LOCUS       sequence-295900-        7201 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 7201)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 7201)
  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-m4133743
FEATURES             Location/Qualifiers
     source          1..7201
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             542..562
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             596..673
                     /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             701..721
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     polyA_signal    1170..1291
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1181..1200)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1235..1254
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      1406..1452
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1482..1618
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1663..1734
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(1960..2548)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2040..2059)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        3048..4257
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     4094..4113
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             4261..4860
                     /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(4261..4280)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4757..4777
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4887..4935
                     /note="synthetic polyadenylation signal"
     CDS             join(5201..5269,5270..6061)
                     /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(5419..5438)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      6074..6210
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    6355..6426
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(6431..6477)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    6495..6543
                     /note="synthetic polyadenylation signal"
     enhancer        6624..6927
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6929..7131
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     7082..7102
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      join(7200..7201,1..8)
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
ORIGIN
        1 caccatggag agccttgtcc ctggtttcaa cgagaaaaca cacgtccaac tcagtttgcc
       61 tgttttacag gttcgcgacg tgctcgtacg tggctttgga gactccgtgg aggaggtctt
      121 atcagaggca cgtcaacatc ttaaagatgg cacttgtggc ttagtagaag ttgaaaaagg
      181 cgttttgcct caacttgaac agccctatgt gttcatcaaa cgttcggatg ctcgaactgc
      241 acctcatggt catgttatgg ttgagctggt agcagaactc gaaggcattc agtacggtcg
      301 tagtggtgag acacttggtg tccttgtccc tcatgtgggc gaaataccag tggcttaccg
      361 caaggttctt cttcgtaaga acggtaataa aggagctggt ggccatagtt acggcgccga
      421 tctaaagtca tttgacttag gcgacgagct tggcactgat ccttatgaag attttcaaga
      481 aaactggaac actaaacata gcagtggtgt tacccgtgaa ctcatgcgtg agcttaacgg
      541 agaaaacctt tactttcagt cgggacggcg aatcccgggg ctgataaatc cctggaaaag
      601 gagatggaag aagaacttta ttgcagtgtc tgcggctaac cggttcaaaa agatcagttc
      661 aagcggcgcc ctagattacg acattcctac gaccgcctcc gagaatctct atttccaggg
      721 tgaatttggc ttggcacaac atgacgaggc cgtcgataac aagtttaata aagaacagca
      781 gaacgctttc tatgagatcc ttcacttacc aaatctgaac gaagagcaac gcaacgcttt
      841 cattcagagc ctgaaagacg atccctcgca gtctgcaaat ctcctggccg aagccaagaa
      901 gctgaatgac gcacaggctc caaaagttga taacaagttt aacaaggagc agcagaatgc
      961 tttctacgag atattgcacc ttcctaatct gaatgaggaa caaagaaatg catttatcca
     1021 gtccctaaaa gacgatccat cacagagtgc caacctgctt gcggaagcca aaaagctcaa
     1081 tgatgcccag gccccgaaag tagacgcgaa ccatcaataa gctagcttga ctgactgaga
