LOCUS       sequence-295256-        6694 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 6694)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 6694)
  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-m4134106
FEATURES             Location/Qualifiers
     source          1..6694
                     /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..610
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     polyA_signal    663..784
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(674..693)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     728..747
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      899..945
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      975..1111
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1156..1227
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(1453..2041)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1533..1552)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2541..3750
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     3587..3606
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             3754..4353
                     /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(3754..3773)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4250..4270
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4380..4428
                     /note="synthetic polyadenylation signal"
     CDS             join(4694..4762,4763..5554)
                     /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(4912..4931)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      5567..5703
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    5848..5919
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(5924..5970)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    5988..6036
                     /note="synthetic polyadenylation signal"
     enhancer        6117..6420
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6422..6624
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     6575..6595
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      join(6693..6694,1..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 agtcgaaaga aaccgcggca gctaagttcg agaggcagca
      601 catggacagt taagctagct tgactgactg agatacagcg taccttcagc tcacagacat
      661 gataagatac attgatgagt ttggacaaac cacaactaga atgcagtgaa aaaaatgctt
      721 tatttgtgaa atttgtgatg ctattgcttt atttgtaacc attataagct gcaataaaca
      781 agttaacaac aacaattgca ttcattttat gtttcaggtt cagggggagg tgtgggaggt
      841 tttttaaagc aagtaaaacc tctacaaatg tggtattggc ccatctctat cggtatcgta
      901 gcataacccc ttggggcctc taaacgggtc ttgaggggtt ttttgtgccc ctcgggccgg
      961 attgctatct accggcattg gcgcagaaaa aaatgcctga tgcgacgctg cgcgtcttat
     1021 actcccacat atgccagatt cagcaacgga tacggcttcc ccaacttgcc cacttccata
     1081 cgtgtcctcc ttaccagaaa tttatcctta aggtcgtcag ctatcctgca ggcgatctct
     1141 cgatttcgat caagacattc ctttaatggt cttttctgga caccactagg ggtcagaagt
     1201 agttcatcaa actttcttcc ctccctaatc tcattggtta ccttgggcta tcgaaactta
     1261 attaaccagt caagtcagct acttggcgag atcgacttgt ctgggtttcg actacgctca
     1321 gaattgcgtc agtcaagttc gatctggtcc ttgctattgc acccgttctc cgattacgag
     1381 tttcatttaa atcatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
     1441 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
     1501 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
     1561 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     1621 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
     1681 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     1741 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     1801 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     1861 gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct
     1921 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     1981 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca
     2041 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
     2101 agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa
     2161 atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg
     2221 cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcatt
     2281 taaatttccg aactctccaa ggccctcgtc ggaaaatctt caaacctttc gtccgatcca
     2341 tcttgcaggc tacctctcga acgaactatc gcaagtctct tggccggcct tgcgccttgg
     2401 ctattgcttg gcagcgccta tcgccaggta ttactccaat cccgaatatc cgagatcggg
     2461 atcacccgag agaagttcaa cctacatcct caatcccgat ctatccgaga tccgaggaat
     2521 atcgaaatcg gggcgcgcct ggcctccgcg ccgggttttg gcgcctcccg cgggcgcccc
     2581 cctcgtcacg gcgagcgctg ccacgtcaga cgaagggcgc aggagcgtcc tgatccttcc
     2641 gcccggacgc tcaggacagc ggcccgctgc tcataagact cggccttaga accccagtat
     2701 cagcagaagg acattttagg acgggacttg ggtgactcta gggcactggt tttctttcca
     2761 gagagcggaa caggcgagga aaagtagtcc cttctcggcg attctgcgga gggatctccg
