LOCUS       sequence-295540-        6727 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 6727)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 6727)
  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-m4134296
FEATURES             Location/Qualifiers
     source          1..6727
                     /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             566..643
                     /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"
     polyA_signal    696..817
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(707..726)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     761..780
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      932..978
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1008..1144
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1189..1260
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(1486..2074)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1566..1585)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2574..3783
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     3620..3639
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             3787..4386
                     /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(3787..3806)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4283..4303
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4413..4461
                     /note="synthetic polyadenylation signal"
     CDS             join(4727..4795,4796..5587)
                     /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(4945..4964)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      5600..5736
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    5881..5952
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(5957..6003)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    6021..6069
                     /note="synthetic polyadenylation signal"
     enhancer        6150..6453
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6455..6657
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     6608..6628
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      join(6726..6727,1..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 agtcgaagag gcgatggaaa aagaatttta tcgcggtgag
      601 tgcagccaac agattcaaaa agatatcctc gtctggtgcc ctctaagcta gcttgactga
      661 ctgagataca gcgtaccttc agctcacaga catgataaga tacattgatg agtttggaca
      721 aaccacaact agaatgcagt gaaaaaaatg ctttatttgt gaaatttgtg atgctattgc
      781 tttatttgta accattataa gctgcaataa acaagttaac aacaacaatt gcattcattt
      841 tatgtttcag gttcaggggg aggtgtggga ggttttttaa agcaagtaaa acctctacaa
      901 atgtggtatt ggcccatctc tatcggtatc gtagcataac cccttggggc ctctaaacgg
      961 gtcttgaggg gttttttgtg cccctcgggc cggattgcta tctaccggca ttggcgcaga
     1021 aaaaaatgcc tgatgcgacg ctgcgcgtct tatactccca catatgccag attcagcaac
     1081 ggatacggct tccccaactt gcccacttcc atacgtgtcc tccttaccag aaatttatcc
     1141 ttaaggtcgt cagctatcct gcaggcgatc tctcgatttc gatcaagaca ttcctttaat
     1201 ggtcttttct ggacaccact aggggtcaga agtagttcat caaactttct tccctcccta
     1261 atctcattgg ttaccttggg ctatcgaaac ttaattaacc agtcaagtca gctacttggc
     1321 gagatcgact tgtctgggtt tcgactacgc tcagaattgc gtcagtcaag ttcgatctgg
     1381 tccttgctat tgcacccgtt ctccgattac gagtttcatt taaatcatgt gagcaaaagg
     1441 ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg
     1501 cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg
     1561 actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac
     1621 cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca
     1681 tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt
     1741 gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc
     1801 caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag
     1861 agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac
     1921 tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt
     1981 tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa
     2041 gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg
     2101 gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa
     2161 aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat
     2221 atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc
     2281 gatctgtcta tttcgttcat ccatagttgc atttaaattt ccgaactctc caaggccctc
     2341 gtcggaaaat cttcaaacct ttcgtccgat ccatcttgca ggctacctct cgaacgaact
     2401 atcgcaagtc tcttggccgg ccttgcgcct tggctattgc ttggcagcgc ctatcgccag
     2461 gtattactcc aatcccgaat atccgagatc gggatcaccc gagagaagtt caacctacat
     2521 cctcaatccc gatctatccg agatccgagg aatatcgaaa tcggggcgcg cctggcctcc
     2581 gcgccgggtt ttggcgcctc ccgcgggcgc ccccctcgtc acggcgagcg ctgccacgtc
     2641 agacgaaggg cgcaggagcg tcctgatcct tccgcccgga cgctcaggac agcggcccgc
     2701 tgctcataag actcggcctt agaaccccag tatcagcaga aggacatttt aggacgggac
     2761 ttgggtgact ctagggcact ggttttcttt ccagagagcg gaacaggcga ggaaaagtag
