LOCUS       sequence-295983-        6517 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Mammalian expression plasmid for SARS-CoV-2 ORF8 protein.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6517)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 6517)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: pGBW-m4133458
FEATURES             Location/Qualifiers
     source          1..6517
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             368..388
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             389..433
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     polyA_signal    486..607
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(497..516)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     551..570
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      722..768
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      798..934
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    979..1050
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(1276..1864)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1356..1375)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2364..3573
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     3410..3429
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             3577..4176
                     /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(3577..3596)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4073..4093
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4203..4251
                     /note="synthetic polyadenylation signal"
     CDS             join(4517..4585,4586..5377)
                     /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(4735..4754)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      5390..5526
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    5671..5742
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(5747..5793)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    5811..5859
                     /note="synthetic polyadenylation signal"
     enhancer        5940..6243
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6245..6447
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     6398..6418
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
ORIGIN
        1 caccatgaaa tttcttgttt tcttaggaat catcacaact gtagctgcat ttcaccaaga
       61 atgtagttta cagtcatgta ctcaacatca accatatgta gttgatgacc cgtgtcctat
      121 tcacttctat tctaaatggt atattagagt aggagctaga aaatcagcac ctttaattga
      181 attgtgcgtg gatgaggctg gttctaaatc acccattcag tacatcgata tcggtaatta
      241 tacagtttcc tgtttacctt ttacaattaa ttgccaggaa cctaaattgg gtagtcttgt
      301 agtgcgttgt tcgttctatg aagacttttt agagtatcat gacgttcgtg ttgttttaga
      361 tttcatcgaa aacctttact ttcagtcgaa agaaaccgcg gcagctaagt tcgagaggca
      421 gcacatggac agttaagcta gcttgactga ctgagataca gcgtaccttc agctcacaga
      481 catgataaga tacattgatg agtttggaca aaccacaact agaatgcagt gaaaaaaatg
      541 ctttatttgt gaaatttgtg atgctattgc tttatttgta accattataa gctgcaataa
      601 acaagttaac aacaacaatt gcattcattt tatgtttcag gttcaggggg aggtgtggga
      661 ggttttttaa agcaagtaaa acctctacaa atgtggtatt ggcccatctc tatcggtatc
      721 gtagcataac cccttggggc ctctaaacgg gtcttgaggg gttttttgtg cccctcgggc
      781 cggattgcta tctaccggca ttggcgcaga aaaaaatgcc tgatgcgacg ctgcgcgtct
      841 tatactccca catatgccag attcagcaac ggatacggct tccccaactt gcccacttcc
      901 atacgtgtcc tccttaccag aaatttatcc ttaaggtcgt cagctatcct gcaggcgatc
      961 tctcgatttc gatcaagaca ttcctttaat ggtcttttct ggacaccact aggggtcaga
     1021 agtagttcat caaactttct tccctcccta atctcattgg ttaccttggg ctatcgaaac
     1081 ttaattaacc agtcaagtca gctacttggc gagatcgact tgtctgggtt tcgactacgc
     1141 tcagaattgc gtcagtcaag ttcgatctgg tccttgctat tgcacccgtt ctccgattac
     1201 gagtttcatt taaatcatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc
     1261 cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg
     1321 ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg
     1381 aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt
     1441 tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt
     1501 gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg
     1561 cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact
     1621 ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt
     1681 cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct
     1741 gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac
     1801 cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc
     1861 tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg
     1921 ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta
     1981 aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca
     2041 atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc
     2101 atttaaattt ccgaactctc caaggccctc gtcggaaaat cttcaaacct ttcgtccgat
     2161 ccatcttgca ggctacctct cgaacgaact atcgcaagtc tcttggccgg ccttgcgcct
     2221 tggctattgc ttggcagcgc ctatcgccag gtattactcc aatcccgaat atccgagatc
     2281 gggatcaccc gagagaagtt caacctacat cctcaatccc gatctatccg agatccgagg
     2341 aatatcgaaa tcggggcgcg cctggcctcc gcgccgggtt ttggcgcctc ccgcgggcgc