     1141 tacagcgtac cttcagctca cagacatgat aagatacatt gatgagtttg gacaaaccac
     1201 aactagaatg cagtgaaaaa aatgctttat ttgtgaaatt tgtgatgcta ttgctttatt
     1261 tgtaaccatt ataagctgca ataaacaagt taacaacaac aattgcattc attttatgtt
     1321 tcaggttcag ggggaggtgt gggaggtttt ttaaagcaag taaaacctct acaaatgtgg
     1381 tattggccca tctctatcgg tatcgtagca taaccccttg gggcctctaa acgggtcttg
     1441 aggggttttt tgtgcccctc gggccggatt gctatctacc ggcattggcg cagaaaaaaa
     1501 tgcctgatgc gacgctgcgc gtcttatact cccacatatg ccagattcag caacggatac
     1561 ggcttcccca acttgcccac ttccatacgt gtcctcctta ccagaaattt atccttaagg
     1621 tcgtcagcta tcctgcaggc gatctctcga tttcgatcaa gacattcctt taatggtctt
     1681 ttctggacac cactaggggt cagaagtagt tcatcaaact ttcttccctc cctaatctca
     1741 ttggttacct tgggctatcg aaacttaatt aaccagtcaa gtcagctact tggcgagatc
     1801 gacttgtctg ggtttcgact acgctcagaa ttgcgtcagt caagttcgat ctggtccttg
     1861 ctattgcacc cgttctccga ttacgagttt catttaaatc atgtgagcaa aaggccagca
     1921 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc
     1981 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata
     2041 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc
     2101 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc
     2161 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga
     2221 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc
     2281 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag
     2341 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag
     2401 aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag
     2461 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca
     2521 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga
     2581 cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat
     2641 cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga
     2701 gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg
     2761 tctatttcgt tcatccatag ttgcatttaa atttccgaac tctccaaggc cctcgtcgga
     2821 aaatcttcaa acctttcgtc cgatccatct tgcaggctac ctctcgaacg aactatcgca
     2881 agtctcttgg ccggccttgc gccttggcta ttgcttggca gcgcctatcg ccaggtatta
     2941 ctccaatccc gaatatccga gatcgggatc acccgagaga agttcaacct acatcctcaa
     3001 tcccgatcta tccgagatcc gaggaatatc gaaatcgggg cgcgcctggc ctccgcgccg
     3061 ggttttggcg cctcccgcgg gcgcccccct cgtcacggcg agcgctgcca cgtcagacga
     3121 agggcgcagg agcgtcctga tccttccgcc cggacgctca ggacagcggc ccgctgctca
     3181 taagactcgg ccttagaacc ccagtatcag cagaaggaca ttttaggacg ggacttgggt
     3241 gactctaggg cactggtttt ctttccagag agcggaacag gcgaggaaaa gtagtccctt
     3301 ctcggcgatt ctgcggaggg atctccgtgg ggcggtgaac gccgatgatt atataaggac
     3361 gcgccgggtg tggcacagct agttccgtcg cagccgggat ttgggtcgcg gttcttgttt
     3421 gtggatcgct gtgatcgtca cttggtgagt agcgggctgc tgggctggcc ggggctttcg
     3481 tggccgccgg gccgctcggt gggacggaag cgtgtggaga gaccgccaag ggctgtagtc
     3541 tgggtccgcg agcaaggttg ccctgaactg ggggttgggg ggagcgcagc aaaatggcgg
     3601 ctgttcccga gtcttgaatg gaagacgctt gtgaggcggg ctgtgaggtc gttgaaacaa
     3661 ggtggggggc atggtgggcg gcaagaaccc aaggtcttga gcccttcgct aatgcgggaa
     3721 agctcttatt cgggtgagat gggctgggca ccatctgggg accctgacgt gaagtttgtc
     3781 actgactgga gaactcggtt tgtcgtctgt tgcgggggcg gcagttatgg cggtgccgtt
     3841 gggcagtgca cccgtacctt tgggagcgcg cgccctcgtc gtgtcgtgac gtcacccgtt
     3901 ctgttggctt ataatgcagg gtggggccac ctgccggtag gtgtgcggta ggcttttctc
     3961 cgtcgcagga cgcagggttc gggcctaggg taggctctcc tgaatcgaca ggcgccggac
     4021 ctctggtgag gggagggata agtgaggcgt cagtttcttt ggtcggtttt atgtacctat
     4081 cttcttaagt agctgaagct ccggttttga actatgcgct cggggttggc gagtgtgttt
     4141 tgtgaagttt tttaggcacc ttttgaaatg taatcatttg ggtcaatatg taattttcag
