LOCUS       sequence-314728-        7991 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression plasmid for SARS-CoV-2 helicase.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7991)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 7991)
  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-m4133197
FEATURES             Location/Qualifiers
     source          1..7991
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..304
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        306..508
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     459..479
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      577..586
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2389..2409
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             2413..2478
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     polyA_signal    2531..2652
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2542..2561)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2596..2615
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      2767..2813
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      2843..2979
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    3024..3095
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(3321..3909)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3401..3420)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        4409..5618
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     5455..5474
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             5622..6221
                     /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(5622..5641)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     6118..6138
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    6248..6296
                     /note="synthetic polyadenylation signal"
     CDS             join(6562..6630,6631..7422)
                     /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(6780..6799)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      7435..7571
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    7723..7794
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(7799..7845)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    7863..7911
                     /note="synthetic polyadenylation signal"
ORIGIN
        1 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
       61 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      121 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      181 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      241 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      301 catgctgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      361 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      421 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      481 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatca gatctttgtc
      541 gatcctacca tccactcgac acacccgcca gcggccgcca ccatggcggt gggggcttgc
      601 gtactatgta acagccagac gtcattgcgc tgcggtgcat gtataagaag gccgtttctg
      661 tgttgcaaat gctgctacga tcatgttatt tcgacttctc acaagcttgt gctcagtgtc
      721 aatccttatg tgtgtaatgc ccccggatgc gacgtcacag atgttaccca actgtacctg
      781 ggcggcatgt cctattattg taagagccac aaaccaccaa tctccttccc cctgtgcgcc
      841 aacggacagg tgtttgggct ctacaaaaac acatgtgtgg gatctgacaa tgtcaccgac
      901 ttcaacgcta tcgccacatg cgattggact aatgcaggcg actacatttt agccaacacc
      961 tgtaccgaac ggctgaagtt atttgcggct gagactctaa aggcaactga ggaaaccttc
     1021 aaactttcat atggtatcgc cacggtacgt gaggtgttgt ctgatcgaga actgcatctc
     1081 agttgggagg tcgggaagcc tagacctccc cttaatcgaa attacgtgtt cacaggttat
     1141 agggttacca agaacagcaa agtccagatc ggcgaatata cttttgagaa gggagattac
     1201 ggtgacgccg tagtgtatcg tggcacaaca acatacaaac tgaatgttgg ggattacttc
     1261 gtgctgacgt cccacaccgt tatgccgctc tcagctccaa ctttggttcc ccaggaacat
     1321 tacgtgcgga ttactggact ctatccaacg ctaaacataa gcgacgagtt ttccagcaac
     1381 gtggctaatt accaaaaggt cggcatgcag aagtactcca cccttcaggg tcctcctggg
