LOCUS       sequence-295756-        7457 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression plasmid for SARS-CoV-2 ORF3a protein.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7457)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 7457)
  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-m4134239
FEATURES             Location/Qualifiers
     source          1..7457
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             830..850
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             851..928
                     /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    997..1118
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1008..1027)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1062..1081
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      1233..1279
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1309..1445
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1490..1561
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(1787..2375)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1867..1886)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2875..4084
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     3921..3940
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             4088..5116
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     polyA_signal    5143..5191
                     /note="synthetic polyadenylation signal"
     CDS             join(5457..5525,5526..6317)
                     /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(5675..5694)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      6330..6466
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    6611..6682
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(6687..6733)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    6751..6799
                     /note="synthetic polyadenylation signal"
     enhancer        6880..7183
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        7185..7387
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     7338..7358
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      join(7456..7457,1..8)
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
ORIGIN
        1 caccatggat ctgtttatgc gtatattcac catcggcacg gttacattga agcaagggga
       61 aattaaagac gcaactccca gtgacttcgt acgggcgacc gctacaattc cgatccaggc
      121 cagcctccct tttggatggc ttatcgtcgg cgtggcccta ctggctgtgt ttcagtcggc
      181 atcaaagata atcactctga aaaagcgatg gcagttagcc cttagcaaag gtgtgcactt
      241 cgtatgcaat ttgctgctcc tgttcgtcac cgtttattcc catctgctcc tggtggctgc
      301 cggattggag gcaccatttt tatacctata cgccctcgtg tattttctgc aatctattaa
      361 cttcgtccgc attatcatga gactctggct ctgttggaag tgcaggtcta aaaatccctt
      421 gttatatgat gccaactact tcctgtgttg gcacactaac tgctatgact actgtattcc
      481 atacaattca gttacctcca gtatagtgat cacatctggg gatggtacga cttcgcctat
      541 ttccgagcat gactatcaga tcggcgggta cacagaaaag tgggagagcg gagtcaagga
      601 ttgcgtggtg cttcactcct atttcaccag tgactactac cagctttata gcacacaact
      661 ttcaacagac accggcgtgg agcatgtgac ttttttcatt tataataaaa tcgttgatga
      721 accggaggaa cacgttcaga tacacactat cgacggcagc tctggtgtcg tcaaccctgt
      781 tatggagccc atttacgatg aaccaacgac cacaacatct gtacctctgg aaaaccttta
      841 ctttcagtcg aagaggcgat ggaaaaagaa ttttatcgcg gtgagtgcag ccaacagatt
      901 caaaaagata tcctcgtctg gtgccctcta aggatccgca ggcctctgct agcttgactg
      961 actgagatac agcgtacctt cagctcacag acatgataag atacattgat gagtttggac
     1021 aaaccacaac tagaatgcag tgaaaaaaat gctttatttg tgaaatttgt gatgctattg
     1081 ctttatttgt aaccattata agctgcaata aacaagttaa caacaacaat tgcattcatt
     1141 ttatgtttca ggttcagggg gaggtgtggg aggtttttta aagcaagtaa aacctctaca
     1201 aatgtggtat tggcccatct ctatcggtat cgtagcataa ccccttgggg cctctaaacg
