LOCUS       sequence-295981-        7204 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression plasmid for SARS-CoV-2 leader protein.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7204)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 7204)
  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-m4133563
FEATURES             Location/Qualifiers
     source          1..7204
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             545..565
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             599..676
                     /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             704..724
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     polyA_signal    1173..1294
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1184..1203)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1238..1257
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      1409..1455
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1485..1621
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1666..1737
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(1963..2551)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2043..2062)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        3051..4260
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     4097..4116
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             4264..4863
                     /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(4264..4283)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4760..4780
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4890..4938
                     /note="synthetic polyadenylation signal"
     CDS             join(5204..5272,5273..6064)
                     /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(5422..5441)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      6077..6213
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    6358..6429
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(6434..6480)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    6498..6546
                     /note="synthetic polyadenylation signal"
     enhancer        6627..6930
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6932..7134
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     7085..7105
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      join(7203..7204,1..8)
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
ORIGIN
        1 caccatggag tctctcgttc ccggctttaa tgaaaaaact catgtccagt tatccttgcc
       61 agtgcttcaa gtaagggatg tgctggtgcg gggtttcgga gacagtgtag aggaagtcct
      121 gtcagaggct agacagcacc tgaaggatgg gacctgtggc ttagtggaag ttgagaaggg
      181 agtcctcccg cagctggaac aaccttatgt gtttataaaa cgcagcgacg cgcgaacagc
      241 acctcacggg catgtaatgg tggagcttgt cgccgagttg gagggcatcc agtacggaag
      301 atcgggtgaa acgctgggag ttctagtgcc ccacgttggc gaaattccgg tggcctaccg
      361 taaggttttg ctcaggaaaa acggaaataa gggggctggt ggccatagct atggggcaga
      421 cctgaagtct ttcgatctgg gcgacgagct gggcaccgac ccatatgagg atttccagga
      481 aaactggaac acaaaacaca gctccggggt aactcgcgag ctaatgcgag aattgaatgg
      541 aggggaaaac ctttactttc agtcgggacg gcgaatcccg gggctgataa atccctggaa
      601 aaggagatgg aagaagaact ttattgcagt gtctgcggct aaccggttca aaaagatcag
      661 ttcaagcggc gccctagatt acgacattcc tacgaccgcc tccgagaatc tctatttcca
      721 gggtgaattt ggcttggcac aacatgacga ggccgtcgat aacaagttta ataaagaaca
      781 gcagaacgct ttctatgaga tccttcactt accaaatctg aacgaagagc aacgcaacgc
      841 tttcattcag agcctgaaag acgatccctc gcagtctgca aatctcctgg ccgaagccaa
      901 gaagctgaat gacgcacagg ctccaaaagt tgataacaag tttaacaagg agcagcagaa
      961 tgctttctac gagatattgc accttcctaa tctgaatgag gaacaaagaa atgcatttat
     1021 ccagtcccta aaagacgatc catcacagag tgccaacctg cttgcggaag ccaaaaagct
