LOCUS       sequence-295742-        8464 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 8464)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 8464)
  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-m4133756
FEATURES             Location/Qualifiers
     source          1..8464
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1805..1825
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             1859..1936
                     /codon_start=1
                     /product="calmodulin-binding peptide"
                     /label=CBP
                     /note="derived from skeletal muscle myosin light chain 
                     kinase; binds calmodulin with nanomolar affinity in the 
                     presence of calcium"
                     /translation="KRRWKKNFIAVSAANRFKKISSSGAL"
     CDS             1964..1984
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     polyA_signal    2433..2554
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2444..2463)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2498..2517
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      2669..2715
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      2745..2881
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    2926..2997
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(3223..3811)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3303..3322)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        4311..5520
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     5357..5376
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             5524..6123
                     /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(5524..5543)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     6020..6040
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    6150..6198
                     /note="synthetic polyadenylation signal"
     CDS             join(6464..6532,6533..7324)
                     /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(6682..6701)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      7337..7473
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    7618..7689
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(7694..7740)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    7758..7806
                     /note="synthetic polyadenylation signal"
     enhancer        7887..8190
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        8192..8394
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     8345..8365
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      join(8463..8464,1..8)
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
ORIGIN
        1 caccatggtt ggggcttgtg ttctttgcaa ttcacagact tcattaagat gtggtgcttg
       61 catacgtaga ccattcttat gttgtaaatg ctgttacgac catgtcatat caacatcaca
      121 taaattagtc ttgtctgtta atccgtatgt ttgcaatgct ccaggttgtg atgtcacaga
      181 tgtgactcaa ctttacttag gaggtatgag ctattattgt aaatcacata aaccacccat
      241 tagttttcca ttgtgtgcta atggacaagt ttttggttta tataaaaata catgtgttgg
      301 tagcgataat gttactgact ttaatgcaat tgcaacatgt gactggacaa atgctggtga
      361 ttacatttta gctaacacct gtactgaaag actcaagctt tttgcagcag aaacgctcaa
      421 agctactgag gagacattta aactgtctta tggtattgct actgtacgtg aagtgctgtc
      481 tgacagagaa ttacatcttt catgggaagt tggtaaacct agaccaccac ttaaccgaaa
      541 ttatgtcttt actggttatc gtgtaactaa aaacagtaaa gtacaaatag gagagtacac
      601 ctttgaaaaa ggtgactatg gtgatgctgt tgtttaccga ggtacaacaa cttacaaatt
      661 aaatgttggt gattattttg tgctgacatc acatacagta atgccattaa gtgcacctac
      721 actagtgcca caagagcact atgttagaat tactggctta tacccaacac tcaatatctc
      781 agatgagttt tctagcaatg ttgcaaatta tcaaaaggtt ggtatgcaaa agtattctac
      841 actccaggga ccacctggta ctggtaagag tcattttgct attggcctag ctctctacta
      901 cccttctgct cgcatagtgt atacagcttg ctctcatgcc gctgttgatg cactatgtga
      961 gaaggcatta aaatatttgc ctatagataa atgtagtaga attatacctg cacgtgctcg
     1021 tgtagagtgt tttgataaat tcaaagtgaa ttcaacatta gaacagtatg tcttttgtac
     1081 tgtaaatgca ttgcctgaga cgacagcaga tatagttgtc tttgatgaaa tttcaatggc
     1141 cacaaattat gatttgagtg ttgtcaatgc cagattacgt gctaagcact atgtgtacat
     1201 tggcgaccct gctcaattac ctgcaccacg cacattgcta actaagggca cactagaacc
     1261 agaatatttc aattcagtgt gtagacttat gaaaactata ggtccagaca tgttcctcgg
     1321 aacttgtcgg cgttgtcctg ctgaaattgt tgacactgtg agtgctttgg tttatgataa
