LOCUS       sequence-297452-        6308 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 6308)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 6308)
  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-m4252843
FEATURES             Location/Qualifiers
     source          1..6308
                     /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..217
                     /codon_start=1
                     /product="calmodulin-binding peptide"
                     /label=CBP
                     /note="derived from skeletal muscle myosin light chain 
                     kinase; binds calmodulin with nanomolar affinity in the 
                     presence of calcium"
                     /translation="KRRWKKNFIAVSAANRFKKISSSGAL"
     polyA_signal    270..391
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(281..300)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     335..354
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      506..552
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      582..718
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    763..834
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(1060..1648)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1140..1159)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2148..3357
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     3194..3213
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             3361..3960
                     /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(3361..3380)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3857..3877
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    3987..4035
                     /note="synthetic polyadenylation signal"
     CDS             join(4301..4369,4370..5161)
                     /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(4519..4538)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      5174..5310
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    5462..5533
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(5538..5584)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    5602..5650
                     /note="synthetic polyadenylation signal"
     enhancer        5731..6034
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6036..6238
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     6189..6209
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      join(6307..6308,1..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 agaggcgatg gaaaaagaat tttatcgcgg tgagtgcagc
      181 caacagattc aaaaagatat cctcgtctgg tgccctctaa gctagcttga ctgactgaga
      241 tacagcgtac cttcagctca cagacatgat aagatacatt gatgagtttg gacaaaccac
      301 aactagaatg cagtgaaaaa aatgctttat ttgtgaaatt tgtgatgcta ttgctttatt
      361 tgtaaccatt ataagctgca ataaacaagt taacaacaac aattgcattc attttatgtt
      421 tcaggttcag ggggaggtgt gggaggtttt ttaaagcaag taaaacctct acaaatgtgg
      481 tattggccca tctctatcgg tatcgtagca taaccccttg gggcctctaa acgggtcttg
      541 aggggttttt tgtgcccctc gggccggatt gctatctacc ggcattggcg cagaaaaaaa
      601 tgcctgatgc gacgctgcgc gtcttatact cccacatatg ccagattcag caacggatac
      661 ggcttcccca acttgcccac ttccatacgt gtcctcctta ccagaaattt atccttaagg
      721 tcgtcagcta tcctgcaggc gatctctcga tttcgatcaa gacattcctt taatggtctt
      781 ttctggacac cactaggggt cagaagtagt tcatcaaact ttcttccctc cctaatctca
      841 ttggttacct tgggctatcg aaacttaatt aaccagtcaa gtcagctact tggcgagatc
      901 gacttgtctg ggtttcgact acgctcagaa ttgcgtcagt caagttcgat ctggtccttg
      961 ctattgcacc cgttctccga ttacgagttt catttaaatc atgtgagcaa aaggccagca
     1021 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc
     1081 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata
     1141 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc
     1201 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc
     1261 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga
     1321 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc
     1381 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag
     1441 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag
     1501 aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag
     1561 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca
     1621 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga
     1681 cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat
     1741 cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga
     1801 gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg
     1861 tctatttcgt tcatccatag ttgcatttaa atttccgaac tctccaaggc cctcgtcgga
     1921 aaatcttcaa acctttcgtc cgatccatct tgcaggctac ctctcgaacg aactatcgca
     1981 agtctcttgg ccggccttgc gccttggcta ttgcttggca gcgcctatcg ccaggtatta
     2041 ctccaatccc gaatatccga gatcgggatc acccgagaga agttcaacct acatcctcaa
     2101 tcccgatcta tccgagatcc gaggaatatc gaaatcgggg cgcgcctggc ctccgcgccg
     2161 ggttttggcg cctcccgcgg gcgcccccct cgtcacggcg agcgctgcca cgtcagacga
     2221 agggcgcagg agcgtcctga tccttccgcc cggacgctca ggacagcggc ccgctgctca
