LOCUS       sequence-317730-        6959 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 6959)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 6959)
  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-m4133508
FEATURES             Location/Qualifiers
     source          1..6959
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..304
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        306..508
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     459..479
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      577..586
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1408..1428
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             1429..1446
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     polyA_signal    1499..1620
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1510..1529)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1564..1583
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      1735..1781
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1811..1947
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1992..2063
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(2289..2877)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2369..2388)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        3377..4586
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     4423..4442
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             4590..5189
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(4590..4609)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5086..5106
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    5216..5264
                     /note="synthetic polyadenylation signal"
     CDS             join(5530..5598,5599..6390)
                     /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(5748..5767)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      6403..6539
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    6691..6762
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(6767..6813)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    6831..6879
                     /note="synthetic polyadenylation signal"
ORIGIN
        1 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
       61 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      121 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      181 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      241 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      301 catgctgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      361 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      421 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      481 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatca gatctttgtc
      541 gatcctacca tccactcgac acacccgcca gcggccgcca ccatggatct gtttatgcgt
      601 atattcacca tcggcacggt tacattgaag caaggggaaa ttaaagacgc aactcccagt
      661 gacttcgtac gggcgaccgc tacaattccg atccaggcca gcctcccttt tggatggctt
      721 atcgtcggcg tggccctact ggctgtgttt cagtcggcat caaagataat cactctgaaa
      781 aagcgatggc agttagccct tagcaaaggt gtgcacttcg tatgcaattt gctgctcctg
      841 ttcgtcaccg tttattccca tctgctcctg gtggctgccg gattggaggc accattttta
      901 tacctatacg ccctcgtgta ttttctgcaa tctattaact tcgtccgcat tatcatgaga
      961 ctctggctct gttggaagtg caggtctaaa aatcccttgt tatatgatgc caactacttc
     1021 ctgtgttggc acactaactg ctatgactac tgtattccat acaattcagt tacctccagt
     1081 atagtgatca catctgggga tggtacgact tcgcctattt ccgagcatga ctatcagatc
     1141 ggcgggtaca cagaaaagtg ggagagcgga gtcaaggatt gcgtggtgct tcactcctat
     1201 ttcaccagtg actactacca gctttatagc acacaacttt caacagacac cggcgtggag
     1261 catgtgactt ttttcattta taataaaatc gttgatgaac cggaggaaca cgttcagata
     1321 cacactatcg acggcagctc tggtgtcgtc aaccctgtta tggagcccat ttacgatgaa
     1381 ccaacgacca caacatctgt acctctggaa aacctttact ttcagtcgca ccatcaccat
     1441 caccattaag ctagcttgac tgactgagat acagcgtacc ttcagctcac agacatgata
     1501 agatacattg atgagtttgg acaaaccaca actagaatgc agtgaaaaaa atgctttatt
     1561 tgtgaaattt gtgatgctat tgctttattt gtaaccatta taagctgcaa taaacaagtt
     1621 aacaacaaca attgcattca ttttatgttt caggttcagg gggaggtgtg ggaggttttt
     1681 taaagcaagt aaaacctcta caaatgtggt attggcccat ctctatcggt atcgtagcat
     1741 aaccccttgg ggcctctaaa cgggtcttga ggggtttttt gtgcccctcg ggccggattg
     1801 ctatctaccg gcattggcgc agaaaaaaat gcctgatgcg acgctgcgcg tcttatactc
     1861 ccacatatgc cagattcagc aacggatacg gcttccccaa cttgcccact tccatacgtg
     1921 tcctccttac cagaaattta tccttaaggt cgtcagctat cctgcaggcg atctctcgat
     1981 ttcgatcaag acattccttt aatggtcttt tctggacacc actaggggtc agaagtagtt
     2041 catcaaactt tcttccctcc ctaatctcat tggttacctt gggctatcga aacttaatta
     2101 accagtcaag tcagctactt ggcgagatcg acttgtctgg gtttcgacta cgctcagaat
     2161 tgcgtcagtc aagttcgatc tggtccttgc tattgcaccc gttctccgat tacgagtttc
     2221 atttaaatca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg
     2281 ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt
     2341 cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc
     2401 ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct
     2461 tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc
     2521 gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta
     2581 tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca
