LOCUS       sequence-297306-        6248 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 6248)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 6248)
  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-m4252788
FEATURES             Location/Qualifiers
     source          1..6248
                     /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             140..157
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     polyA_signal    210..331
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(221..240)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     275..294
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      446..492
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      522..658
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    703..774
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(1000..1588)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1080..1099)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2088..3297
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     3134..3153
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             3301..3900
                     /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(3301..3320)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3797..3817
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    3927..3975
                     /note="synthetic polyadenylation signal"
     CDS             join(4241..4309,4310..5101)
                     /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(4459..4478)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      5114..5250
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    5402..5473
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(5478..5524)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    5542..5590
                     /note="synthetic polyadenylation signal"
     enhancer        5671..5974
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        5976..6178
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     6129..6149
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      join(6247..6248,1..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 tttcagtcgc accatcacca tcaccattaa gctagcttga ctgactgaga
      181 tacagcgtac cttcagctca cagacatgat aagatacatt gatgagtttg gacaaaccac
      241 aactagaatg cagtgaaaaa aatgctttat ttgtgaaatt tgtgatgcta ttgctttatt
      301 tgtaaccatt ataagctgca ataaacaagt taacaacaac aattgcattc attttatgtt
      361 tcaggttcag ggggaggtgt gggaggtttt ttaaagcaag taaaacctct acaaatgtgg
      421 tattggccca tctctatcgg tatcgtagca taaccccttg gggcctctaa acgggtcttg
      481 aggggttttt tgtgcccctc gggccggatt gctatctacc ggcattggcg cagaaaaaaa
      541 tgcctgatgc gacgctgcgc gtcttatact cccacatatg ccagattcag caacggatac
      601 ggcttcccca acttgcccac ttccatacgt gtcctcctta ccagaaattt atccttaagg
      661 tcgtcagcta tcctgcaggc gatctctcga tttcgatcaa gacattcctt taatggtctt
      721 ttctggacac cactaggggt cagaagtagt tcatcaaact ttcttccctc cctaatctca
      781 ttggttacct tgggctatcg aaacttaatt aaccagtcaa gtcagctact tggcgagatc
      841 gacttgtctg ggtttcgact acgctcagaa ttgcgtcagt caagttcgat ctggtccttg
      901 ctattgcacc cgttctccga ttacgagttt catttaaatc atgtgagcaa aaggccagca
      961 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc
     1021 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata
     1081 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc
     1141 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc
     1201 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga
     1261 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc
     1321 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag
     1381 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag
     1441 aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag
     1501 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca
     1561 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga
     1621 cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat
     1681 cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga
     1741 gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg
     1801 tctatttcgt tcatccatag ttgcatttaa atttccgaac tctccaaggc cctcgtcgga
     1861 aaatcttcaa acctttcgtc cgatccatct tgcaggctac ctctcgaacg aactatcgca
     1921 agtctcttgg ccggccttgc gccttggcta ttgcttggca gcgcctatcg ccaggtatta
     1981 ctccaatccc gaatatccga gatcgggatc acccgagaga agttcaacct acatcctcaa
     2041 tcccgatcta tccgagatcc gaggaatatc gaaatcgggg cgcgcctggc ctccgcgccg
     2101 ggttttggcg cctcccgcgg gcgcccccct cgtcacggcg agcgctgcca cgtcagacga
     2161 agggcgcagg agcgtcctga tccttccgcc cggacgctca ggacagcggc ccgctgctca
     2221 taagactcgg ccttagaacc ccagtatcag cagaaggaca ttttaggacg ggacttgggt
     2281 gactctaggg cactggtttt ctttccagag agcggaacag gcgaggaaaa gtagtccctt
     2341 ctcggcgatt ctgcggaggg atctccgtgg ggcggtgaac gccgatgatt atataaggac
