LOCUS       sequence-295960-        7769 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 7769)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 7769)
  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-m4133785
FEATURES             Location/Qualifiers
     source          1..7769
                     /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             1199..1216
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     polyA_signal    1309..1430
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1320..1339)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1374..1393
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      1545..1591
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1621..1757
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1802..1873
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(2099..2687)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2179..2198)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        3187..4396
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     4233..4252
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             4400..5428
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     polyA_signal    5455..5503
                     /note="synthetic polyadenylation signal"
     CDS             join(5769..5837,5838..6629)
                     /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(5987..6006)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      6642..6778
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    6923..6994
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(6999..7045)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    7063..7111
                     /note="synthetic polyadenylation signal"
     enhancer        7192..7495
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        7497..7699
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     7650..7670
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      join(7768..7769,1..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 atgagtgaca aaatcattca tctaaccgat gactcctttg atacagacgt
      901 gttgaaggcc gatggtgcga tactggtaga tttctgggct gaatggtgtg gcccgtgcaa
      961 gatgatcgca cccattctgg acgagatcgc cgacgagtat caaggaaaat taactgtcgc
     1021 taagcttaat atagatcaga acccagggac ggcacctaaa tacggcatca ggggaatacc
     1081 taccctcctg ctctttaaga atggggaagt tgccgcgaca aaagtgggag ccctgtctaa
     1141 gggccagcta aaagagttct tggacgctaa cctggcaggt tcaggctcgg ggcacatgca
     1201 ccatcaccat caccatagca gcgggctcgt gccacgcggc taaggatccg caggcctctg
     1261 ctagcttgac tgactgagat acagcgtacc ttcagctcac agacatgata agatacattg
     1321 atgagtttgg acaaaccaca actagaatgc agtgaaaaaa atgctttatt tgtgaaattt
     1381 gtgatgctat tgctttattt gtaaccatta taagctgcaa taaacaagtt aacaacaaca
     1441 attgcattca ttttatgttt caggttcagg gggaggtgtg ggaggttttt taaagcaagt
     1501 aaaacctcta caaatgtggt attggcccat ctctatcggt atcgtagcat aaccccttgg
     1561 ggcctctaaa cgggtcttga ggggtttttt gtgcccctcg ggccggattg ctatctaccg
     1621 gcattggcgc agaaaaaaat gcctgatgcg acgctgcgcg tcttatactc ccacatatgc
     1681 cagattcagc aacggatacg gcttccccaa cttgcccact tccatacgtg tcctccttac
     1741 cagaaattta tccttaaggt cgtcagctat cctgcaggcg atctctcgat ttcgatcaag
     1801 acattccttt aatggtcttt tctggacacc actaggggtc agaagtagtt catcaaactt
     1861 tcttccctcc ctaatctcat tggttacctt gggctatcga aacttaatta accagtcaag
     1921 tcagctactt ggcgagatcg acttgtctgg gtttcgacta cgctcagaat tgcgtcagtc
     1981 aagttcgatc tggtccttgc tattgcaccc gttctccgat tacgagtttc atttaaatca