     1081 caatgatgcc caggccccga aagtagacgc gaaccatcaa taagctagct tgactgactg
     1141 agatacagcg taccttcagc tcacagacat gataagatac attgatgagt ttggacaaac
     1201 cacaactaga atgcagtgaa aaaaatgctt tatttgtgaa atttgtgatg ctattgcttt
     1261 atttgtaacc attataagct gcaataaaca agttaacaac aacaattgca ttcattttat
     1321 gtttcaggtt cagggggagg tgtgggaggt tttttaaagc aagtaaaacc tctacaaatg
     1381 tggtattggc ccatctctat cggtatcgta gcataacccc ttggggcctc taaacgggtc
     1441 ttgaggggtt ttttgtgccc ctcgggccgg attgctatct accggcattg gcgcagaaaa
     1501 aaatgcctga tgcgacgctg cgcgtcttat actcccacat atgccagatt cagcaacgga
     1561 tacggcttcc ccaacttgcc cacttccata cgtgtcctcc ttaccagaaa tttatcctta
     1621 aggtcgtcag ctatcctgca ggcgatctct cgatttcgat caagacattc ctttaatggt
     1681 cttttctgga caccactagg ggtcagaagt agttcatcaa actttcttcc ctccctaatc
     1741 tcattggtta ccttgggcta tcgaaactta attaaccagt caagtcagct acttggcgag
     1801 atcgacttgt ctgggtttcg actacgctca gaattgcgtc agtcaagttc gatctggtcc
     1861 ttgctattgc acccgttctc cgattacgag tttcatttaa atcatgtgag caaaaggcca
     1921 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc
     1981 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
     2041 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
     2101 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag
     2161 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca
     2221 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa
     2281 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc
     2341 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag
     2401 aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg
     2461 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca
     2521 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc
     2581 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag
     2641 gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata
     2701 tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat
     2761 ctgtctattt cgttcatcca tagttgcatt taaatttccg aactctccaa ggccctcgtc
     2821 ggaaaatctt caaacctttc gtccgatcca tcttgcaggc tacctctcga acgaactatc
     2881 gcaagtctct tggccggcct tgcgccttgg ctattgcttg gcagcgccta tcgccaggta
     2941 ttactccaat cccgaatatc cgagatcggg atcacccgag agaagttcaa cctacatcct
     3001 caatcccgat ctatccgaga tccgaggaat atcgaaatcg gggcgcgcct ggcctccgcg
     3061 ccgggttttg gcgcctcccg cgggcgcccc cctcgtcacg gcgagcgctg ccacgtcaga
     3121 cgaagggcgc aggagcgtcc tgatccttcc gcccggacgc tcaggacagc ggcccgctgc
     3181 tcataagact cggccttaga accccagtat cagcagaagg acattttagg acgggacttg
     3241 ggtgactcta gggcactggt tttctttcca gagagcggaa caggcgagga aaagtagtcc
     3301 cttctcggcg attctgcgga gggatctccg tggggcggtg aacgccgatg attatataag
     3361 gacgcgccgg gtgtggcaca gctagttccg tcgcagccgg gatttgggtc gcggttcttg
     3421 tttgtggatc gctgtgatcg tcacttggtg agtagcgggc tgctgggctg gccggggctt
     3481 tcgtggccgc cgggccgctc ggtgggacgg aagcgtgtgg agagaccgcc aagggctgta
     3541 gtctgggtcc gcgagcaagg ttgccctgaa ctgggggttg gggggagcgc agcaaaatgg
     3601 cggctgttcc cgagtcttga atggaagacg cttgtgaggc gggctgtgag gtcgttgaaa
     3661 caaggtgggg ggcatggtgg gcggcaagaa cccaaggtct tgagcccttc gctaatgcgg
     3721 gaaagctctt attcgggtga gatgggctgg gcaccatctg gggaccctga cgtgaagttt
     3781 gtcactgact ggagaactcg gtttgtcgtc tgttgcgggg gcggcagtta tggcggtgcc
     3841 gttgggcagt gcacccgtac ctttgggagc gcgcgccctc gtcgtgtcgt gacgtcaccc
     3901 gttctgttgg cttataatgc agggtggggc cacctgccgg taggtgtgcg gtaggctttt
     3961 ctccgtcgca ggacgcaggg ttcgggccta gggtaggctc tcctgaatcg acaggcgccg
     4021 gacctctggt gaggggaggg ataagtgagg cgtcagtttc tttggtcggt tttatgtacc
     4081 tatcttctta agtagctgaa gctccggttt tgaactatgc gctcggggtt ggcgagtgtg
