LOCUS       sequence-300142-        6864 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Mammalian expression plasmid for SARS-CoV-2 E (envelope).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6864)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 6864)
  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-m4252954
FEATURES             Location/Qualifiers
     source          1..6864
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             230..250
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             251..328
                     /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    397..518
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(408..427)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     462..481
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      633..679
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      709..845
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    890..961
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(1187..1775)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1267..1286)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2275..3484
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     3321..3340
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             3488..4516
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     polyA_signal    4543..4591
                     /note="synthetic polyadenylation signal"
     CDS             join(4857..4925,4926..5717)
                     /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(5075..5094)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      5730..5866
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    6018..6089
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(6094..6140)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    6158..6206
                     /note="synthetic polyadenylation signal"
     enhancer        6287..6590
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6592..6794
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     6745..6765
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
ORIGIN
        1 caccatgtac agctttgtat ctgaagagac aggcactctc atcgttaatt cagtgttgct
       61 gttcctggca tttgtggtct tccttttagt gaccctagcg atactgacgg ccctccggct
      121 gtgcgcttat tgttgtaaca ttgtcaacgt tagtctcgtg aagccctcgt tctacgttta
      181 ttcccgcgtg aaaaatctaa acagctcccg tgtgcctgat cttcttgtcg aaaaccttta
      241 ctttcagtcg aagaggcgat ggaaaaagaa ttttatcgcg gtgagtgcag ccaacagatt
      301 caaaaagata tcctcgtctg gtgccctcta aggatccgca ggcctctgct agcttgactg
      361 actgagatac agcgtacctt cagctcacag acatgataag atacattgat gagtttggac
      421 aaaccacaac tagaatgcag tgaaaaaaat gctttatttg tgaaatttgt gatgctattg
      481 ctttatttgt aaccattata agctgcaata aacaagttaa caacaacaat tgcattcatt
      541 ttatgtttca ggttcagggg gaggtgtggg aggtttttta aagcaagtaa aacctctaca
      601 aatgtggtat tggcccatct ctatcggtat cgtagcataa ccccttgggg cctctaaacg
      661 ggtcttgagg ggttttttgt gcccctcggg ccggattgct atctaccggc attggcgcag
      721 aaaaaaatgc ctgatgcgac gctgcgcgtc ttatactccc acatatgcca gattcagcaa
      781 cggatacggc ttccccaact tgcccacttc catacgtgtc ctccttacca gaaatttatc
      841 cttaaggtcg tcagctatcc tgcaggcgat ctctcgattt cgatcaagac attcctttaa
      901 tggtcttttc tggacaccac taggggtcag aagtagttca tcaaactttc ttccctccct
      961 aatctcattg gttaccttgg gctatcgaaa cttaattaac cagtcaagtc agctacttgg
     1021 cgagatcgac ttgtctgggt ttcgactacg ctcagaattg cgtcagtcaa gttcgatctg
     1081 gtccttgcta ttgcacccgt tctccgatta cgagtttcat ttaaatcatg tgagcaaaag
     1141 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc
     1201 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
     1261 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
     1321 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
     1381 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
     1441 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
     1501 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
     1561 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
     1621 ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
     1681 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
     1741 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg
     1801 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa
     1861 aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta
     1921 tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag
     1981 cgatctgtct atttcgttca tccatagttg catttaaatt tccgaactct ccaaggccct
     2041 cgtcggaaaa tcttcaaacc tttcgtccga tccatcttgc aggctacctc tcgaacgaac
     2101 tatcgcaagt ctcttggccg gccttgcgcc ttggctattg cttggcagcg cctatcgcca
     2161 ggtattactc caatcccgaa tatccgagat cgggatcacc cgagagaagt tcaacctaca
     2221 tcctcaatcc cgatctatcc gagatccgag gaatatcgaa atcggggcgc gcctggcctc
     2281 cgcgccgggt tttggcgcct cccgcgggcg cccccctcgt cacggcgagc gctgccacgt
     2341 cagacgaagg gcgcaggagc gtcctgatcc ttccgcccgg acgctcagga cagcggcccg
     2401 ctgctcataa gactcggcct tagaacccca gtatcagcag aaggacattt taggacggga
     2461 cttgggtgac tctagggcac tggttttctt tccagagagc ggaacaggcg aggaaaagta
     2521 gtcccttctc ggcgattctg cggagggatc tccgtggggc ggtgaacgcc gatgattata
     2581 taaggacgcg ccgggtgtgg cacagctagt tccgtcgcag ccgggatttg ggtcgcggtt
