LOCUS       sequence-295186-        6613 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Mammalian expression plasmid for SARS-CoV-2 ORF10 protein.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6613)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 6613)
  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-m4133664
FEATURES             Location/Qualifiers
     source          1..6613
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             119..139
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             488..505
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     polyA_signal    582..703
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(593..612)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     647..666
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      818..864
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      894..1030
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1075..1146
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(1372..1960)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1452..1471)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2460..3669
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     3506..3525
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             3673..4272
                     /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(3673..3692)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4169..4189
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4299..4347
                     /note="synthetic polyadenylation signal"
     CDS             join(4613..4681,4682..5473)
                     /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(4831..4850)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      5486..5622
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    5767..5838
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(5843..5889)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    5907..5955
                     /note="synthetic polyadenylation signal"
     enhancer        6036..6339
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6341..6543
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     6494..6514
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      join(6612..6613,1..8)
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
ORIGIN
        1 caccatgggc tatataaacg ttttcgcttt tccgtttacg atatatagtc tactcttgtg
       61 cagaatgaat tctcgtaact acatagcaca agtagatgta gttaacttta atctcacaga
      121 aaacctttac tttcagtcga tgagtgacaa aatcattcat ctaaccgatg actcctttga
      181 tacagacgtg ttgaaggccg atggtgcgat actggtagat ttctgggctg aatggtgtgg
      241 cccgtgcaag atgatcgcac ccattctgga cgagatcgcc gacgagtatc aaggaaaatt
      301 aactgtcgct aagcttaata tagatcagaa cccagggacg gcacctaaat acggcatcag
      361 gggaatacct accctcctgc tctttaagaa tggggaagtt gccgcgacaa aagtgggagc
      421 cctgtctaag ggccagctaa aagagttctt ggacgctaac ctggcaggtt caggctcggg
      481 gcacatgcac catcaccatc accatagcag cgggctcgtg ccacgcggct aagctagctt
      541 gactgactga gatacagcgt accttcagct cacagacatg ataagataca ttgatgagtt
      601 tggacaaacc acaactagaa tgcagtgaaa aaaatgcttt atttgtgaaa tttgtgatgc
      661 tattgcttta tttgtaacca ttataagctg caataaacaa gttaacaaca acaattgcat
      721 tcattttatg tttcaggttc agggggaggt gtgggaggtt ttttaaagca agtaaaacct
      781 ctacaaatgt ggtattggcc catctctatc ggtatcgtag cataacccct tggggcctct
      841 aaacgggtct tgaggggttt tttgtgcccc tcgggccgga ttgctatcta ccggcattgg
      901 cgcagaaaaa aatgcctgat gcgacgctgc gcgtcttata ctcccacata tgccagattc
      961 agcaacggat acggcttccc caacttgccc acttccatac gtgtcctcct taccagaaat
     1021 ttatccttaa ggtcgtcagc tatcctgcag gcgatctctc gatttcgatc aagacattcc
     1081 tttaatggtc ttttctggac accactaggg gtcagaagta gttcatcaaa ctttcttccc
     1141 tccctaatct cattggttac cttgggctat cgaaacttaa ttaaccagtc aagtcagcta
     1201 cttggcgaga tcgacttgtc tgggtttcga ctacgctcag aattgcgtca gtcaagttcg
     1261 atctggtcct tgctattgca cccgttctcc gattacgagt ttcatttaaa tcatgtgagc
     1321 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag
     1381 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc
     1441 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt
     1501 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
     1561 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
     1621 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
     1681 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat
     1741 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg
     1801 ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa
     1861 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt
     1921 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc
     1981 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt
     2041 atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta
     2101 aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat
     2161 ctcagcgatc tgtctatttc gttcatccat agttgcattt aaatttccga actctccaag
     2221 gccctcgtcg gaaaatcttc aaacctttcg tccgatccat cttgcaggct acctctcgaa
     2281 cgaactatcg caagtctctt ggccggcctt gcgccttggc tattgcttgg cagcgcctat
     2341 cgccaggtat tactccaatc ccgaatatcc gagatcggga tcacccgaga gaagttcaac