     2821 tcccttctcg gcgattctgc ggagggatct ccgtggggcg gtgaacgccg atgattatat
     2881 aaggacgcgc cgggtgtggc acagctagtt ccgtcgcagc cgggatttgg gtcgcggttc
     2941 ttgtttgtgg atcgctgtga tcgtcacttg gtgagtagcg ggctgctggg ctggccgggg
     3001 ctttcgtggc cgccgggccg ctcggtggga cggaagcgtg tggagagacc gccaagggct
     3061 gtagtctggg tccgcgagca aggttgccct gaactggggg ttggggggag cgcagcaaaa
     3121 tggcggctgt tcccgagtct tgaatggaag acgcttgtga ggcgggctgt gaggtcgttg
     3181 aaacaaggtg gggggcatgg tgggcggcaa gaacccaagg tcttgagccc ttcgctaatg
     3241 cgggaaagct cttattcggg tgagatgggc tgggcaccat ctggggaccc tgacgtgaag
     3301 tttgtcactg actggagaac tcggtttgtc gtctgttgcg ggggcggcag ttatggcggt
     3361 gccgttgggc agtgcacccg tacctttggg agcgcgcgcc ctcgtcgtgt cgtgacgtca
     3421 cccgttctgt tggcttataa tgcagggtgg ggccacctgc cggtaggtgt gcggtaggct
     3481 tttctccgtc gcaggacgca gggttcgggc ctagggtagg ctctcctgaa tcgacaggcg
     3541 ccggacctct ggtgagggga gggataagtg aggcgtcagt ttctttggtc ggttttatgt
     3601 acctatcttc ttaagtagct gaagctccgg ttttgaacta tgcgctcggg gttggcgagt
     3661 gtgttttgtg aagtttttta ggcacctttt gaaatgtaat catttgggtc aatatgtaat
     3721 tttcagtgtt agacttgtaa attgtccgct aaattctggc cgtttttggc ttttttgtta
     3781 gacaacatga ccgagtacaa gcccacggtg cgcctcgcca cccgcgacga cgtccccagg
     3841 gccgtacgca ccctcgccgc cgcgttcgcc gactaccccg ccacgcgcca caccgtcgat
     3901 ccggaccgcc acatcgagcg ggtcaccgag ctgcaagaac tcttcctcac gcgcgtcggg
     3961 ctcgacatcg gcaaggtgtg ggtcgcggac gacggcgccg cggtggcggt ctggaccacg
     4021 ccggagagcg tcgaagcggg ggcggtgttc gccgagatcg gcccgcgcat ggccgagttg
     4081 agcggttccc ggctggccgc gcagcaacag atggagggcc tcctggcgcc gcaccggccc
     4141 aaggagcccg cgtggttcct ggccaccgtc ggcgtctcgc ccgaccacca gggcaagggt
     4201 ctgggcagcg ccgtcgtgct ccccggagtg gaggccgccg agcgcgccgg ggtgcccgcc
     4261 ttcctggaga cctccgcgcc ccgcaacctc cccttctacg agcggctcgg cttcaccgtc
     4321 accgccgacg tcgaggtgcc cgaaggaccg cgcacctggt gcatgacccg caagcccggt
     4381 gcctaggcta gtatgtaagc ctagtcttag ataataaaat cgctatccat cgaagatgga
     4441 tgtgtgttgg ttttttgtgt gtgtaacgct aggcgcgcct ggtgtaccga gaacgatcct
     4501 ctcagtgcga gtctcgacga tccatatcgt tgcttggcag tcagccagtc ggaatccagc
     4561 ttgggaccca ggaagtccaa tcgtcagata ttgtactcaa gcctggtcac ggcagcgtac
     4621 cgatctgttt aaacctagat attgatagtc tgatcggtca acgtataatc gagtcctagc
     4681 ttttgcaaac atctatcaag agacaggatc agcaggaggc tttcgcatga gtattcaaca
     4741 tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt ttgctcaccc
     4801 agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcgcgag tgggttacat
     4861 cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag aacgctttcc
     4921 aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta ttgacgccgg
     4981 gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg agtattcacc
     5041 agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca gtgctgccat
     5101 aaccatgagt gataacactg cggccaactt acttctgaca acgattggag gaccgaagga
     5161 gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc gttgggaacc
     5221 ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg tagcaatggc
     5281 aacaaccttg cgtaaactat taactggcga actacttact ctagcttccc ggcaacagtt
     5341 gatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg cccttccggc
     5401 tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg gtatcattgc
     5461 agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga cggggagtca
     5521 ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac tgattaagca
     5581 ttggtaaccg attctaggtg cattggcgca gaaaaaaatg cctgatgcga cgctgcgcgt
     5641 cttatactcc cacatatgcc agattcagca acggatacgg cttccccaac ttgcccactt
     5701 ccatacgtgt cctccttacc agaaatttat ccttaagatc gtttaaactc gactctggct
     5761 ctatcgaatc tccgtcgttt cgagcttacg cgaacagccg tggcgctcat ttgctcgtcg
     5821 ggcatcgaat ctcgtcagct atcgtcagct tacctttttg gcagcgatcg cggctcccga
     5881 catcttggac cattagctcc acaggtatct tcttccctct agtggtcata acagcagctt
     5941 cagctacctc tcaattcaaa aaacccctca agacccgttt agaggcccca aggggttatg
     6001 ctatcaatcg ttgcgttaca cacacaaaaa accaacacac atccatcttc gatggatagc
     6061 gattttatta tctaactgct gatcgagtgt agccagatct agtaatcaat tacggggtca
     6121 ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct
     6181 ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta
     6241 acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac
     6301 ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt
     6361 aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag
     6421 tacatctacg tattagtcat cgctattacc atgctgatgc ggttttggca gtacatcaat
     6481 gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat tgacgtcaat
     6541 gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa caactccgcc
     6601 ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag cagagctggt
     6661 ttagtgaacc gtcagatcag atctttgtcg atcctaccat ccactcgaca cacccgccag
     6721 cggccgc
//