     1381 taagcttaaa gcacataaag acaaatcagc tcaatgcttt aaaatgtttt ataagggtgt
     1441 tatcacgcat gatgtttcat ctgcaattaa caggccacaa ataggcgtgg taagagaatt
     1501 ccttacacgt aaccctgctt ggagaaaagc tgtctttatt tcaccttata attcacagaa
     1561 tgctgtagcc tcaaagattt tgggactacc aactcaaact gttgattcat cacagggctc
     1621 agaatatgac tatgtcatat tcactcaaac cactgaaaca gctcactctt gtaatgtaaa
     1681 cagatttaat gttgctatta ccagagcaaa agtaggcata ctttgcataa tgtctgatag
     1741 agacctttat gacaagttgc aatttacaag tcttgaaatt ccacgtagga atgtggcaac
     1801 tttagaaaac ctttactttc agtcgggacg gcgaatcccg gggctgataa atccctggaa
     1861 aaggagatgg aagaagaact ttattgcagt gtctgcggct aaccggttca aaaagatcag
     1921 ttcaagcggc gccctagatt acgacattcc tacgaccgcc tccgagaatc tctatttcca
     1981 gggtgaattt ggcttggcac aacatgacga ggccgtcgat aacaagttta ataaagaaca
     2041 gcagaacgct ttctatgaga tccttcactt accaaatctg aacgaagagc aacgcaacgc
     2101 tttcattcag agcctgaaag acgatccctc gcagtctgca aatctcctgg ccgaagccaa
     2161 gaagctgaat gacgcacagg ctccaaaagt tgataacaag tttaacaagg agcagcagaa
     2221 tgctttctac gagatattgc accttcctaa tctgaatgag gaacaaagaa atgcatttat
     2281 ccagtcccta aaagacgatc catcacagag tgccaacctg cttgcggaag ccaaaaagct
     2341 caatgatgcc caggccccga aagtagacgc gaaccatcaa taagctagct tgactgactg
     2401 agatacagcg taccttcagc tcacagacat gataagatac attgatgagt ttggacaaac
     2461 cacaactaga atgcagtgaa aaaaatgctt tatttgtgaa atttgtgatg ctattgcttt
     2521 atttgtaacc attataagct gcaataaaca agttaacaac aacaattgca ttcattttat
     2581 gtttcaggtt cagggggagg tgtgggaggt tttttaaagc aagtaaaacc tctacaaatg
     2641 tggtattggc ccatctctat cggtatcgta gcataacccc ttggggcctc taaacgggtc
     2701 ttgaggggtt ttttgtgccc ctcgggccgg attgctatct accggcattg gcgcagaaaa
     2761 aaatgcctga tgcgacgctg cgcgtcttat actcccacat atgccagatt cagcaacgga
     2821 tacggcttcc ccaacttgcc cacttccata cgtgtcctcc ttaccagaaa tttatcctta
     2881 aggtcgtcag ctatcctgca ggcgatctct cgatttcgat caagacattc ctttaatggt
     2941 cttttctgga caccactagg ggtcagaagt agttcatcaa actttcttcc ctccctaatc
     3001 tcattggtta ccttgggcta tcgaaactta attaaccagt caagtcagct acttggcgag
     3061 atcgacttgt ctgggtttcg actacgctca gaattgcgtc agtcaagttc gatctggtcc
     3121 ttgctattgc acccgttctc cgattacgag tttcatttaa atcatgtgag caaaaggcca
     3181 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc
     3241 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
     3301 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
     3361 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag
     3421 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca
     3481 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa
     3541 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc
     3601 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag
     3661 aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg
     3721 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca
     3781 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc
     3841 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag
     3901 gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata
     3961 tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat
     4021 ctgtctattt cgttcatcca tagttgcatt taaatttccg aactctccaa ggccctcgtc
     4081 ggaaaatctt caaacctttc gtccgatcca tcttgcaggc tacctctcga acgaactatc
     4141 gcaagtctct tggccggcct tgcgccttgg ctattgcttg gcagcgccta tcgccaggta
     4201 ttactccaat cccgaatatc cgagatcggg atcacccgag agaagttcaa cctacatcct
     4261 caatcccgat ctatccgaga tccgaggaat atcgaaatcg gggcgcgcct ggcctccgcg
     4321 ccgggttttg gcgcctcccg cgggcgcccc cctcgtcacg gcgagcgctg ccacgtcaga
     4381 cgaagggcgc aggagcgtcc tgatccttcc gcccggacgc tcaggacagc ggcccgctgc
     4441 tcataagact cggccttaga accccagtat cagcagaagg acattttagg acgggacttg
     4501 ggtgactcta gggcactggt tttctttcca gagagcggaa caggcgagga aaagtagtcc
     4561 cttctcggcg attctgcgga gggatctccg tggggcggtg aacgccgatg attatataag
     4621 gacgcgccgg gtgtggcaca gctagttccg tcgcagccgg gatttgggtc gcggttcttg
     4681 tttgtggatc gctgtgatcg tcacttggtg agtagcgggc tgctgggctg gccggggctt
     4741 tcgtggccgc cgggccgctc ggtgggacgg aagcgtgtgg agagaccgcc aagggctgta
     4801 gtctgggtcc gcgagcaagg ttgccctgaa ctgggggttg gggggagcgc agcaaaatgg
     4861 cggctgttcc cgagtcttga atggaagacg cttgtgaggc gggctgtgag gtcgttgaaa
