LOCUS       sequence-295943-        7397 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 7397)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 7397)
  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-m4133673
FEATURES             Location/Qualifiers
     source          1..7397
                     /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..868
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     polyA_signal    937..1058
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(948..967)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1002..1021
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      1173..1219
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1249..1385
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1430..1501
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(1727..2315)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1807..1826)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2815..4024
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     3861..3880
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             4028..5056
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     polyA_signal    5083..5131
                     /note="synthetic polyadenylation signal"
     CDS             join(5397..5465,5466..6257)
                     /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(5615..5634)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      6270..6406
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    6551..6622
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(6627..6673)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    6691..6739
                     /note="synthetic polyadenylation signal"
     enhancer        6820..7123
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        7125..7327
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     7278..7298
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      join(7396..7397,1..8)
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
ORIGIN
        1 caccatggat ttgtttatga gaatcttcac aattggaact gtaactttga agcaaggtga
       61 aatcaaggat gctactcctt cagattttgt tcgcgctact gcaacgatac cgatacaagc
      121 ctcactccct ttcggatggc ttattgttgg cgttgcactt cttgctgttt ttcagagcgc
      181 ttccaaaatc ataaccctca aaaagagatg gcaactagca ctctccaagg gtgttcactt
      241 tgtttgcaac ttgctgttgt tgtttgtaac agtttactca caccttttgc tcgttgctgc
      301 tggccttgaa gccccttttc tctatcttta tgctttagtc tacttcttgc agagtataaa
      361 ctttgtaaga ataataatga ggctttggct ttgctggaaa tgccgttcca aaaacccatt
      421 actttatgat gccaactatt ttctttgctg gcatactaat tgttacgact attgtatacc
      481 ttacaatagt gtaacttctt caattgtcat tacttcaggt gatggcacaa caagtcctat
      541 ttctgaacat gactaccaga ttggtggtta tactgaaaaa tgggaatctg gagtaaaaga
      601 ctgtgttgta ttacacagtt acttcacttc agactattac cagctgtact caactcaatt
      661 gagtacagac actggtgttg aacatgttac cttcttcatc tacaataaaa ttgttgatga
      721 gcctgaagaa catgtccaaa ttcacacaat cgacggttca tccggagttg ttaatccagt
      781 aatggaacca atttatgatg aaccgacgac gactactagc gtgcctttgg aaaaccttta
      841 ctttcagtcg caccatcacc atcaccatta aggatccgca ggcctctgct agcttgactg
      901 actgagatac agcgtacctt cagctcacag acatgataag atacattgat gagtttggac
      961 aaaccacaac tagaatgcag tgaaaaaaat gctttatttg tgaaatttgt gatgctattg
     1021 ctttatttgt aaccattata agctgcaata aacaagttaa caacaacaat tgcattcatt
     1081 ttatgtttca ggttcagggg gaggtgtggg aggtttttta aagcaagtaa aacctctaca
     1141 aatgtggtat tggcccatct ctatcggtat cgtagcataa ccccttgggg cctctaaacg
     1201 ggtcttgagg ggttttttgt gcccctcggg ccggattgct atctaccggc attggcgcag
     1261 aaaaaaatgc ctgatgcgac gctgcgcgtc ttatactccc acatatgcca gattcagcaa