     1261 ggtcttgagg ggttttttgt gcccctcggg ccggattgct atctaccggc attggcgcag
     1321 aaaaaaatgc ctgatgcgac gctgcgcgtc ttatactccc acatatgcca gattcagcaa
     1381 cggatacggc ttccccaact tgcccacttc catacgtgtc ctccttacca gaaatttatc
     1441 cttaaggtcg tcagctatcc tgcaggcgat ctctcgattt cgatcaagac attcctttaa
     1501 tggtcttttc tggacaccac taggggtcag aagtagttca tcaaactttc ttccctccct
     1561 aatctcattg gttaccttgg gctatcgaaa cttaattaac cagtcaagtc agctacttgg
     1621 cgagatcgac ttgtctgggt ttcgactacg ctcagaattg cgtcagtcaa gttcgatctg
     1681 gtccttgcta ttgcacccgt tctccgatta cgagtttcat ttaaatcatg tgagcaaaag
     1741 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc
     1801 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
     1861 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
     1921 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
     1981 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
     2041 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
     2101 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
     2161 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
     2221 ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
     2281 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
     2341 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg
     2401 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa
     2461 aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta
     2521 tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag
     2581 cgatctgtct atttcgttca tccatagttg catttaaatt tccgaactct ccaaggccct
     2641 cgtcggaaaa tcttcaaacc tttcgtccga tccatcttgc aggctacctc tcgaacgaac
     2701 tatcgcaagt ctcttggccg gccttgcgcc ttggctattg cttggcagcg cctatcgcca
     2761 ggtattactc caatcccgaa tatccgagat cgggatcacc cgagagaagt tcaacctaca
     2821 tcctcaatcc cgatctatcc gagatccgag gaatatcgaa atcggggcgc gcctggcctc
     2881 cgcgccgggt tttggcgcct cccgcgggcg cccccctcgt cacggcgagc gctgccacgt
     2941 cagacgaagg gcgcaggagc gtcctgatcc ttccgcccgg acgctcagga cagcggcccg
     3001 ctgctcataa gactcggcct tagaacccca gtatcagcag aaggacattt taggacggga
     3061 cttgggtgac tctagggcac tggttttctt tccagagagc ggaacaggcg aggaaaagta
     3121 gtcccttctc ggcgattctg cggagggatc tccgtggggc ggtgaacgcc gatgattata
     3181 taaggacgcg ccgggtgtgg cacagctagt tccgtcgcag ccgggatttg ggtcgcggtt
     3241 cttgtttgtg gatcgctgtg atcgtcactt ggtgagtagc gggctgctgg gctggccggg
     3301 gctttcgtgg ccgccgggcc gctcggtggg acggaagcgt gtggagagac cgccaagggc
     3361 tgtagtctgg gtccgcgagc aaggttgccc tgaactgggg gttgggggga gcgcagcaaa
     3421 atggcggctg ttcccgagtc ttgaatggaa gacgcttgtg aggcgggctg tgaggtcgtt
     3481 gaaacaaggt ggggggcatg gtgggcggca agaacccaag gtcttgagcc cttcgctaat
     3541 gcgggaaagc tcttattcgg gtgagatggg ctgggcacca tctggggacc ctgacgtgaa
     3601 gtttgtcact gactggagaa ctcggtttgt cgtctgttgc gggggcggca gttatggcgg
     3661 tgccgttggg cagtgcaccc gtacctttgg gagcgcgcgc cctcgtcgtg tcgtgacgtc
     3721 acccgttctg ttggcttata atgcagggtg gggccacctg ccggtaggtg tgcggtaggc
     3781 ttttctccgt cgcaggacgc agggttcggg cctagggtag gctctcctga atcgacaggc
     3841 gccggacctc tggtgagggg agggataagt gaggcgtcag tttctttggt cggttttatg
     3901 tacctatctt cttaagtagc tgaagctccg gttttgaact atgcgctcgg ggttggcgag
     3961 tgtgttttgt gaagtttttt aggcaccttt tgaaatgtaa tcatttgggt caatatgtaa
     4021 ttttcagtgt tagacttgta aattgtccgc taaattctgg ccgtttttgg cttttttgtt
     4081 agacaacatg ggtaaaaagc ctgaactcac cgcgacgtct gtcgagaagt ttctgatcga
     4141 aaagttcgac agcgtctccg acctgatgca gctctcggag ggcgaagaat ctcgtgcttt
     4201 cagcttcgat gtaggagggc gtggatatgt cctgcgggta aatagctgcg ccgatggttt
     4261 ctacaaagat cgttatgttt atcggcactt tgcatccgcc gcgctcccga ttccggaagt
     4321 gcttgacatt ggggagttca gcgagagcct gacctattgc atctcccgcc gtgcacaggg