     2401 ccccctcgtc acggcgagcg ctgccacgtc agacgaaggg cgcaggagcg tcctgatcct
     2461 tccgcccgga cgctcaggac agcggcccgc tgctcataag actcggcctt agaaccccag
     2521 tatcagcaga aggacatttt aggacgggac ttgggtgact ctagggcact ggttttcttt
     2581 ccagagagcg gaacaggcga ggaaaagtag tcccttctcg gcgattctgc ggagggatct
     2641 ccgtggggcg gtgaacgccg atgattatat aaggacgcgc cgggtgtggc acagctagtt
     2701 ccgtcgcagc cgggatttgg gtcgcggttc ttgtttgtgg atcgctgtga tcgtcacttg
     2761 gtgagtagcg ggctgctggg ctggccgggg ctttcgtggc cgccgggccg ctcggtggga
     2821 cggaagcgtg tggagagacc gccaagggct gtagtctggg tccgcgagca aggttgccct
     2881 gaactggggg ttggggggag cgcagcaaaa tggcggctgt tcccgagtct tgaatggaag
     2941 acgcttgtga ggcgggctgt gaggtcgttg aaacaaggtg gggggcatgg tgggcggcaa
     3001 gaacccaagg tcttgagccc ttcgctaatg cgggaaagct cttattcggg tgagatgggc
     3061 tgggcaccat ctggggaccc tgacgtgaag tttgtcactg actggagaac tcggtttgtc
     3121 gtctgttgcg ggggcggcag ttatggcggt gccgttgggc agtgcacccg tacctttggg
     3181 agcgcgcgcc ctcgtcgtgt cgtgacgtca cccgttctgt tggcttataa tgcagggtgg
     3241 ggccacctgc cggtaggtgt gcggtaggct tttctccgtc gcaggacgca gggttcgggc
     3301 ctagggtagg ctctcctgaa tcgacaggcg ccggacctct ggtgagggga gggataagtg
     3361 aggcgtcagt ttctttggtc ggttttatgt acctatcttc ttaagtagct gaagctccgg
     3421 ttttgaacta tgcgctcggg gttggcgagt gtgttttgtg aagtttttta ggcacctttt
     3481 gaaatgtaat catttgggtc aatatgtaat tttcagtgtt agacttgtaa attgtccgct
     3541 aaattctggc cgtttttggc ttttttgtta gacaacatga ccgagtacaa gcccacggtg
     3601 cgcctcgcca cccgcgacga cgtccccagg gccgtacgca ccctcgccgc cgcgttcgcc
     3661 gactaccccg ccacgcgcca caccgtcgat ccggaccgcc acatcgagcg ggtcaccgag
     3721 ctgcaagaac tcttcctcac gcgcgtcggg ctcgacatcg gcaaggtgtg ggtcgcggac
     3781 gacggcgccg cggtggcggt ctggaccacg ccggagagcg tcgaagcggg ggcggtgttc
     3841 gccgagatcg gcccgcgcat ggccgagttg agcggttccc ggctggccgc gcagcaacag
     3901 atggagggcc tcctggcgcc gcaccggccc aaggagcccg cgtggttcct ggccaccgtc
     3961 ggcgtctcgc ccgaccacca gggcaagggt ctgggcagcg ccgtcgtgct ccccggagtg
     4021 gaggccgccg agcgcgccgg ggtgcccgcc ttcctggaga cctccgcgcc ccgcaacctc
     4081 cccttctacg agcggctcgg cttcaccgtc accgccgacg tcgaggtgcc cgaaggaccg
     4141 cgcacctggt gcatgacccg caagcccggt gcctaggcta gtatgtaagc ctagtcttag
     4201 ataataaaat cgctatccat cgaagatgga tgtgtgttgg ttttttgtgt gtgtaacgct
     4261 aggcgcgcct ggtgtaccga gaacgatcct ctcagtgcga gtctcgacga tccatatcgt
     4321 tgcttggcag tcagccagtc ggaatccagc ttgggaccca ggaagtccaa tcgtcagata
     4381 ttgtactcaa gcctggtcac ggcagcgtac cgatctgttt aaacctagat attgatagtc
     4441 tgatcggtca acgtataatc gagtcctagc ttttgcaaac atctatcaag agacaggatc
     4501 agcaggaggc tttcgcatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc
     4561 ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga
     4621 agatcagttg ggtgcgcgag tgggttacat cgaactggat ctcaacagcg gtaagatcct
     4681 tgagagtttt cgccccgaag aacgctttcc aatgatgagc acttttaaag ttctgctatg
     4741 tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta
     4801 ttctcagaat gacttggttg agtattcacc agtcacagaa aagcatctta cggatggcat
     4861 gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt
     4921 acttctgaca acgattggag gaccgaagga gctaaccgct tttttgcaca acatggggga
     4981 tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga
     5041 gcgtgacacc acgatgcctg tagcaatggc aacaaccttg cgtaaactat taactggcga
     5101 actacttact ctagcttccc ggcaacagtt gatagactgg atggaggcgg ataaagttgc
     5161 aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc
     5221 cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg
     5281 tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat
     5341 cgctgagata ggtgcctcac tgattaagca ttggtaaccg attctaggtg cattggcgca
     5401 gaaaaaaatg cctgatgcga cgctgcgcgt cttatactcc cacatatgcc agattcagca
     5461 acggatacgg cttccccaac ttgcccactt ccatacgtgt cctccttacc agaaatttat
     5521 ccttaagatc gtttaaactc gactctggct ctatcgaatc tccgtcgttt cgagcttacg
     5581 cgaacagccg tggcgctcat ttgctcgtcg ggcatcgaat ctcgtcagct atcgtcagct
     5641 tacctttttg gcagcgatcg cggctcccga catcttggac cattagctcc acaggtatct
     5701 tcttccctct agtggtcata acagcagctt cagctacctc tcaattcaaa aaacccctca
     5761 agacccgttt agaggcccca aggggttatg ctatcaatcg ttgcgttaca cacacaaaaa
     5821 accaacacac atccatcttc gatggatagc gattttatta tctaactgct gatcgagtgt
     5881 agccagatct agtaatcaat tacggggtca ttagttcata gcccatatat ggagttccgc
     5941 gttacataac ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg
     6001 acgtcaataa tgacgtatgt tcccatagta acgccaatag ggactttcca ttgacgtcaa
     6061 tgggtggagt atttacggta aactgcccac ttggcagtac atcaagtgta tcatatgcca
     6121 agtacgcccc ctattgacgt caatgacggt aaatggcccg cctggcatta tgcccagtac
     6181 atgaccttat gggactttcc tacttggcag tacatctacg tattagtcat cgctattacc
     6241 atgctgatgc ggttttggca gtacatcaat gggcgtggat agcggtttga ctcacgggga
     6301 tttccaagtc tccaccccat tgacgtcaat gggagtttgt tttggcacca aaatcaacgg
     6361 gactttccaa aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg taggcgtgta
     6421 cggtgggagg tctatataag cagagctggt ttagtgaacc gtcagatcag atctttgtcg
     6481 atcctaccat ccactcgaca cacccgccag cggccgc
//