     2281 taagactcgg ccttagaacc ccagtatcag cagaaggaca ttttaggacg ggacttgggt
     2341 gactctaggg cactggtttt ctttccagag agcggaacag gcgaggaaaa gtagtccctt
     2401 ctcggcgatt ctgcggaggg atctccgtgg ggcggtgaac gccgatgatt atataaggac
     2461 gcgccgggtg tggcacagct agttccgtcg cagccgggat ttgggtcgcg gttcttgttt
     2521 gtggatcgct gtgatcgtca cttggtgagt agcgggctgc tgggctggcc ggggctttcg
     2581 tggccgccgg gccgctcggt gggacggaag cgtgtggaga gaccgccaag ggctgtagtc
     2641 tgggtccgcg agcaaggttg ccctgaactg ggggttgggg ggagcgcagc aaaatggcgg
     2701 ctgttcccga gtcttgaatg gaagacgctt gtgaggcggg ctgtgaggtc gttgaaacaa
     2761 ggtggggggc atggtgggcg gcaagaaccc aaggtcttga gcccttcgct aatgcgggaa
     2821 agctcttatt cgggtgagat gggctgggca ccatctgggg accctgacgt gaagtttgtc
     2881 actgactgga gaactcggtt tgtcgtctgt tgcgggggcg gcagttatgg cggtgccgtt
     2941 gggcagtgca cccgtacctt tgggagcgcg cgccctcgtc gtgtcgtgac gtcacccgtt
     3001 ctgttggctt ataatgcagg gtggggccac cagccggtag gtgtgcggta ggcttttctc
     3061 cgtcgcagga cgcagggttc gggcctaggg taggctctcc tgaatcgaca ggcgccggac
     3121 ctctggtgag gggagggata agtgaggcgt cagtttcttt ggtcggtttt atgtacctat
     3181 cttcttaagt agctgaagct ccggttttga actatgcgct cggggttggc gagtgtgttt
     3241 tgtgaagttt tttaggcacc ttttgaaatg taatcatttg ggtcaatatg taattttcag
     3301 tgttagactt gtaaattgtc cgctaaattc tggccgtttt tggctttttt gttagacaac
     3361 atgaccgagt acaagcccac ggtgcgcctc gccacccgcg acgacgtccc cagggccgta
     3421 cgcaccctcg ccgccgcgtt cgccgactac cccgccacgc gccacaccgt cgatccggac
     3481 cgccacatcg agcgggtcac cgagctgcaa gaactcttcc tcacgcgcgt cgggctcgac
     3541 atcggcaagg tgtgggtcgc ggacgacggc gccgcggtgg cggtctggac cacgccggag
     3601 agcgtcgaag cgggggcggt gttcgccgag atcggcccgc gcatggccga gttgagcggt
     3661 tcccggctgg ccgcgcagca acagatggag ggcctcctgg cgccgcaccg gcccaaggag
     3721 cccgcgtggt tcctggccac cgtcggcgtc tcgcccgacc accagggcaa gggtctgggc
     3781 agcgccgtcg tgctccccgg agtggaggcc gccgagcgcg ccggggtgcc cgccttcctg
     3841 gagacctccg cgccccgcaa cctccccttc tacgagcggc tcggcttcac cgtcaccgcc
     3901 gacgtcgagg tgcccgaagg accgcgcacc tggtgcatga cccgcaagcc cggtgcctag
     3961 gctagtatgt aagcctagtc ttagataata aaatcgctat ccatcgaaga tggatgtgtg
     4021 ttggtttttt gtgtgtgtaa cgctaggcgc gcctggtgta ccgagaacga tcctctcagt
     4081 gcgagtctcg acgatccata tcgttgcttg gcagtcagcc agtcggaatc cagcttggga
     4141 cccaggaagt ccaatcgtca gatattgtac tcaagcctgg tcacggcagc gtaccgatct
     4201 gtttaaacct agatattgat agtctgatcg gtcaacgtat aatcgagtcc tagcttttgc
     4261 aaacatctat caagagacag gatcagcagg aggctttcgc atgagtattc aacatttccg
     4321 tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc acccagaaac
     4381 gctggtgaaa gtaaaagatg ctgaagatca gttgggtgcg cgagtgggtt acatcgaact
     4441 ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgct ttccaatgat
     4501 gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga
     4561 gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtatt caccagtcac
     4621 agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg ccataaccat
     4681 gagtgataac actgcggcca acttacttct gacaacgatt ggaggaccga aggagctaac
     4741 cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct
     4801 gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgtagcaa tggcaacaac
     4861 cttgcgtaaa ctattaactg gcgaactact tactctagct tcccggcaac agttgataga
     4921 ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg
     4981 gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca ttgcagcact
     5041 ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac
     5101 tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta
     5161 accgattcta ggtgcattgg cgcagaaaaa aatgcctgat gcgacgctgc gcgtcttata
     5221 ctcccacata tgccagattc agcaacggat acggcttccc caacttgccc acttccatac
     5281 gtgtcctcct taccagaaat ttatccttaa gatcccgaat cgtttaaact cgactctggc
     5341 tctatcgaat ctccgtcgtt tcgagcttac gcgaacagcc gtggcgctca tttgctcgtc
     5401 gggcatcgaa tctcgtcagc tatcgtcagc ttaccttttt ggcagcgatc gcggctcccg
     5461 acatcttgga ccattagctc cacaggtatc ttcttccctc tagtggtcat aacagcagct
     5521 tcagctacct ctcaattcaa aaaacccctc aagacccgtt tagaggcccc aaggggttat
     5581 gctatcaatc gttgcgttac acacacaaaa aaccaacaca catccatctt cgatggatag
     5641 cgattttatt atctaactgc tgatcgagtg tagccagatc tagtaatcaa ttacggggtc
     5701 attagttcat agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc
     5761 tggctgaccg cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt
     5821 aacgccaata gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca
     5881 cttggcagta catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg
     5941 taaatggccc gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca
     6001 gtacatctac gtattagtca tcgctattac catgctgatg cggttttggc agtacatcaa
     6061 tgggcgtgga tagcggtttg actcacgggg atttccaagt ctccacccca ttgacgtcaa
     6121 tgggagtttg ttttggcacc aaaatcaacg ggactttcca aaatgtcgta acaactccgc
     6181 cccattgacg caaatgggcg gtaggcgtgt acggtgggag gtctatataa gcagagctgg
     6241 tttagtgaac cgtcagatca gatctttgtc gatcctacca tccactcgac acacccgcca
     6301 gcggccgc
//