     1321 cggatacggc ttccccaact tgcccacttc catacgtgtc ctccttacca gaaatttatc
     1381 cttaaggtcg tcagctatcc tgcaggcgat ctctcgattt cgatcaagac attcctttaa
     1441 tggtcttttc tggacaccac taggggtcag aagtagttca tcaaactttc ttccctccct
     1501 aatctcattg gttaccttgg gctatcgaaa cttaattaac cagtcaagtc agctacttgg
     1561 cgagatcgac ttgtctgggt ttcgactacg ctcagaattg cgtcagtcaa gttcgatctg
     1621 gtccttgcta ttgcacccgt tctccgatta cgagtttcat ttaaatcatg tgagcaaaag
     1681 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc
     1741 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
     1801 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
     1861 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
     1921 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
     1981 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
     2041 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
     2101 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
     2161 ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
     2221 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
     2281 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg
     2341 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa
     2401 aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta
     2461 tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag
     2521 cgatctgtct atttcgttca tccatagttg catttaaatt tccgaactct ccaaggccct
     2581 cgtcggaaaa tcttcaaacc tttcgtccga tccatcttgc aggctacctc tcgaacgaac
     2641 tatcgcaagt ctcttggccg gccttgcgcc ttggctattg cttggcagcg cctatcgcca
     2701 ggtattactc caatcccgaa tatccgagat cgggatcacc cgagagaagt tcaacctaca
     2761 tcctcaatcc cgatctatcc gagatccgag gaatatcgaa atcggggcgc gcctggcctc
     2821 cgcgccgggt tttggcgcct cccgcgggcg cccccctcgt cacggcgagc gctgccacgt
     2881 cagacgaagg gcgcaggagc gtcctgatcc ttccgcccgg acgctcagga cagcggcccg
     2941 ctgctcataa gactcggcct tagaacccca gtatcagcag aaggacattt taggacggga
     3001 cttgggtgac tctagggcac tggttttctt tccagagagc ggaacaggcg aggaaaagta
     3061 gtcccttctc ggcgattctg cggagggatc tccgtggggc ggtgaacgcc gatgattata
     3121 taaggacgcg ccgggtgtgg cacagctagt tccgtcgcag ccgggatttg ggtcgcggtt
     3181 cttgtttgtg gatcgctgtg atcgtcactt ggtgagtagc gggctgctgg gctggccggg
     3241 gctttcgtgg ccgccgggcc gctcggtggg acggaagcgt gtggagagac cgccaagggc
     3301 tgtagtctgg gtccgcgagc aaggttgccc tgaactgggg gttgggggga gcgcagcaaa
     3361 atggcggctg ttcccgagtc ttgaatggaa gacgcttgtg aggcgggctg tgaggtcgtt
     3421 gaaacaaggt ggggggcatg gtgggcggca agaacccaag gtcttgagcc cttcgctaat
     3481 gcgggaaagc tcttattcgg gtgagatggg ctgggcacca tctggggacc ctgacgtgaa
     3541 gtttgtcact gactggagaa ctcggtttgt cgtctgttgc gggggcggca gttatggcgg
     3601 tgccgttggg cagtgcaccc gtacctttgg gagcgcgcgc cctcgtcgtg tcgtgacgtc
     3661 acccgttctg ttggcttata atgcagggtg gggccacctg ccggtaggtg tgcggtaggc
     3721 ttttctccgt cgcaggacgc agggttcggg cctagggtag gctctcctga atcgacaggc
     3781 gccggacctc tggtgagggg agggataagt gaggcgtcag tttctttggt cggttttatg
     3841 tacctatctt cttaagtagc tgaagctccg gttttgaact atgcgctcgg ggttggcgag
     3901 tgtgttttgt gaagtttttt aggcaccttt tgaaatgtaa tcatttgggt caatatgtaa
     3961 ttttcagtgt tagacttgta aattgtccgc taaattctgg ccgtttttgg cttttttgtt
     4021 agacaacatg ggtaaaaagc ctgaactcac cgcgacgtct gtcgagaagt ttctgatcga
     4081 aaagttcgac agcgtctccg acctgatgca gctctcggag ggcgaagaat ctcgtgcttt
     4141 cagcttcgat gtaggagggc gtggatatgt cctgcgggta aatagctgcg ccgatggttt
     4201 ctacaaagat cgttatgttt atcggcactt tgcatccgcc gcgctcccga ttccggaagt
     4261 gcttgacatt ggggagttca gcgagagcct gacctattgc atctcccgcc gtgcacaggg
     4321 tgtcacgttg caagacctgc ctgaaaccga actgcccgct gttctgcagc cggtcgcgga