     2041 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
     2101 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
     2161 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
     2221 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
     2281 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
     2341 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
     2401 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
     2461 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta
     2521 actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct
     2581 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt
     2641 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga
     2701 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca
     2761 tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat
     2821 caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg
     2881 cacctatctc agcgatctgt ctatttcgtt catccatagt tgcatttaaa tttccgaact
     2941 ctccaaggcc ctcgtcggaa aatcttcaaa cctttcgtcc gatccatctt gcaggctacc
     3001 tctcgaacga actatcgcaa gtctcttggc cggccttgcg ccttggctat tgcttggcag
     3061 cgcctatcgc caggtattac tccaatcccg aatatccgag atcgggatca cccgagagaa
     3121 gttcaaccta catcctcaat cccgatctat ccgagatccg aggaatatcg aaatcggggc
     3181 gcgcctggcc tccgcgccgg gttttggcgc ctcccgcggg cgcccccctc gtcacggcga
     3241 gcgctgccac gtcagacgaa gggcgcagga gcgtcctgat ccttccgccc ggacgctcag
     3301 gacagcggcc cgctgctcat aagactcggc cttagaaccc cagtatcagc agaaggacat
     3361 tttaggacgg gacttgggtg actctagggc actggttttc tttccagaga gcggaacagg
     3421 cgaggaaaag tagtcccttc tcggcgattc tgcggaggga tctccgtggg gcggtgaacg
     3481 ccgatgatta tataaggacg cgccgggtgt ggcacagcta gttccgtcgc agccgggatt
     3541 tgggtcgcgg ttcttgtttg tggatcgctg tgatcgtcac ttggtgagta gcgggctgct
     3601 gggctggccg gggctttcgt ggccgccggg ccgctcggtg ggacggaagc gtgtggagag
     3661 accgccaagg gctgtagtct gggtccgcga gcaaggttgc cctgaactgg gggttggggg
     3721 gagcgcagca aaatggcggc tgttcccgag tcttgaatgg aagacgcttg tgaggcgggc
     3781 tgtgaggtcg ttgaaacaag gtggggggca tggtgggcgg caagaaccca aggtcttgag
     3841 cccttcgcta atgcgggaaa gctcttattc gggtgagatg ggctgggcac catctgggga
     3901 ccctgacgtg aagtttgtca ctgactggag aactcggttt gtcgtctgtt gcgggggcgg
     3961 cagttatggc ggtgccgttg ggcagtgcac ccgtaccttt gggagcgcgc gccctcgtcg
     4021 tgtcgtgacg tcacccgttc tgttggctta taatgcaggg tggggccacc tgccggtagg
     4081 tgtgcggtag gcttttctcc gtcgcaggac gcagggttcg ggcctagggt aggctctcct
     4141 gaatcgacag gcgccggacc tctggtgagg ggagggataa gtgaggcgtc agtttctttg
     4201 gtcggtttta tgtacctatc ttcttaagta gctgaagctc cggttttgaa ctatgcgctc
     4261 ggggttggcg agtgtgtttt gtgaagtttt ttaggcacct tttgaaatgt aatcatttgg
     4321 gtcaatatgt aattttcagt gttagacttg taaattgtcc gctaaattct ggccgttttt
     4381 ggcttttttg ttagacaaca tgggtaaaaa gcctgaactc accgcgacgt ctgtcgagaa
     4441 gtttctgatc gaaaagttcg acagcgtctc cgacctgatg cagctctcgg agggcgaaga
     4501 atctcgtgct ttcagcttcg atgtaggagg gcgtggatat gtcctgcggg taaatagctg
     4561 cgccgatggt ttctacaaag atcgttatgt ttatcggcac tttgcatccg ccgcgctccc
     4621 gattccggaa gtgcttgaca ttggggagtt cagcgagagc ctgacctatt gcatctcccg
     4681 ccgtgcacag ggtgtcacgt tgcaagacct gcctgaaacc gaactgcccg ctgttctgca
     4741 gccggtcgcg gaggcaatgg atgccatcgc tgccgccgat cttagccaga cgagcgggtt
     4801 cggcccattc ggaccgcaag gaatcggtca atacactaca tggcgtgatt tcatatgcgc
     4861 gattgctgat ccccatgtgt atcactggca aactgtgatg gacgacaccg tcagtgcgtc
