LOCUS       sequence-295530-        7424 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 7424)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 7424)
  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-m4133905
FEATURES             Location/Qualifiers
     source          1..7424
                     /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..895
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     polyA_signal    964..1085
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(975..994)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1029..1048
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      1200..1246
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1276..1412
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1457..1528
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(1754..2342)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1834..1853)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2842..4051
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     3888..3907
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             4055..5083
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     polyA_signal    5110..5158
                     /note="synthetic polyadenylation signal"
     CDS             join(5424..5492,5493..6284)
                     /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(5642..5661)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      6297..6433
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    6578..6649
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(6654..6700)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    6718..6766
                     /note="synthetic polyadenylation signal"
     enhancer        6847..7150
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        7152..7354
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     7305..7325
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      join(7423..7424,1..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 aaagaaaccg cggcagctaa gttcgagagg cagcacatgg acagttaagg
      901 atccgcaggc ctctgctagc ttgactgact gagatacagc gtaccttcag ctcacagaca
      961 tgataagata cattgatgag tttggacaaa ccacaactag aatgcagtga aaaaaatgct
     1021 ttatttgtga aatttgtgat gctattgctt tatttgtaac cattataagc tgcaataaac
     1081 aagttaacaa caacaattgc attcatttta tgtttcaggt tcagggggag gtgtgggagg
     1141 ttttttaaag caagtaaaac ctctacaaat gtggtattgg cccatctcta tcggtatcgt
     1201 agcataaccc cttggggcct ctaaacgggt cttgaggggt tttttgtgcc cctcgggccg
     1261 gattgctatc taccggcatt ggcgcagaaa aaaatgcctg atgcgacgct gcgcgtctta
     1321 tactcccaca tatgccagat tcagcaacgg atacggcttc cccaacttgc ccacttccat
     1381 acgtgtcctc cttaccagaa atttatcctt aaggtcgtca gctatcctgc aggcgatctc
     1441 tcgatttcga tcaagacatt cctttaatgg tcttttctgg acaccactag gggtcagaag
     1501 tagttcatca aactttcttc cctccctaat ctcattggtt accttgggct atcgaaactt
     1561 aattaaccag tcaagtcagc tacttggcga gatcgacttg tctgggtttc gactacgctc
     1621 agaattgcgt cagtcaagtt cgatctggtc cttgctattg cacccgttct ccgattacga
     1681 gtttcattta aatcatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     1741 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     1801 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     1861 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     1921 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     1981 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     2041 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     2101 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     2161 tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc
     2221 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     2281 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     2341 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     2401 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     2461 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     2521 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcat
     2581 ttaaatttcc gaactctcca aggccctcgt cggaaaatct tcaaaccttt cgtccgatcc
     2641 atcttgcagg ctacctctcg aacgaactat cgcaagtctc ttggccggcc ttgcgccttg
     2701 gctattgctt ggcagcgcct atcgccaggt attactccaa tcccgaatat ccgagatcgg
     2761 gatcacccga gagaagttca acctacatcc tcaatcccga tctatccgag atccgaggaa
     2821 tatcgaaatc ggggcgcgcc tggcctccgc gccgggtttt ggcgcctccc gcgggcgccc
     2881 ccctcgtcac ggcgagcgct gccacgtcag acgaagggcg caggagcgtc ctgatccttc
     2941 cgcccggacg ctcaggacag cggcccgctg ctcataagac tcggccttag aaccccagta
     3001 tcagcagaag gacattttag gacgggactt gggtgactct agggcactgg ttttctttcc
     3061 agagagcgga acaggcgagg aaaagtagtc ccttctcggc gattctgcgg agggatctcc
     3121 gtggggcggt gaacgccgat gattatataa ggacgcgccg ggtgtggcac agctagttcc
     3181 gtcgcagccg ggatttgggt cgcggttctt gtttgtggat cgctgtgatc gtcacttggt
     3241 gagtagcggg ctgctgggct ggccggggct ttcgtggccg ccgggccgct cggtgggacg
     3301 gaagcgtgtg gagagaccgc caagggctgt agtctgggtc cgcgagcaag gttgccctga