     2821 tggggcggtg aacgccgatg attatataag gacgcgccgg gtgtggcaca gctagttccg
     2881 tcgcagccgg gatttgggtc gcggttcttg tttgtggatc gctgtgatcg tcacttggtg
     2941 agtagcgggc tgctgggctg gccggggctt tcgtggccgc cgggccgctc ggtgggacgg
     3001 aagcgtgtgg agagaccgcc aagggctgta gtctgggtcc gcgagcaagg ttgccctgaa
     3061 ctgggggttg gggggagcgc agcaaaatgg cggctgttcc cgagtcttga atggaagacg
     3121 cttgtgaggc gggctgtgag gtcgttgaaa caaggtgggg ggcatggtgg gcggcaagaa
     3181 cccaaggtct tgagcccttc gctaatgcgg gaaagctctt attcgggtga gatgggctgg
     3241 gcaccatctg gggaccctga cgtgaagttt gtcactgact ggagaactcg gtttgtcgtc
     3301 tgttgcgggg gcggcagtta tggcggtgcc gttgggcagt gcacccgtac ctttgggagc
     3361 gcgcgccctc gtcgtgtcgt gacgtcaccc gttctgttgg cttataatgc agggtggggc
     3421 cacctgccgg taggtgtgcg gtaggctttt ctccgtcgca ggacgcaggg ttcgggccta
     3481 gggtaggctc tcctgaatcg acaggcgccg gacctctggt gaggggaggg ataagtgagg
     3541 cgtcagtttc tttggtcggt tttatgtacc tatcttctta agtagctgaa gctccggttt
     3601 tgaactatgc gctcggggtt ggcgagtgtg ttttgtgaag ttttttaggc accttttgaa
     3661 atgtaatcat ttgggtcaat atgtaatttt cagtgttaga cttgtaaatt gtccgctaaa
     3721 ttctggccgt ttttggcttt tttgttagac aacatgaccg agtacaagcc cacggtgcgc
     3781 ctcgccaccc gcgacgacgt ccccagggcc gtacgcaccc tcgccgccgc gttcgccgac
     3841 taccccgcca cgcgccacac cgtcgatccg gaccgccaca tcgagcgggt caccgagctg
     3901 caagaactct tcctcacgcg cgtcgggctc gacatcggca aggtgtgggt cgcggacgac
     3961 ggcgccgcgg tggcggtctg gaccacgccg gagagcgtcg aagcgggggc ggtgttcgcc
     4021 gagatcggcc cgcgcatggc cgagttgagc ggttcccggc tggccgcgca gcaacagatg
     4081 gagggcctcc tggcgccgca ccggcccaag gagcccgcgt ggttcctggc caccgtcggc
     4141 gtctcgcccg accaccaggg caagggtctg ggcagcgccg tcgtgctccc cggagtggag
     4201 gccgccgagc gcgccggggt gcccgccttc ctggagacct ccgcgccccg caacctcccc
     4261 ttctacgagc ggctcggctt caccgtcacc gccgacgtcg aggtgcccga aggaccgcgc
     4321 acctggtgca tgacccgcaa gcccggtgcc taggctagta tgtaagccta gtcttagata
     4381 ataaaatcgc tatccatcga agatggatgt gtgttggttt tttgtgtgtg taacgctagg
     4441 cgcgcctggt gtaccgagaa cgatcctctc agtgcgagtc tcgacgatcc atatcgttgc
     4501 ttggcagtca gccagtcgga atccagcttg ggacccagga agtccaatcg tcagatattg
     4561 tactcaagcc tggtcacggc agcgtaccga tctgtttaaa cctagatatt gatagtctga
     4621 tcggtcaacg tataatcgag tcctagcttt tgcaaacatc tatcaagaga caggatcagc
     4681 aggaggcttt cgcatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc
     4741 attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga
     4801 tcagttgggt gcgcgagtgg gttacatcga actggatctc aacagcggta agatccttga
     4861 gagttttcgc cccgaagaac gctttccaat gatgagcact tttaaagttc tgctatgtgg
     4921 cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc
     4981 tcagaatgac ttggttgagt attcaccagt cacagaaaag catcttacgg atggcatgac
     5041 agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact
     5101 tctgacaacg attggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca
     5161 tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg
     5221 tgacaccacg atgcctgtag caatggcaac aaccttgcgt aaactattaa ctggcgaact
     5281 acttactcta gcttcccggc aacagttgat agactggatg gaggcggata aagttgcagg
     5341 accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg
     5401 tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat
     5461 cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc
     5521 tgagataggt gcctcactga ttaagcattg gtaaccgatt ctaggtgcat tggcgcagaa
     5581 aaaaatgcct gatgcgacgc tgcgcgtctt atactcccac atatgccaga ttcagcaacg
     5641 gatacggctt ccccaacttg cccacttcca tacgtgtcct ccttaccaga aatttatcct
     5701 taagatcgtt taaactcgac tctggctcta tcgaatctcc gtcgtttcga gcttacgcga
     5761 acagccgtgg cgctcatttg ctcgtcgggc atcgaatctc gtcagctatc gtcagcttac
     5821 ctttttggca gcgatcgcgg ctcccgacat cttggaccat tagctccaca ggtatcttct
     5881 tccctctagt ggtcataaca gcagcttcag ctacctctca attcaaaaaa cccctcaaga
     5941 cccgtttaga ggccccaagg ggttatgcta tcaatcgttg cgttacacac acaaaaaacc
     6001 aacacacatc catcttcgat ggatagcgat tttattatct aactgctgat cgagtgtagc
     6061 cagatctagt aatcaattac ggggtcatta gttcatagcc catatatgga gttccgcgtt
     6121 acataactta cggtaaatgg cccgcctggc tgaccgccca acgacccccg cccattgacg
     6181 tcaataatga cgtatgttcc catagtaacg ccaataggga ctttccattg acgtcaatgg
     6241 gtggagtatt tacggtaaac tgcccacttg gcagtacatc aagtgtatca tatgccaagt
     6301 acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc ccagtacatg
     6361 accttatggg actttcctac ttggcagtac atctacgtat tagtcatcgc tattaccatg
     6421 ctgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt
     6481 ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac
     6541 tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg
     6601 tgggaggtct atataagcag agctggttta gtgaaccgtc agatcagatc tttgtcgatc
     6661 ctaccatcca ctcgacacac ccgccagcgg ccgc
//