     4201 tgttagactt gtaaattgtc cgctaaattc tggccgtttt tggctttttt gttagacaac
     4261 atgaccgagt acaagcccac ggtgcgcctc gccacccgcg acgacgtccc cagggccgta
     4321 cgcaccctcg ccgccgcgtt cgccgactac cccgccacgc gccacaccgt cgatccggac
     4381 cgccacatcg agcgggtcac cgagctgcaa gaactcttcc tcacgcgcgt cgggctcgac
     4441 atcggcaagg tgtgggtcgc ggacgacggc gccgcggtgg cggtctggac cacgccggag
     4501 agcgtcgaag cgggggcggt gttcgccgag atcggcccgc gcatggccga gttgagcggt
     4561 tcccggctgg ccgcgcagca acagatggag ggcctcctgg cgccgcaccg gcccaaggag
     4621 cccgcgtggt tcctggccac cgtcggcgtc tcgcccgacc accagggcaa gggtctgggc
     4681 agcgccgtcg tgctccccgg agtggaggcc gccgagcgcg ccggggtgcc cgccttcctg
     4741 gagacctccg cgccccgcaa cctccccttc tacgagcggc tcggcttcac cgtcaccgcc
     4801 gacgtcgagg tgcccgaagg accgcgcacc tggtgcatga cccgcaagcc cggtgcctag
     4861 gctagtatgt aagcctagtc ttagataata aaatcgctat ccatcgaaga tggatgtgtg
     4921 ttggtttttt gtgtgtgtaa cgctaggcgc gcctggtgta ccgagaacga tcctctcagt
     4981 gcgagtctcg acgatccata tcgttgcttg gcagtcagcc agtcggaatc cagcttggga
     5041 cccaggaagt ccaatcgtca gatattgtac tcaagcctgg tcacggcagc gtaccgatct
     5101 gtttaaacct agatattgat agtctgatcg gtcaacgtat aatcgagtcc tagcttttgc
     5161 aaacatctat caagagacag gatcagcagg aggctttcgc atgagtattc aacatttccg
     5221 tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc acccagaaac
     5281 gctggtgaaa gtaaaagatg ctgaagatca gttgggtgcg cgagtgggtt acatcgaact
     5341 ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgct ttccaatgat
     5401 gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga
     5461 gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtatt caccagtcac
     5521 agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg ccataaccat
     5581 gagtgataac actgcggcca acttacttct gacaacgatt ggaggaccga aggagctaac
     5641 cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct
     5701 gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgtagcaa tggcaacaac
     5761 cttgcgtaaa ctattaactg gcgaactact tactctagct tcccggcaac agttgataga
     5821 ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg
     5881 gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca ttgcagcact
     5941 ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac
     6001 tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta
     6061 accgattcta ggtgcattgg cgcagaaaaa aatgcctgat gcgacgctgc gcgtcttata
     6121 ctcccacata tgccagattc agcaacggat acggcttccc caacttgccc acttccatac
     6181 gtgtcctcct taccagaaat ttatccttaa gatcgtttaa actcgactct ggctctatcg
     6241 aatctccgtc gtttcgagct tacgcgaaca gccgtggcgc tcatttgctc gtcgggcatc
     6301 gaatctcgtc agctatcgtc agcttacctt tttggcagcg atcgcggctc ccgacatctt
     6361 ggaccattag ctccacaggt atcttcttcc ctctagtggt cataacagca gcttcagcta
     6421 cctctcaatt caaaaaaccc ctcaagaccc gtttagaggc cccaaggggt tatgctatca
     6481 atcgttgcgt tacacacaca aaaaaccaac acacatccat cttcgatgga tagcgatttt
     6541 attatctaac tgctgatcga gtgtagccag atctagtaat caattacggg gtcattagtt
     6601 catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga
     6661 ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca
     6721 atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca
     6781 gtacatcaag tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg
     6841 cccgcctggc attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc
     6901 tacgtattag tcatcgctat taccatgctg atgcggtttt ggcagtacat caatgggcgt
     6961 ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt
     7021 ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg
     7081 acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagc tggtttagtg
     7141 aaccgtcaga tcagatcttt gtcgatccta ccatccactc gacacacccg ccagcggccg
     7201 c
//