     2401 ctacatcctc aatcccgatc tatccgagat ccgaggaata tcgaaatcgg ggcgcgcctg
     2461 gcctccgcgc cgggttttgg cgcctcccgc gggcgccccc ctcgtcacgg cgagcgctgc
     2521 cacgtcagac gaagggcgca ggagcgtcct gatccttccg cccggacgct caggacagcg
     2581 gcccgctgct cataagactc ggccttagaa ccccagtatc agcagaagga cattttagga
     2641 cgggacttgg gtgactctag ggcactggtt ttctttccag agagcggaac aggcgaggaa
     2701 aagtagtccc ttctcggcga ttctgcggag ggatctccgt ggggcggtga acgccgatga
     2761 ttatataagg acgcgccggg tgtggcacag ctagttccgt cgcagccggg atttgggtcg
     2821 cggttcttgt ttgtggatcg ctgtgatcgt cacttggtga gtagcgggct gctgggctgg
     2881 ccggggcttt cgtggccgcc gggccgctcg gtgggacgga agcgtgtgga gagaccgcca
     2941 agggctgtag tctgggtccg cgagcaaggt tgccctgaac tgggggttgg ggggagcgca
     3001 gcaaaatggc ggctgttccc gagtcttgaa tggaagacgc ttgtgaggcg ggctgtgagg
     3061 tcgttgaaac aaggtggggg gcatggtggg cggcaagaac ccaaggtctt gagcccttcg
     3121 ctaatgcggg aaagctctta ttcgggtgag atgggctggg caccatctgg ggaccctgac
     3181 gtgaagtttg tcactgactg gagaactcgg tttgtcgtct gttgcggggg cggcagttat
     3241 ggcggtgccg ttgggcagtg cacccgtacc tttgggagcg cgcgccctcg tcgtgtcgtg
     3301 acgtcacccg ttctgttggc ttataatgca gggtggggcc acctgccggt aggtgtgcgg
     3361 taggcttttc tccgtcgcag gacgcagggt tcgggcctag ggtaggctct cctgaatcga
     3421 caggcgccgg acctctggtg aggggaggga taagtgaggc gtcagtttct ttggtcggtt
     3481 ttatgtacct atcttcttaa gtagctgaag ctccggtttt gaactatgcg ctcggggttg
     3541 gcgagtgtgt tttgtgaagt tttttaggca ccttttgaaa tgtaatcatt tgggtcaata
     3601 tgtaattttc agtgttagac ttgtaaattg tccgctaaat tctggccgtt tttggctttt
     3661 ttgttagaca acatgaccga gtacaagccc acggtgcgcc tcgccacccg cgacgacgtc
     3721 cccagggccg tacgcaccct cgccgccgcg ttcgccgact accccgccac gcgccacacc
     3781 gtcgatccgg accgccacat cgagcgggtc accgagctgc aagaactctt cctcacgcgc
     3841 gtcgggctcg acatcggcaa ggtgtgggtc gcggacgacg gcgccgcggt ggcggtctgg
     3901 accacgccgg agagcgtcga agcgggggcg gtgttcgccg agatcggccc gcgcatggcc
     3961 gagttgagcg gttcccggct ggccgcgcag caacagatgg agggcctcct ggcgccgcac
     4021 cggcccaagg agcccgcgtg gttcctggcc accgtcggcg tctcgcccga ccaccagggc
     4081 aagggtctgg gcagcgccgt cgtgctcccc ggagtggagg ccgccgagcg cgccggggtg
     4141 cccgccttcc tggagacctc cgcgccccgc aacctcccct tctacgagcg gctcggcttc
     4201 accgtcaccg ccgacgtcga ggtgcccgaa ggaccgcgca cctggtgcat gacccgcaag
     4261 cccggtgcct aggctagtat gtaagcctag tcttagataa taaaatcgct atccatcgaa
     4321 gatggatgtg tgttggtttt ttgtgtgtgt aacgctaggc gcgcctggtg taccgagaac
     4381 gatcctctca gtgcgagtct cgacgatcca tatcgttgct tggcagtcag ccagtcggaa
     4441 tccagcttgg gacccaggaa gtccaatcgt cagatattgt actcaagcct ggtcacggca
     4501 gcgtaccgat ctgtttaaac ctagatattg atagtctgat cggtcaacgt ataatcgagt
     4561 cctagctttt gcaaacatct atcaagagac aggatcagca ggaggctttc gcatgagtat
     4621 tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc
     4681 tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg cgcgagtggg
     4741 ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg
     4801 ctttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga
     4861 cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta
     4921 ttcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc
     4981 tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga ttggaggacc
     5041 gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg
     5101 ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc
     5161 aatggcaaca accttgcgta aactattaac tggcgaacta cttactctag cttcccggca
     5221 acagttgata gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct
     5281 tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat
     5341 cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg
     5401 gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat
     5461 taagcattgg taaccgattc taggtgcatt ggcgcagaaa aaaatgcctg atgcgacgct
     5521 gcgcgtctta tactcccaca tatgccagat tcagcaacgg atacggcttc cccaacttgc
     5581 ccacttccat acgtgtcctc cttaccagaa atttatcctt aagatcgttt aaactcgact
     5641 ctggctctat cgaatctccg tcgtttcgag cttacgcgaa cagccgtggc gctcatttgc
     5701 tcgtcgggca tcgaatctcg tcagctatcg tcagcttacc tttttggcag cgatcgcggc
     5761 tcccgacatc ttggaccatt agctccacag gtatcttctt ccctctagtg gtcataacag
     5821 cagcttcagc tacctctcaa ttcaaaaaac ccctcaagac ccgtttagag gccccaaggg
     5881 gttatgctat caatcgttgc gttacacaca caaaaaacca acacacatcc atcttcgatg
     5941 gatagcgatt ttattatcta actgctgatc gagtgtagcc agatctagta atcaattacg
     6001 gggtcattag ttcatagccc atatatggag ttccgcgtta cataacttac ggtaaatggc
     6061 ccgcctggct gaccgcccaa cgacccccgc ccattgacgt caataatgac gtatgttccc
     6121 atagtaacgc caatagggac tttccattga cgtcaatggg tggagtattt acggtaaact
     6181 gcccacttgg cagtacatca agtgtatcat atgccaagta cgccccctat tgacgtcaat
     6241 gacggtaaat ggcccgcctg gcattatgcc cagtacatga ccttatggga ctttcctact
     6301 tggcagtaca tctacgtatt agtcatcgct attaccatgc tgatgcggtt ttggcagtac
     6361 atcaatgggc gtggatagcg gtttgactca cggggatttc caagtctcca ccccattgac
     6421 gtcaatggga gtttgttttg gcaccaaaat caacgggact ttccaaaatg tcgtaacaac
     6481 tccgccccat tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta tataagcaga
     6541 gctggtttag tgaaccgtca gatcagatct ttgtcgatcc taccatccac tcgacacacc
     6601 cgccagcggc cgc
//