     4141 ttttgtgaag ttttttaggc accttttgaa atgtaatcat ttgggtcaat atgtaatttt
     4201 cagtgttaga cttgtaaatt gtccgctaaa ttctggccgt ttttggcttt tttgttagac
     4261 aacatgaccg agtacaagcc cacggtgcgc ctcgccaccc gcgacgacgt ccccagggcc
     4321 gtacgcaccc tcgccgccgc gttcgccgac taccccgcca cgcgccacac cgtcgatccg
     4381 gaccgccaca tcgagcgggt caccgagctg caagaactct tcctcacgcg cgtcgggctc
     4441 gacatcggca aggtgtgggt cgcggacgac ggcgccgcgg tggcggtctg gaccacgccg
     4501 gagagcgtcg aagcgggggc ggtgttcgcc gagatcggcc cgcgcatggc cgagttgagc
     4561 ggttcccggc tggccgcgca gcaacagatg gagggcctcc tggcgccgca ccggcccaag
     4621 gagcccgcgt ggttcctggc caccgtcggc gtctcgcccg accaccaggg caagggtctg
     4681 ggcagcgccg tcgtgctccc cggagtggag gccgccgagc gcgccggggt gcccgccttc
     4741 ctggagacct ccgcgccccg caacctcccc ttctacgagc ggctcggctt caccgtcacc
     4801 gccgacgtcg aggtgcccga aggaccgcgc acctggtgca tgacccgcaa gcccggtgcc
     4861 taggctagta tgtaagccta gtcttagata ataaaatcgc tatccatcga agatggatgt
     4921 gtgttggttt tttgtgtgtg taacgctagg cgcgcctggt gtaccgagaa cgatcctctc
     4981 agtgcgagtc tcgacgatcc atatcgttgc ttggcagtca gccagtcgga atccagcttg
     5041 ggacccagga agtccaatcg tcagatattg tactcaagcc tggtcacggc agcgtaccga
     5101 tctgtttaaa cctagatatt gatagtctga tcggtcaacg tataatcgag tcctagcttt
     5161 tgcaaacatc tatcaagaga caggatcagc aggaggcttt cgcatgagta ttcaacattt
     5221 ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga
     5281 aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcgcgagtgg gttacatcga
     5341 actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac gctttccaat
     5401 gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca
     5461 agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt attcaccagt
     5521 cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac
     5581 catgagtgat aacactgcgg ccaacttact tctgacaacg attggaggac cgaaggagct
     5641 aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga
     5701 gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag caatggcaac
     5761 aaccttgcgt aaactattaa ctggcgaact acttactcta gcttcccggc aacagttgat
     5821 agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg
     5881 ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc
     5941 actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc
     6001 aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg
     6061 gtaaccgatt ctaggtgcat tggcgcagaa aaaaatgcct gatgcgacgc tgcgcgtctt
     6121 atactcccac atatgccaga ttcagcaacg gatacggctt ccccaacttg cccacttcca
     6181 tacgtgtcct ccttaccaga aatttatcct taagatcgtt taaactcgac tctggctcta
     6241 tcgaatctcc gtcgtttcga gcttacgcga acagccgtgg cgctcatttg ctcgtcgggc
     6301 atcgaatctc gtcagctatc gtcagcttac ctttttggca gcgatcgcgg ctcccgacat
     6361 cttggaccat tagctccaca ggtatcttct tccctctagt ggtcataaca gcagcttcag
     6421 ctacctctca attcaaaaaa cccctcaaga cccgtttaga ggccccaagg ggttatgcta
     6481 tcaatcgttg cgttacacac acaaaaaacc aacacacatc catcttcgat ggatagcgat
     6541 tttattatct aactgctgat cgagtgtagc cagatctagt aatcaattac ggggtcatta
     6601 gttcatagcc catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc
     6661 tgaccgccca acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg
     6721 ccaataggga ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg
     6781 gcagtacatc aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa
     6841 tggcccgcct ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac
     6901 atctacgtat tagtcatcgc tattaccatg ctgatgcggt tttggcagta catcaatggg
     6961 cgtggatagc ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg
     7021 agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca
     7081 ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctggttta
     7141 gtgaaccgtc agatcagatc tttgtcgatc ctaccatcca ctcgacacac ccgccagcgg
     7201 ccgc
//