     4381 tgtcacgttg caagacctgc ctgaaaccga actgcccgct gttctgcagc cggtcgcgga
     4441 ggcaatggat gccatcgctg ccgccgatct tagccagacg agcgggttcg gcccattcgg
     4501 accgcaagga atcggtcaat acactacatg gcgtgatttc atatgcgcga ttgctgatcc
     4561 ccatgtgtat cactggcaaa ctgtgatgga cgacaccgtc agtgcgtccg tcgcgcaggc
     4621 tctcgatgag ctgatgcttt gggccgagga ctgccccgaa gtccggcacc tcgtgcacgc
     4681 ggatttcggc tccaacaatg tcctgacgga caatggccgc ataacagcgg tcattgactg
     4741 gagcgaggcg atgttcgggg attcccaata cgaggtcgcc aacatcttct tctggaggcc
     4801 gtggttggct tgtatggagc agcagacgcg ctacttcgag cggaggcatc cggagcttgc
     4861 aggatcgccc cggctccggg cgtatatgct ccgcattggt cttgaccaac tctatcagag
     4921 cttggttgac ggcaatttcg atgatgcagc ttgggcgcag ggtcgatgcg acgcaatcgt
     4981 ccgatccgga gccgggactg tcgggcgtac acaaatcgcc cgcagaagcg ccgccgtctg
     5041 gaccgatggc tgtgtagaag tactcgccga tagtggaaac cgacgcccca gcactcgtcc
     5101 gagggcaaag gaataagcta gtatgtaagc ctagtcttag ataataaaat cgctatccat
     5161 cgaagatgga tgtgtgttgg ttttttgtgt gtgtaacgct aggcgcgcct ggtgtaccga
     5221 gaacgatcct ctcagtgcga gtctcgacga tccatatcgt tgcttggcag tcagccagtc
     5281 ggaatccagc ttgggaccca ggaagtccaa tcgtcagata ttgtactcaa gcctggtcac
     5341 ggcagcgtac cgatctgttt aaacctagat attgatagtc tgatcggtca acgtataatc
     5401 gagtcctagc ttttgcaaac atctatcaag agacaggatc agcaggaggc tttcgcatga
     5461 gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt
     5521 ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcgcgag
     5581 tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag
     5641 aacgctttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta
     5701 ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg
     5761 agtattcacc agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca
     5821 gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca acgattggag
     5881 gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc
     5941 gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg
     6001 tagcaatggc aacaaccttg cgtaaactat taactggcga actacttact ctagcttccc
     6061 ggcaacagtt gatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg
     6121 cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg
     6181 gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga
     6241 cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac
     6301 tgattaagca ttggtaaccg attctaggtg cattggcgca gaaaaaaatg cctgatgcga
     6361 cgctgcgcgt cttatactcc cacatatgcc agattcagca acggatacgg cttccccaac
     6421 ttgcccactt ccatacgtgt cctccttacc agaaatttat ccttaagatc gtttaaactc
     6481 gactctggct ctatcgaatc tccgtcgttt cgagcttacg cgaacagccg tggcgctcat
     6541 ttgctcgtcg ggcatcgaat ctcgtcagct atcgtcagct tacctttttg gcagcgatcg
     6601 cggctcccga catcttggac cattagctcc acaggtatct tcttccctct agtggtcata
     6661 acagcagctt cagctacctc tcaattcaaa aaacccctca agacccgttt agaggcccca
     6721 aggggttatg ctatcaatcg ttgcgttaca cacacaaaaa accaacacac atccatcttc
     6781 gatggatagc gattttatta tctaactgct gatcgagtgt agccagatct agtaatcaat
     6841 tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa
     6901 tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt
     6961 tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta
     7021 aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt
     7081 caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc
     7141 tacttggcag tacatctacg tattagtcat cgctattacc atgctgatgc ggttttggca
     7201 gtacatcaat gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat
     7261 tgacgtcaat gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa
     7321 caactccgcc ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag
     7381 cagagctggt ttagtgaacc gtcagatcag atctttgtcg atcctaccat ccactcgaca
     7441 cacccgccag cggccgc
//