     4921 caaggtgggg ggcatggtgg gcggcaagaa cccaaggtct tgagcccttc gctaatgcgg
     4981 gaaagctctt attcgggtga gatgggctgg gcaccatctg gggaccctga cgtgaagttt
     5041 gtcactgact ggagaactcg gtttgtcgtc tgttgcgggg gcggcagtta tggcggtgcc
     5101 gttgggcagt gcacccgtac ctttgggagc gcgcgccctc gtcgtgtcgt gacgtcaccc
     5161 gttctgttgg cttataatgc agggtggggc cacctgccgg taggtgtgcg gtaggctttt
     5221 ctccgtcgca ggacgcaggg ttcgggccta gggtaggctc tcctgaatcg acaggcgccg
     5281 gacctctggt gaggggaggg ataagtgagg cgtcagtttc tttggtcggt tttatgtacc
     5341 tatcttctta agtagctgaa gctccggttt tgaactatgc gctcggggtt ggcgagtgtg
     5401 ttttgtgaag ttttttaggc accttttgaa atgtaatcat ttgggtcaat atgtaatttt
     5461 cagtgttaga cttgtaaatt gtccgctaaa ttctggccgt ttttggcttt tttgttagac
     5521 aacatgaccg agtacaagcc cacggtgcgc ctcgccaccc gcgacgacgt ccccagggcc
     5581 gtacgcaccc tcgccgccgc gttcgccgac taccccgcca cgcgccacac cgtcgatccg
     5641 gaccgccaca tcgagcgggt caccgagctg caagaactct tcctcacgcg cgtcgggctc
     5701 gacatcggca aggtgtgggt cgcggacgac ggcgccgcgg tggcggtctg gaccacgccg
     5761 gagagcgtcg aagcgggggc ggtgttcgcc gagatcggcc cgcgcatggc cgagttgagc
     5821 ggttcccggc tggccgcgca gcaacagatg gagggcctcc tggcgccgca ccggcccaag
     5881 gagcccgcgt ggttcctggc caccgtcggc gtctcgcccg accaccaggg caagggtctg
     5941 ggcagcgccg tcgtgctccc cggagtggag gccgccgagc gcgccggggt gcccgccttc
     6001 ctggagacct ccgcgccccg caacctcccc ttctacgagc ggctcggctt caccgtcacc
     6061 gccgacgtcg aggtgcccga aggaccgcgc acctggtgca tgacccgcaa gcccggtgcc
     6121 taggctagta tgtaagccta gtcttagata ataaaatcgc tatccatcga agatggatgt
     6181 gtgttggttt tttgtgtgtg taacgctagg cgcgcctggt gtaccgagaa cgatcctctc
     6241 agtgcgagtc tcgacgatcc atatcgttgc ttggcagtca gccagtcgga atccagcttg
     6301 ggacccagga agtccaatcg tcagatattg tactcaagcc tggtcacggc agcgtaccga
     6361 tctgtttaaa cctagatatt gatagtctga tcggtcaacg tataatcgag tcctagcttt
     6421 tgcaaacatc tatcaagaga caggatcagc aggaggcttt cgcatgagta ttcaacattt
     6481 ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga
     6541 aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcgcgagtgg gttacatcga
     6601 actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac gctttccaat
     6661 gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca
     6721 agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt attcaccagt
     6781 cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac
     6841 catgagtgat aacactgcgg ccaacttact tctgacaacg attggaggac cgaaggagct
     6901 aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga
     6961 gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag caatggcaac
     7021 aaccttgcgt aaactattaa ctggcgaact acttactcta gcttcccggc aacagttgat
     7081 agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg
     7141 ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc
     7201 actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc
     7261 aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg
     7321 gtaaccgatt ctaggtgcat tggcgcagaa aaaaatgcct gatgcgacgc tgcgcgtctt
     7381 atactcccac atatgccaga ttcagcaacg gatacggctt ccccaacttg cccacttcca
     7441 tacgtgtcct ccttaccaga aatttatcct taagatcgtt taaactcgac tctggctcta
     7501 tcgaatctcc gtcgtttcga gcttacgcga acagccgtgg cgctcatttg ctcgtcgggc
     7561 atcgaatctc gtcagctatc gtcagcttac ctttttggca gcgatcgcgg ctcccgacat
     7621 cttggaccat tagctccaca ggtatcttct tccctctagt ggtcataaca gcagcttcag
     7681 ctacctctca attcaaaaaa cccctcaaga cccgtttaga ggccccaagg ggttatgcta
     7741 tcaatcgttg cgttacacac acaaaaaacc aacacacatc catcttcgat ggatagcgat
     7801 tttattatct aactgctgat cgagtgtagc cagatctagt aatcaattac ggggtcatta
     7861 gttcatagcc catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc
     7921 tgaccgccca acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg
     7981 ccaataggga ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg
     8041 gcagtacatc aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa
     8101 tggcccgcct ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac
     8161 atctacgtat tagtcatcgc tattaccatg ctgatgcggt tttggcagta catcaatggg
     8221 cgtggatagc ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg
     8281 agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca
     8341 ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctggttta
     8401 gtgaaccgtc agatcagatc tttgtcgatc ctaccatcca ctcgacacac ccgccagcgg
     8461 ccgc
//