     4381 ggcaatggat gccatcgctg ccgccgatct tagccagacg agcgggttcg gcccattcgg
     4441 accgcaagga atcggtcaat acactacatg gcgtgatttc atatgcgcga ttgctgatcc
     4501 ccatgtgtat cactggcaaa ctgtgatgga cgacaccgtc agtgcgtccg tcgcgcaggc
     4561 tctcgatgag ctgatgcttt gggccgagga ctgccccgaa gtccggcacc tcgtgcacgc
     4621 ggatttcggc tccaacaatg tcctgacgga caatggccgc ataacagcgg tcattgactg
     4681 gagcgaggcg atgttcgggg attcccaata cgaggtcgcc aacatcttct tctggaggcc
     4741 gtggttggct tgtatggagc agcagacgcg ctacttcgag cggaggcatc cggagcttgc
     4801 aggatcgccc cggctccggg cgtatatgct ccgcattggt cttgaccaac tctatcagag
     4861 cttggttgac ggcaatttcg atgatgcagc ttgggcgcag ggtcgatgcg acgcaatcgt
     4921 ccgatccgga gccgggactg tcgggcgtac acaaatcgcc cgcagaagcg ccgccgtctg
     4981 gaccgatggc tgtgtagaag tactcgccga tagtggaaac cgacgcccca gcactcgtcc
     5041 gagggcaaag gaataagcta gtatgtaagc ctagtcttag ataataaaat cgctatccat
     5101 cgaagatgga tgtgtgttgg ttttttgtgt gtgtaacgct aggcgcgcct ggtgtaccga
     5161 gaacgatcct ctcagtgcga gtctcgacga tccatatcgt tgcttggcag tcagccagtc
     5221 ggaatccagc ttgggaccca ggaagtccaa tcgtcagata ttgtactcaa gcctggtcac
     5281 ggcagcgtac cgatctgttt aaacctagat attgatagtc tgatcggtca acgtataatc
     5341 gagtcctagc ttttgcaaac atctatcaag agacaggatc agcaggaggc tttcgcatga
     5401 gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt
     5461 ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcgcgag
     5521 tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag
     5581 aacgctttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta
     5641 ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg
     5701 agtattcacc agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca
     5761 gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca acgattggag
     5821 gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc
     5881 gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg
     5941 tagcaatggc aacaaccttg cgtaaactat taactggcga actacttact ctagcttccc
     6001 ggcaacagtt gatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg
     6061 cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg
     6121 gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga
     6181 cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac
     6241 tgattaagca ttggtaaccg attctaggtg cattggcgca gaaaaaaatg cctgatgcga
     6301 cgctgcgcgt cttatactcc cacatatgcc agattcagca acggatacgg cttccccaac
     6361 ttgcccactt ccatacgtgt cctccttacc agaaatttat ccttaagatc gtttaaactc
     6421 gactctggct ctatcgaatc tccgtcgttt cgagcttacg cgaacagccg tggcgctcat
     6481 ttgctcgtcg ggcatcgaat ctcgtcagct atcgtcagct tacctttttg gcagcgatcg
     6541 cggctcccga catcttggac cattagctcc acaggtatct tcttccctct agtggtcata
     6601 acagcagctt cagctacctc tcaattcaaa aaacccctca agacccgttt agaggcccca
     6661 aggggttatg ctatcaatcg ttgcgttaca cacacaaaaa accaacacac atccatcttc
     6721 gatggatagc gattttatta tctaactgct gatcgagtgt agccagatct agtaatcaat
     6781 tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa
     6841 tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt
     6901 tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta
     6961 aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt
     7021 caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc
     7081 tacttggcag tacatctacg tattagtcat cgctattacc atgctgatgc ggttttggca
     7141 gtacatcaat gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat
     7201 tgacgtcaat gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa
     7261 caactccgcc ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag
     7321 cagagctggt ttagtgaacc gtcagatcag atctttgtcg atcctaccat ccactcgaca
     7381 cacccgccag cggccgc
//