     2401 gcgccgggtg tggcacagct agttccgtcg cagccgggat ttgggtcgcg gttcttgttt
     2461 gtggatcgct gtgatcgtca cttggtgagt agcgggctgc tgggctggcc ggggctttcg
     2521 tggccgccgg gccgctcggt gggacggaag cgtgtggaga gaccgccaag ggctgtagtc
     2581 tgggtccgcg agcaaggttg ccctgaactg ggggttgggg ggagcgcagc aaaatggcgg
     2641 ctgttcccga gtcttgaatg gaagacgctt gtgaggcggg ctgtgaggtc gttgaaacaa
     2701 ggtggggggc atggtgggcg gcaagaaccc aaggtcttga gcccttcgct aatgcgggaa
     2761 agctcttatt cgggtgagat gggctgggca ccatctgggg accctgacgt gaagtttgtc
     2821 actgactgga gaactcggtt tgtcgtctgt tgcgggggcg gcagttatgg cggtgccgtt
     2881 gggcagtgca cccgtacctt tgggagcgcg cgccctcgtc gtgtcgtgac gtcacccgtt
     2941 ctgttggctt ataatgcagg gtggggccac cagccggtag gtgtgcggta ggcttttctc
     3001 cgtcgcagga cgcagggttc gggcctaggg taggctctcc tgaatcgaca ggcgccggac
     3061 ctctggtgag gggagggata agtgaggcgt cagtttcttt ggtcggtttt atgtacctat
     3121 cttcttaagt agctgaagct ccggttttga actatgcgct cggggttggc gagtgtgttt
     3181 tgtgaagttt tttaggcacc ttttgaaatg taatcatttg ggtcaatatg taattttcag
     3241 tgttagactt gtaaattgtc cgctaaattc tggccgtttt tggctttttt gttagacaac
     3301 atgaccgagt acaagcccac ggtgcgcctc gccacccgcg acgacgtccc cagggccgta
     3361 cgcaccctcg ccgccgcgtt cgccgactac cccgccacgc gccacaccgt cgatccggac
     3421 cgccacatcg agcgggtcac cgagctgcaa gaactcttcc tcacgcgcgt cgggctcgac
     3481 atcggcaagg tgtgggtcgc ggacgacggc gccgcggtgg cggtctggac cacgccggag
     3541 agcgtcgaag cgggggcggt gttcgccgag atcggcccgc gcatggccga gttgagcggt
     3601 tcccggctgg ccgcgcagca acagatggag ggcctcctgg cgccgcaccg gcccaaggag
     3661 cccgcgtggt tcctggccac cgtcggcgtc tcgcccgacc accagggcaa gggtctgggc
     3721 agcgccgtcg tgctccccgg agtggaggcc gccgagcgcg ccggggtgcc cgccttcctg
     3781 gagacctccg cgccccgcaa cctccccttc tacgagcggc tcggcttcac cgtcaccgcc
     3841 gacgtcgagg tgcccgaagg accgcgcacc tggtgcatga cccgcaagcc cggtgcctag
     3901 gctagtatgt aagcctagtc ttagataata aaatcgctat ccatcgaaga tggatgtgtg
     3961 ttggtttttt gtgtgtgtaa cgctaggcgc gcctggtgta ccgagaacga tcctctcagt
     4021 gcgagtctcg acgatccata tcgttgcttg gcagtcagcc agtcggaatc cagcttggga
     4081 cccaggaagt ccaatcgtca gatattgtac tcaagcctgg tcacggcagc gtaccgatct
     4141 gtttaaacct agatattgat agtctgatcg gtcaacgtat aatcgagtcc tagcttttgc
     4201 aaacatctat caagagacag gatcagcagg aggctttcgc atgagtattc aacatttccg
     4261 tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc acccagaaac
     4321 gctggtgaaa gtaaaagatg ctgaagatca gttgggtgcg cgagtgggtt acatcgaact
     4381 ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgct ttccaatgat
     4441 gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga
     4501 gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtatt caccagtcac
     4561 agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg ccataaccat
     4621 gagtgataac actgcggcca acttacttct gacaacgatt ggaggaccga aggagctaac
     4681 cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct
     4741 gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgtagcaa tggcaacaac
     4801 cttgcgtaaa ctattaactg gcgaactact tactctagct tcccggcaac agttgataga
     4861 ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg
     4921 gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca ttgcagcact
     4981 ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac
     5041 tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta
     5101 accgattcta ggtgcattgg cgcagaaaaa aatgcctgat gcgacgctgc gcgtcttata
     5161 ctcccacata tgccagattc agcaacggat acggcttccc caacttgccc acttccatac
     5221 gtgtcctcct taccagaaat ttatccttaa gatcccgaat cgtttaaact cgactctggc
     5281 tctatcgaat ctccgtcgtt tcgagcttac gcgaacagcc gtggcgctca tttgctcgtc
     5341 gggcatcgaa tctcgtcagc tatcgtcagc ttaccttttt ggcagcgatc gcggctcccg
     5401 acatcttgga ccattagctc cacaggtatc ttcttccctc tagtggtcat aacagcagct
     5461 tcagctacct ctcaattcaa aaaacccctc aagacccgtt tagaggcccc aaggggttat
     5521 gctatcaatc gttgcgttac acacacaaaa aaccaacaca catccatctt cgatggatag
     5581 cgattttatt atctaactgc tgatcgagtg tagccagatc tagtaatcaa ttacggggtc
     5641 attagttcat agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc
     5701 tggctgaccg cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt
     5761 aacgccaata gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca
     5821 cttggcagta catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg
     5881 taaatggccc gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca
     5941 gtacatctac gtattagtca tcgctattac catgctgatg cggttttggc agtacatcaa
     6001 tgggcgtgga tagcggtttg actcacgggg atttccaagt ctccacccca ttgacgtcaa
     6061 tgggagtttg ttttggcacc aaaatcaacg ggactttcca aaatgtcgta acaactccgc
     6121 cccattgacg caaatgggcg gtaggcgtgt acggtgggag gtctatataa gcagagctgg
     6181 tttagtgaac cgtcagatca gatctttgtc gatcctacca tccactcgac acacccgcca
     6241 gcggccgc
//
