 gtccgcgagc aaggttgccc tgaactgggg gttgggggga gcgcagcaaa atggcggctg
     3421 ttcccgagtc ttgaatggaa gacgcttgtg aggcgggctg tgaggtcgtt gaaacaaggt
     3481 ggggggcatg gtgggcggca agaacccaag gtcttgagcc cttcgctaat gcgggaaagc
     3541 tcttattcgg gtgagatggg ctgggcacca tctggggacc ctgacgtgaa gtttgtcact
     3601 gactggagaa ctcggtttgt cgtctgttgc gggggcggca gttatggcgg tgccgttggg
     3661 cagtgcaccc gtacctttgg gagcgcgcgc cctcgtcgtg tcgtgacgtc acccgttctg
     3721 ttggcttata atgcagggtg gggccacctg ccggtaggtg tgcggtaggc ttttctccgt
     3781 cgcaggacgc agggttcggg cctagggtag gctctcctga atcgacaggc gccggacctc
     3841 tggtgagggg agggataagt gaggcgtcag tttctttggt cggttttatg tacctatctt
     3901 cttaagtagc tgaagctccg gttttgaact atgcgctcgg ggttggcgag tgtgttttgt
     3961 gaagtttttt aggcaccttt tgaaatgtaa tcatttgggt caatatgtaa ttttcagtgt
     4021 tagacttgta aattgtccgc taaattctgg ccgtttttgg cttttttgtt agacaacatg
     4081 accgagtaca agcccacggt gcgcctcgcc acccgcgacg acgtccccag ggccgtacgc
     4141 accctcgccg ccgcgttcgc cgactacccc gccacgcgcc acaccgtcga tccggaccgc
     4201 cacatcgagc gggtcaccga gctgcaagaa ctcttcctca cgcgcgtcgg gctcgacatc
     4261 ggcaaggtgt gggtcgcgga cgacggcgcc gcggtggcgg tctggaccac gccggagagc
     4321 gtcgaagcgg gggcggtgtt cgccgagatc ggcccgcgca tggccgagtt gagcggttcc
     4381 cggctggccg cgcagcaaca gatggagggc ctcctggcgc cgcaccggcc caaggagccc
     4441 gcgtggttcc tggccaccgt cggcgtctcg cccgaccacc agggcaaggg tctgggcagc
     4501 gccgtcgtgc tccccggagt ggaggccgcc gagcgcgccg gggtgcccgc cttcctggag
     4561 acctccgcgc cccgcaacct ccccttctac gagcggctcg gcttcaccgt caccgccgac
     4621 gtcgaggtgc ccgaaggacc gcgcacctgg tgcatgaccc gcaagcccgg tgcctaggct
     4681 agtatgtaag cctagtctta gataataaaa tcgctatcca tcgaagatgg atgtgtgttg
     4741 gttttttgtg tgtgtaacgc taggcgcgcc tggtgtaccg agaacgatcc tctcagtgcg
     4801 agtctcgacg atccatatcg ttgcttggca gtcagccagt cggaatccag cttgggaccc
     4861 aggaagtcca atcgtcagat attgtactca agcctggtca cggcagcgta ccgatctgtt
     4921 taaacctaga tattgatagt ctgatcggtc aacgtataat cgagtcctag cttttgcaaa
     4981 catctatcaa gagacaggat cagcaggagg ctttcgcatg agtattcaac atttccgtgt
     5041 cgcccttatt cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct
     5101 ggtgaaagta aaagatgctg aagatcagtt gggtgcgcga gtgggttaca tcgaactgga
     5161 tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa gaacgctttc caatgatgag
     5221 cacttttaaa gttctgctat gtggcgcggt attatcccgt attgacgccg ggcaagagca
     5281 actcggtcgc cgcatacact attctcagaa tgacttggtt gagtattcac cagtcacaga
     5341 aaagcatctt acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag
     5401 tgataacact gcggccaact tacttctgac aacgattgga ggaccgaagg agctaaccgc
     5461 ttttttgcac aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa
     5521 tgaagccata ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg caacaacctt
     5581 gcgtaaacta ttaactggcg aactacttac tctagcttcc cggcaacagt tgatagactg
     5641 gatggaggcg gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt
     5701 tattgctgat aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg
     5761 gccagatggt aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat
     5821 ggatgaacga aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaacc
     5881 gattctaggt gcattggcgc agaaaaaaat gcctgatgcg acgctgcgcg tcttatactc
     5941 ccacatatgc cagattcagc aacggatacg gcttccccaa cttgcccact tccatacgtg
     6001 tcctccttac cagaaattta tccttaagat cgtttaaact cgactctggc tctatcgaat
     6061 ctccgtcgtt tcgagcttac gcgaacagcc gtggcgctca tttgctcgtc gggcatcgaa
     6121 tctcgtcagc tatcgtcagc ttaccttttt ggcagcgatc gcggctcccg acatcttgga
     6181 ccattagctc cacaggtatc ttcttccctc tagtggtcat aacagcagct tcagctacct
     6241 ctcaattcaa aaaacccctc aagacccgtt tagaggcccc aaggggttat gctatcaatc
     6301 gttgcgttac acacacaaaa aaccaacaca catccatctt cgatggatag cgattttatt
     6361 atctaactgc tgatcgagtg tagccagatc tagtaatcaa ttacggggtc attagttcat
     6421 agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg
     6481 cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata
     6541 gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta
     6601 catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc
     6661 gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca gtacatctac
     6721 gtattagtca tcgctattac catgctgatg cggttttggc agtacatcaa tgggcgtgga
     6781 tagcggtttg actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg
     6841 ttttggcacc aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg
     6901 caaatgggcg gtaggcgtgt acggtgggag gtctatataa gcagagctgg tttagtgaac
     6961 cgtcagatca gatctttgtc gatcctacca tccactcgac acacccgcca gcggccgc
//