     4921 cgtcgcgcag gctctcgatg agctgatgct ttgggccgag gactgccccg aagtccggca
     4981 cctcgtgcac gcggatttcg gctccaacaa tgtcctgacg gacaatggcc gcataacagc
     5041 ggtcattgac tggagcgagg cgatgttcgg ggattcccaa tacgaggtcg ccaacatctt
     5101 cttctggagg ccgtggttgg cttgtatgga gcagcagacg cgctacttcg agcggaggca
     5161 tccggagctt gcaggatcgc cccggctccg ggcgtatatg ctccgcattg gtcttgacca
     5221 actctatcag agcttggttg acggcaattt cgatgatgca gcttgggcgc agggtcgatg
     5281 cgacgcaatc gtccgatccg gagccgggac tgtcgggcgt acacaaatcg cccgcagaag
     5341 cgccgccgtc tggaccgatg gctgtgtaga agtactcgcc gatagtggaa accgacgccc
     5401 cagcactcgt ccgagggcaa aggaataagc tagtatgtaa gcctagtctt agataataaa
     5461 atcgctatcc atcgaagatg gatgtgtgtt ggttttttgt gtgtgtaacg ctaggcgcgc
     5521 ctggtgtacc gagaacgatc ctctcagtgc gagtctcgac gatccatatc gttgcttggc
     5581 agtcagccag tcggaatcca gcttgggacc caggaagtcc aatcgtcaga tattgtactc
     5641 aagcctggtc acggcagcgt accgatctgt ttaaacctag atattgatag tctgatcggt
     5701 caacgtataa tcgagtccta gcttttgcaa acatctatca agagacagga tcagcaggag
     5761 gctttcgcat gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt
     5821 gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt
     5881 tgggtgcgcg agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt
     5941 ttcgccccga agaacgcttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg
     6001 tattatcccg tattgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga
     6061 atgacttggt tgagtattca ccagtcacag aaaagcatct tacggatggc atgacagtaa
     6121 gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga
     6181 caacgattgg aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa
     6241 ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca
     6301 ccacgatgcc tgtagcaatg gcaacaacct tgcgtaaact attaactggc gaactactta
     6361 ctctagcttc ccggcaacag ttgatagact ggatggaggc ggataaagtt gcaggaccac
     6421 ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc
     6481 gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag
     6541 ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga
     6601 taggtgcctc actgattaag cattggtaac cgattctagg tgcattggcg cagaaaaaaa
     6661 tgcctgatgc gacgctgcgc gtcttatact cccacatatg ccagattcag caacggatac
     6721 ggcttcccca acttgcccac ttccatacgt gtcctcctta ccagaaattt atccttaaga
     6781 tcgtttaaac tcgactctgg ctctatcgaa tctccgtcgt ttcgagctta cgcgaacagc
     6841 cgtggcgctc atttgctcgt cgggcatcga atctcgtcag ctatcgtcag cttacctttt
     6901 tggcagcgat cgcggctccc gacatcttgg accattagct ccacaggtat cttcttccct
     6961 ctagtggtca taacagcagc ttcagctacc tctcaattca aaaaacccct caagacccgt
     7021 ttagaggccc caaggggtta tgctatcaat cgttgcgtta cacacacaaa aaaccaacac
     7081 acatccatct tcgatggata gcgattttat tatctaactg ctgatcgagt gtagccagat
     7141 ctagtaatca attacggggt cattagttca tagcccatat atggagttcc gcgttacata
     7201 acttacggta aatggcccgc ctggctgacc gcccaacgac ccccgcccat tgacgtcaat
     7261 aatgacgtat gttcccatag taacgccaat agggactttc cattgacgtc aatgggtgga
     7321 gtatttacgg taaactgccc acttggcagt acatcaagtg tatcatatgc caagtacgcc
     7381 ccctattgac gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt acatgacctt
     7441 atgggacttt cctacttggc agtacatcta cgtattagtc atcgctatta ccatgctgat
     7501 gcggttttgg cagtacatca atgggcgtgg atagcggttt gactcacggg gatttccaag
     7561 tctccacccc attgacgtca atgggagttt gttttggcac caaaatcaac gggactttcc
     7621 aaaatgtcgt aacaactccg ccccattgac gcaaatgggc ggtaggcgtg tacggtggga
     7681 ggtctatata agcagagctg gtttagtgaa ccgtcagatc agatctttgt cgatcctacc
     7741 atccactcga cacacccgcc agcggccgc
//