     3361 actgggggtt ggggggagcg cagcaaaatg gcggctgttc ccgagtcttg aatggaagac
     3421 gcttgtgagg cgggctgtga ggtcgttgaa acaaggtggg gggcatggtg ggcggcaaga
     3481 acccaaggtc ttgagccctt cgctaatgcg ggaaagctct tattcgggtg agatgggctg
     3541 ggcaccatct ggggaccctg acgtgaagtt tgtcactgac tggagaactc ggtttgtcgt
     3601 ctgttgcggg ggcggcagtt atggcggtgc cgttgggcag tgcacccgta cctttgggag
     3661 cgcgcgccct cgtcgtgtcg tgacgtcacc cgttctgttg gcttataatg cagggtgggg
     3721 ccacctgccg gtaggtgtgc ggtaggcttt tctccgtcgc aggacgcagg gttcgggcct
     3781 agggtaggct ctcctgaatc gacaggcgcc ggacctctgg tgaggggagg gataagtgag
     3841 gcgtcagttt ctttggtcgg ttttatgtac ctatcttctt aagtagctga agctccggtt
     3901 ttgaactatg cgctcggggt tggcgagtgt gttttgtgaa gttttttagg caccttttga
     3961 aatgtaatca tttgggtcaa tatgtaattt tcagtgttag acttgtaaat tgtccgctaa
     4021 attctggccg tttttggctt ttttgttaga caacatgggt aaaaagcctg aactcaccgc
     4081 gacgtctgtc gagaagtttc tgatcgaaaa gttcgacagc gtctccgacc tgatgcagct
     4141 ctcggagggc gaagaatctc gtgctttcag cttcgatgta ggagggcgtg gatatgtcct
     4201 gcgggtaaat agctgcgccg atggtttcta caaagatcgt tatgtttatc ggcactttgc
     4261 atccgccgcg ctcccgattc cggaagtgct tgacattggg gagttcagcg agagcctgac
     4321 ctattgcatc tcccgccgtg cacagggtgt cacgttgcaa gacctgcctg aaaccgaact
     4381 gcccgctgtt ctgcagccgg tcgcggaggc aatggatgcc atcgctgccg ccgatcttag
     4441 ccagacgagc gggttcggcc cattcggacc gcaaggaatc ggtcaataca ctacatggcg
     4501 tgatttcata tgcgcgattg ctgatcccca tgtgtatcac tggcaaactg tgatggacga
     4561 caccgtcagt gcgtccgtcg cgcaggctct cgatgagctg atgctttggg ccgaggactg
     4621 ccccgaagtc cggcacctcg tgcacgcgga tttcggctcc aacaatgtcc tgacggacaa
     4681 tggccgcata acagcggtca ttgactggag cgaggcgatg ttcggggatt cccaatacga
     4741 ggtcgccaac atcttcttct ggaggccgtg gttggcttgt atggagcagc agacgcgcta
     4801 cttcgagcgg aggcatccgg agcttgcagg atcgccccgg ctccgggcgt atatgctccg
     4861 cattggtctt gaccaactct atcagagctt ggttgacggc aatttcgatg atgcagcttg
     4921 ggcgcagggt cgatgcgacg caatcgtccg atccggagcc gggactgtcg ggcgtacaca
     4981 aatcgcccgc agaagcgccg ccgtctggac cgatggctgt gtagaagtac tcgccgatag
     5041 tggaaaccga cgccccagca ctcgtccgag ggcaaaggaa taagctagta tgtaagccta
     5101 gtcttagata ataaaatcgc tatccatcga agatggatgt gtgttggttt tttgtgtgtg
     5161 taacgctagg cgcgcctggt gtaccgagaa cgatcctctc agtgcgagtc tcgacgatcc
     5221 atatcgttgc ttggcagtca gccagtcgga atccagcttg ggacccagga agtccaatcg
     5281 tcagatattg tactcaagcc tggtcacggc agcgtaccga tctgtttaaa cctagatatt
     5341 gatagtctga tcggtcaacg tataatcgag tcctagcttt tgcaaacatc tatcaagaga
     5401 caggatcagc aggaggcttt cgcatgagta ttcaacattt ccgtgtcgcc cttattccct
     5461 tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag
     5521 atgctgaaga tcagttgggt gcgcgagtgg gttacatcga actggatctc aacagcggta
     5581 agatccttga gagttttcgc cccgaagaac gctttccaat gatgagcact tttaaagttc
     5641 tgctatgtgg cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca
     5701 tacactattc tcagaatgac ttggttgagt attcaccagt cacagaaaag catcttacgg
     5761 atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg
     5821 ccaacttact tctgacaacg attggaggac cgaaggagct aaccgctttt ttgcacaaca
     5881 tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa
     5941 acgacgagcg tgacaccacg atgcctgtag caatggcaac aaccttgcgt aaactattaa
     6001 ctggcgaact acttactcta gcttcccggc aacagttgat agactggatg gaggcggata
     6061 aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat
     6121 ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc
     6181 cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata
     6241 gacagatcgc tgagataggt gcctcactga ttaagcattg gtaaccgatt ctaggtgcat
     6301 tggcgcagaa aaaaatgcct gatgcgacgc tgcgcgtctt atactcccac atatgccaga
     6361 ttcagcaacg gatacggctt ccccaacttg cccacttcca tacgtgtcct ccttaccaga
     6421 aatttatcct taagatcgtt taaactcgac tctggctcta tcgaatctcc gtcgtttcga
     6481 gcttacgcga acagccgtgg cgctcatttg ctcgtcgggc atcgaatctc gtcagctatc
     6541 gtcagcttac ctttttggca gcgatcgcgg ctcccgacat cttggaccat tagctccaca
     6601 ggtatcttct tccctctagt ggtcataaca gcagcttcag ctacctctca attcaaaaaa
     6661 cccctcaaga cccgtttaga ggccccaagg ggttatgcta tcaatcgttg cgttacacac
     6721 acaaaaaacc aacacacatc catcttcgat ggatagcgat tttattatct aactgctgat
     6781 cgagtgtagc cagatctagt aatcaattac ggggtcatta gttcatagcc catatatgga
     6841 gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca acgacccccg
     6901 cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga ctttccattg
     6961 acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc aagtgtatca
     7021 tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc
     7081 ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat tagtcatcgc
     7141 tattaccatg ctgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc
     7201 acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa
     7261 tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag
     7321 gcgtgtacgg tgggaggtct atataagcag agctggttta gtgaaccgtc agatcagatc
     7381 tttgtcgatc ctaccatcca ctcgacacac ccgccagcgg ccgc
//