     1441 actggcaaaa gtcacttcgc catcggactg gcactttact atcccagtgc gcgcattgta
     1501 tataccgcct gctcacatgc ggcagtggac gccctgtgtg agaaggcttt aaagtatctg
     1561 ccaatagaca aatgctctcg cattatccct gccagggcca gagtggagtg ctttgataag
     1621 tttaaagtga actcgaccct cgaacagtat gtcttctgca cagttaacgc attgcccgaa
     1681 acgaccgccg atatagttgt gttcgacgaa atctcaatgg ctaccaatta tgatctctcc
     1741 gtcgtcaatg cacggctgcg ggccaaacac tacgtctaca tcggggaccc cgcgcagtta
     1801 ccagcaccgc gaaccctact gactaagggc acacttgagc cagagtactt caacagcgta
     1861 tgtagactga tgaaaaccat tggccctgat atgttcctgg gaacatgtcg gaggtgcccc
     1921 gctgaaatag tggacaccgt ttcagcactt gtgtacgaca ataagctcaa agcacacaag
     1981 gataaatccg cacaatgttt taagatgttt tacaaagggg ttattaccca tgatgtgtct
     2041 agcgctatca atcgccctca gattggcgtg gtgagagagt ttttgacgcg aaatcccgcc
     2101 tggcgaaagg ccgtattcat cagtccttac aactcgcaaa atgccgtcgc tagtaagatt
     2161 ttaggactcc ctactcagac agttgatagc tctcaaggtt ctgaatacga ttatgtgatt
     2221 tttactcaga ctaccgaaac ggcacattct tgtaacgtca accgttttaa cgtggccatt
     2281 acaagggcca aggttggcat cttgtgtatc atgtcagacc gggacctgta tgataaactg
     2341 caattcacat ctttggagat accgcgccgc aatgtcgcta cccttcagga aaacctttac
     2401 tttcagtcga tggattataa ggaccacgat ggagactaca aagatcatga cattgattac
     2461 aaggacgacg atgacaaata agctagcttg actgactgag atacagcgta ccttcagctc
     2521 acagacatga taagatacat tgatgagttt ggacaaacca caactagaat gcagtgaaaa
     2581 aaatgcttta tttgtgaaat ttgtgatgct attgctttat ttgtaaccat tataagctgc
     2641 aataaacaag ttaacaacaa caattgcatt cattttatgt ttcaggttca gggggaggtg
     2701 tgggaggttt tttaaagcaa gtaaaacctc tacaaatgtg gtattggccc atctctatcg
     2761 gtatcgtagc ataacccctt ggggcctcta aacgggtctt gaggggtttt ttgtgcccct
     2821 cgggccggat tgctatctac cggcattggc gcagaaaaaa atgcctgatg cgacgctgcg
     2881 cgtcttatac tcccacatat gccagattca gcaacggata cggcttcccc aacttgccca
     2941 cttccatacg tgtcctcctt accagaaatt tatccttaag gtcgtcagct atcctgcagg
     3001 cgatctctcg atttcgatca agacattcct ttaatggtct tttctggaca ccactagggg
     3061 tcagaagtag ttcatcaaac tttcttccct ccctaatctc attggttacc ttgggctatc
     3121 gaaacttaat taaccagtca agtcagctac ttggcgagat cgacttgtct gggtttcgac
     3181 tacgctcaga attgcgtcag tcaagttcga tctggtcctt gctattgcac ccgttctccg
     3241 attacgagtt tcatttaaat catgtgagca aaaggccagc aaaaggccag gaaccgtaaa
     3301 aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat
     3361 cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc
     3421 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc
     3481 gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt
     3541 tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac
     3601 cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg
     3661 ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca
     3721 gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc
     3781 gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa
     3841 accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa
     3901 ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac
     3961 tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta
     4021 aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt
     4081 taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata
     4141 gttgcattta aatttccgaa ctctccaagg ccctcgtcgg aaaatcttca aacctttcgt
     4201 ccgatccatc ttgcaggcta cctctcgaac gaactatcgc aagtctcttg gccggccttg
     4261 cgccttggct attgcttggc agcgcctatc gccaggtatt actccaatcc cgaatatccg
     4321 agatcgggat cacccgagag aagttcaacc tacatcctca atcccgatct atccgagatc
     4381 cgaggaatat cgaaatcggg gcgcgcctgg cctccgcgcc gggttttggc gcctcccgcg
     4441 ggcgcccccc tcgtcacggc gagcgctgcc acgtcagacg aagggcgcag gagcgtcctg
     4501 atccttccgc ccggacgctc aggacagcgg cccgctgctc ataagactcg gccttagaac
     4561 cccagtatca gcagaaggac attttaggac gggacttggg tgactctagg gcactggttt
     4621 tctttccaga gagcggaaca ggcgaggaaa agtagtccct tctcggcgat tctgcggagg
     4681 gatctccgtg gggcggtgaa cgccgatgat tatataagga cgcgccgggt gtggcacagc
     4741 tagttccgtc gcagccggga tttgggtcgc ggttcttgtt tgtggatcgc tgtgatcgtc
