LOCUS       sequence-307259-        7404 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 7404)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 7404)
  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-m4134277
FEATURES             Location/Qualifiers
     source          1..7404
                     /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    6558..6629
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(6634..6680)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    6698..6746
                     /note="synthetic polyadenylation signal"
     enhancer        6827..7130
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        7132..7334
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     7285..7305
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      join(7403..7404,1..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 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 ccgaatcgtt
     6421 taaactcgac tctggctcta tcgaatctcc gtcgtttcga gcttacgcga acagccgtgg
     6481 cgctcatttg ctcgtcgggc atcgaatctc gtcagctatc gtcagcttac ctttttggca
     6541 gcgatcgcgg ctcccgacat cttggaccat tagctccaca ggtatcttct tccctctagt
     6601 ggtcataaca gcagcttcag ctacctctca attcaaaaaa cccctcaaga cccgtttaga
     6661 ggccccaagg ggttatgcta tcaatcgttg cgttacacac acaaaaaacc aacacacatc
     6721 catcttcgat ggatagcgat tttattatct aactgctgat cgagtgtagc cagatctagt
     6781 aatcaattac ggggtcatta gttcatagcc catatatgga gttccgcgtt acataactta
     6841 cggtaaatgg cccgcctggc tgaccgccca acgacccccg cccattgacg tcaataatga
     6901 cgtatgttcc catagtaacg ccaataggga ctttccattg acgtcaatgg gtggagtatt
     6961 tacggtaaac tgcccacttg gcagtacatc aagtgtatca tatgccaagt acgcccccta
     7021 ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc ccagtacatg accttatggg
     7081 actttcctac ttggcagtac atctacgtat tagtcatcgc tattaccatg ctgatgcggt
     7141 tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt ccaagtctcc
     7201 accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat
     7261 gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct
     7321 atataagcag agctggttta gtgaaccgtc agatcagatc tttgtcgatc ctaccatcca
     7381 ctcgacacac ccgccagcgg ccgc
//