     2641 gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag
     2701 tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag
     2761 ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt
     2821 agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa
     2881 gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg
     2941 attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga
     3001 agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta
     3061 atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt tgcatttaaa
     3121 tttccgaact ctccaaggcc ctcgtcggaa aatcttcaaa cctttcgtcc gatccatctt
     3181 gcaggctacc tctcgaacga actatcgcaa gtctcttggc cggccttgcg ccttggctat
     3241 tgcttggcag cgcctatcgc caggtattac tccaatcccg aatatccgag atcgggatca
     3301 cccgagagaa gttcaaccta catcctcaat cccgatctat ccgagatccg aggaatatcg
     3361 aaatcggggc gcgcctggcc tccgcgccgg gttttggcgc ctcccgcggg cgcccccctc
     3421 gtcacggcga gcgctgccac gtcagacgaa gggcgcagga gcgtcctgat ccttccgccc
     3481 ggacgctcag gacagcggcc cgctgctcat aagactcggc cttagaaccc cagtatcagc
     3541 agaaggacat tttaggacgg gacttgggtg actctagggc actggttttc tttccagaga
     3601 gcggaacagg cgaggaaaag tagtcccttc tcggcgattc tgcggaggga tctccgtggg
     3661 gcggtgaacg ccgatgatta tataaggacg cgccgggtgt ggcacagcta gttccgtcgc
     3721 agccgggatt tgggtcgcgg ttcttgtttg tggatcgctg tgatcgtcac ttggtgagta
     3781 gcgggctgct gggctggccg gggctttcgt ggccgccggg ccgctcggtg ggacggaagc
     3841 gtgtggagag accgccaagg gctgtagtct gggtccgcga gcaaggttgc cctgaactgg
     3901 gggttggggg gagcgcagca aaatggcggc tgttcccgag tcttgaatgg aagacgcttg
     3961 tgaggcgggc tgtgaggtcg ttgaaacaag gtggggggca tggtgggcgg caagaaccca
     4021 aggtcttgag cccttcgcta atgcgggaaa gctcttattc gggtgagatg ggctgggcac
     4081 catctgggga ccctgacgtg aagtttgtca ctgactggag aactcggttt gtcgtctgtt
     4141 gcgggggcgg cagttatggc ggtgccgttg ggcagtgcac ccgtaccttt gggagcgcgc
     4201 gccctcgtcg tgtcgtgacg tcacccgttc tgttggctta taatgcaggg tggggccacc
     4261 tgccggtagg tgtgcggtag gcttttctcc gtcgcaggac gcagggttcg ggcctagggt
     4321 aggctctcct gaatcgacag gcgccggacc tctggtgagg ggagggataa gtgaggcgtc
     4381 agtttctttg gtcggtttta tgtacctatc ttcttaagta gctgaagctc cggttttgaa
     4441 ctatgcgctc ggggttggcg agtgtgtttt gtgaagtttt ttaggcacct tttgaaatgt
     4501 aatcatttgg gtcaatatgt aattttcagt gttagacttg taaattgtcc gctaaattct
     4561 ggccgttttt ggcttttttg ttagacaaca tgaccgagta caagcccacg gtgcgcctcg
     4621 ccacccgcga cgacgtcccc agggccgtac gcaccctcgc cgccgcgttc gccgactacc
     4681 ccgccacgcg ccacaccgtc gatccggacc gccacatcga gcgggtcacc gagctgcaag
     4741 aactcttcct cacgcgcgtc gggctcgaca tcggcaaggt gtgggtcgcg gacgacggcg
     4801 ccgcggtggc ggtctggacc acgccggaga gcgtcgaagc gggggcggtg ttcgccgaga
     4861 tcggcccgcg catggccgag ttgagcggtt cccggctggc cgcgcagcaa cagatggagg
     4921 gcctcctggc gccgcaccgg cccaaggagc ccgcgtggtt cctggccacc gtcggcgtct
     4981 cgcccgacca ccagggcaag ggtctgggca gcgccgtcgt gctccccgga gtggaggccg
     5041 ccgagcgcgc cggggtgccc gccttcctgg agacctccgc gccccgcaac ctccccttct
     5101 acgagcggct cggcttcacc gtcaccgccg acgtcgaggt gcccgaagga ccgcgcacct
     5161 ggtgcatgac ccgcaagccc ggtgcctagg ctagtatgta agcctagtct tagataataa
     5221 aatcgctatc catcgaagat ggatgtgtgt tggttttttg tgtgtgtaac gctaggcgcg
     5281 cctggtgtac cgagaacgat cctctcagtg cgagtctcga cgatccatat cgttgcttgg
     5341 cagtcagcca gtcggaatcc agcttgggac ccaggaagtc caatcgtcag atattgtact
     5401 caagcctggt cacggcagcg taccgatctg tttaaaccta gatattgata gtctgatcgg
     5461 tcaacgtata atcgagtcct agcttttgca aacatctatc aagagacagg atcagcagga
     5521 ggctttcgca tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt
     5581 tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag
     5641 ttgggtgcgc gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt
     5701 tttcgccccg aagaacgctt tccaatgatg agcactttta aagttctgct atgtggcgcg
     5761 gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag
     5821 aatgacttgg ttgagtattc accagtcaca gaaaagcatc ttacggatgg catgacagta
     5881 agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg
     5941 acaacgattg gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta
     6001 actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac
     6061 accacgatgc ctgtagcaat ggcaacaacc ttgcgtaaac tattaactgg cgaactactt
     6121 actctagctt cccggcaaca gttgatagac tggatggagg cggataaagt tgcaggacca
     6181 cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag
     6241 cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta
     6301 gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag
     6361 ataggtgcct cactgattaa gcattggtaa ccgattctag gtgcattggc gcagaaaaaa
     6421 atgcctgatg cgacgctgcg cgtcttatac tcccacatat gccagattca gcaacggata
     6481 cggcttcccc aacttgccca cttccatacg tgtcctcctt accagaaatt tatccttaag
     6541 atcccgaatc gtttaaactc gactctggct ctatcgaatc tccgtcgttt cgagcttacg
     6601 cgaacagccg tggcgctcat ttgctcgtcg ggcatcgaat ctcgtcagct atcgtcagct
     6661 tacctttttg gcagcgatcg cggctcccga catcttggac cattagctcc acaggtatct
     6721 tcttccctct agtggtcata acagcagctt cagctacctc tcaattcaaa aaacccctca
     6781 agacccgttt agaggcccca aggggttatg ctatcaatcg ttgcgttaca cacacaaaaa
     6841 accaacacac atccatcttc gatggatagc gattttatta tctaactgct gatcgagtgt
     6901 agccagatct agtaatcaat tacggggtca ttagttcata gcccatatat ggagttccg
//