     4801 acttggtgag tagcgggctg ctgggctggc cggggctttc gtggccgccg ggccgctcgg
     4861 tgggacggaa gcgtgtggag agaccgccaa gggctgtagt ctgggtccgc gagcaaggtt
     4921 gccctgaact gggggttggg gggagcgcag caaaatggcg gctgttcccg agtcttgaat
     4981 ggaagacgct tgtgaggcgg gctgtgaggt cgttgaaaca aggtgggggg catggtgggc
     5041 ggcaagaacc caaggtcttg agcccttcgc taatgcggga aagctcttat tcgggtgaga
     5101 tgggctgggc accatctggg gaccctgacg tgaagtttgt cactgactgg agaactcggt
     5161 ttgtcgtctg ttgcgggggc ggcagttatg gcggtgccgt tgggcagtgc acccgtacct
     5221 ttgggagcgc gcgccctcgt cgtgtcgtga cgtcacccgt tctgttggct tataatgcag
     5281 ggtggggcca cctgccggta ggtgtgcggt aggcttttct ccgtcgcagg acgcagggtt
     5341 cgggcctagg gtaggctctc ctgaatcgac aggcgccgga cctctggtga ggggagggat
     5401 aagtgaggcg tcagtttctt tggtcggttt tatgtaccta tcttcttaag tagctgaagc
     5461 tccggttttg aactatgcgc tcggggttgg cgagtgtgtt ttgtgaagtt ttttaggcac
     5521 cttttgaaat gtaatcattt gggtcaatat gtaattttca gtgttagact tgtaaattgt
     5581 ccgctaaatt ctggccgttt ttggcttttt tgttagacaa catgaccgag tacaagccca
     5641 cggtgcgcct cgccacccgc gacgacgtcc ccagggccgt acgcaccctc gccgccgcgt
     5701 tcgccgacta ccccgccacg cgccacaccg tcgatccgga ccgccacatc gagcgggtca
     5761 ccgagctgca agaactcttc ctcacgcgcg tcgggctcga catcggcaag gtgtgggtcg
     5821 cggacgacgg cgccgcggtg gcggtctgga ccacgccgga gagcgtcgaa gcgggggcgg
     5881 tgttcgccga gatcggcccg cgcatggccg agttgagcgg ttcccggctg gccgcgcagc
     5941 aacagatgga gggcctcctg gcgccgcacc ggcccaagga gcccgcgtgg ttcctggcca
     6001 ccgtcggcgt ctcgcccgac caccagggca agggtctggg cagcgccgtc gtgctccccg
     6061 gagtggaggc cgccgagcgc gccggggtgc ccgccttcct ggagacctcc gcgccccgca
     6121 acctcccctt ctacgagcgg ctcggcttca ccgtcaccgc cgacgtcgag gtgcccgaag
     6181 gaccgcgcac ctggtgcatg acccgcaagc ccggtgccta ggctagtatg taagcctagt
     6241 cttagataat aaaatcgcta tccatcgaag atggatgtgt gttggttttt tgtgtgtgta
     6301 acgctaggcg cgcctggtgt accgagaacg atcctctcag tgcgagtctc gacgatccat
     6361 atcgttgctt ggcagtcagc cagtcggaat ccagcttggg acccaggaag tccaatcgtc
     6421 agatattgta ctcaagcctg gtcacggcag cgtaccgatc tgtttaaacc tagatattga
     6481 tagtctgatc ggtcaacgta taatcgagtc ctagcttttg caaacatcta tcaagagaca
     6541 ggatcagcag gaggctttcg catgagtatt caacatttcc gtgtcgccct tattcccttt
     6601 tttgcggcat tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat
     6661 gctgaagatc agttgggtgc gcgagtgggt tacatcgaac tggatctcaa cagcggtaag
     6721 atccttgaga gttttcgccc cgaagaacgc tttccaatga tgagcacttt taaagttctg
     6781 ctatgtggcg cggtattatc ccgtattgac gccgggcaag agcaactcgg tcgccgcata
     6841 cactattctc agaatgactt ggttgagtat tcaccagtca cagaaaagca tcttacggat
     6901 ggcatgacag taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc
     6961 aacttacttc tgacaacgat tggaggaccg aaggagctaa ccgctttttt gcacaacatg
     7021 ggggatcatg taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac
     7081 gacgagcgtg acaccacgat gcctgtagca atggcaacaa ccttgcgtaa actattaact
     7141 ggcgaactac ttactctagc ttcccggcaa cagttgatag actggatgga ggcggataaa
     7201 gttgcaggac cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct
     7261 ggagccggtg agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc
     7321 tcccgtatcg tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga
     7381 cagatcgctg agataggtgc ctcactgatt aagcattggt aaccgattct aggtgcattg
     7441 gcgcagaaaa aaatgcctga tgcgacgctg cgcgtcttat actcccacat atgccagatt
     7501 cagcaacgga tacggcttcc ccaacttgcc cacttccata cgtgtcctcc ttaccagaaa
     7561 tttatcctta agatcccgaa tcgtttaaac tcgactctgg ctctatcgaa tctccgtcgt
     7621 ttcgagctta cgcgaacagc cgtggcgctc atttgctcgt cgggcatcga atctcgtcag
     7681 ctatcgtcag cttacctttt tggcagcgat cgcggctccc gacatcttgg accattagct
     7741 ccacaggtat cttcttccct ctagtggtca taacagcagc ttcagctacc tctcaattca
     7801 aaaaacccct caagacccgt ttagaggccc caaggggtta tgctatcaat cgttgcgtta
     7861 cacacacaaa aaaccaacac acatccatct tcgatggata gcgattttat tatctaactg
     7921 ctgatcgagt gtagccagat ctagtaatca attacggggt cattagttca tagcccatat
     7981 atggagttcc g
//