     2641 cttgtttgtg gatcgctgtg atcgtcactt ggtgagtagc gggctgctgg gctggccggg
     2701 gctttcgtgg ccgccgggcc gctcggtggg acggaagcgt gtggagagac cgccaagggc
     2761 tgtagtctgg gtccgcgagc aaggttgccc tgaactgggg gttgggggga gcgcagcaaa
     2821 atggcggctg ttcccgagtc ttgaatggaa gacgcttgtg aggcgggctg tgaggtcgtt
     2881 gaaacaaggt ggggggcatg gtgggcggca agaacccaag gtcttgagcc cttcgctaat
     2941 gcgggaaagc tcttattcgg gtgagatggg ctgggcacca tctggggacc ctgacgtgaa
     3001 gtttgtcact gactggagaa ctcggtttgt cgtctgttgc gggggcggca gttatggcgg
     3061 tgccgttggg cagtgcaccc gtacctttgg gagcgcgcgc cctcgtcgtg tcgtgacgtc
     3121 acccgttctg ttggcttata atgcagggtg gggccaccag ccggtaggtg tgcggtaggc
     3181 ttttctccgt cgcaggacgc agggttcggg cctagggtag gctctcctga atcgacaggc
     3241 gccggacctc tggtgagggg agggataagt gaggcgtcag tttctttggt cggttttatg
     3301 tacctatctt cttaagtagc tgaagctccg gttttgaact atgcgctcgg ggttggcgag
     3361 tgtgttttgt gaagtttttt aggcaccttt tgaaatgtaa tcatttgggt caatatgtaa
     3421 ttttcagtgt tagacttgta aattgtccgc taaattctgg ccgtttttgg cttttttgtt
     3481 agacaacatg ggtaaaaagc ctgaactcac cgcgacgtct gtcgagaagt ttctgatcga
     3541 aaagttcgac agcgtctccg acctgatgca gctctcggag ggcgaagaat ctcgtgcttt
     3601 cagcttcgat gtaggagggc gtggatatgt cctgcgggta aatagctgcg ccgatggttt
     3661 ctacaaagat cgttatgttt atcggcactt tgcatccgcc gcgctcccga ttccggaagt
     3721 gcttgacatt ggggagttca gcgagagcct gacctattgc atctcccgcc gtgcacaggg
     3781 tgtcacgttg caagacctgc ctgaaaccga actgcccgct gttctgcagc cggtcgcgga
     3841 ggcaatggat gccatcgctg ccgccgatct tagccagacg agcgggttcg gcccattcgg
     3901 accgcaagga atcggtcaat acactacatg gcgtgatttc atatgcgcga ttgctgatcc
     3961 ccatgtgtat cactggcaaa ctgtgatgga cgacaccgtc agtgcgtccg tcgcgcaggc
     4021 tctcgatgag ctgatgcttt gggccgagga ctgccccgaa gtccggcacc tcgtgcacgc
     4081 ggatttcggc tccaacaatg tcctgacgga caatggccgc ataacagcgg tcattgactg
     4141 gagcgaggcg atgttcgggg attcccaata cgaggtcgcc aacatcttct tctggaggcc
     4201 gtggttggct tgtatggagc agcagacgcg ctacttcgag cggaggcatc cggagcttgc
     4261 aggatcgccc cggctccggg cgtatatgct ccgcattggt cttgaccaac tctatcagag
     4321 cttggttgac ggcaatttcg atgatgcagc ttgggcgcag ggtcgatgcg acgcaatcgt
     4381 ccgatccgga gccgggactg tcgggcgtac acaaatcgcc cgcagaagcg ccgccgtctg
     4441 gaccgatggc tgtgtagaag tactcgccga tagtggaaac cgacgcccca gcactcgtcc
     4501 gagggcaaag gaataagcta gtatgtaagc ctagtcttag ataataaaat cgctatccat
     4561 cgaagatgga tgtgtgttgg ttttttgtgt gtgtaacgct aggcgcgcct ggtgtaccga
     4621 gaacgatcct ctcagtgcga gtctcgacga tccatatcgt tgcttggcag tcagccagtc
     4681 ggaatccagc ttgggaccca ggaagtccaa tcgtcagata ttgtactcaa gcctggtcac
     4741 ggcagcgtac cgatctgttt aaacctagat attgatagtc tgatcggtca acgtataatc
     4801 gagtcctagc ttttgcaaac atctatcaag agacaggatc agcaggaggc tttcgcatga
     4861 gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt
     4921 ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcgcgag
     4981 tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag
     5041 aacgctttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta
     5101 ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg
     5161 agtattcacc agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca
     5221 gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca acgattggag
     5281 gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc
     5341 gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg
     5401 tagcaatggc aacaaccttg cgtaaactat taactggcga actacttact ctagcttccc
     5461 ggcaacagtt gatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg
     5521 cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg
     5581 gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga
     5641 cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac
     5701 tgattaagca ttggtaaccg attctaggtg cattggcgca gaaaaaaatg cctgatgcga
     5761 cgctgcgcgt cttatactcc cacatatgcc agattcagca acggatacgg cttccccaac
     5821 ttgcccactt ccatacgtgt cctccttacc agaaatttat ccttaagatc ccgaatcgtt
     5881 taaactcgac tctggctcta tcgaatctcc gtcgtttcga gcttacgcga acagccgtgg
     5941 cgctcatttg ctcgtcgggc atcgaatctc gtcagctatc gtcagcttac ctttttggca
     6001 gcgatcgcgg ctcccgacat cttggaccat tagctccaca ggtatcttct tccctctagt
     6061 ggtcataaca gcagcttcag ctacctctca attcaaaaaa cccctcaaga cccgtttaga
     6121 ggccccaagg ggttatgcta tcaatcgttg cgttacacac acaaaaaacc aacacacatc
     6181 catcttcgat ggatagcgat tttattatct aactgctgat cgagtgtagc cagatctagt
     6241 aatcaattac ggggtcatta gttcatagcc catatatgga gttccgcgtt acataactta
     6301 cggtaaatgg cccgcctggc tgaccgccca acgacccccg cccattgacg tcaataatga
     6361 cgtatgttcc catagtaacg ccaataggga ctttccattg acgtcaatgg gtggagtatt
     6421 tacggtaaac tgcccacttg gcagtacatc aagtgtatca tatgccaagt acgcccccta
     6481 ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc ccagtacatg accttatggg
     6541 actttcctac ttggcagtac atctacgtat tagtcatcgc tattaccatg ctgatgcggt
     6601 tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt ccaagtctcc
     6661 accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat
     6721 gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct
     6781 atataagcag agctggttta gtgaaccgtc agatcagatc tttgtcgatc ctaccatcca
     6841 ctcgacacac ccgccagcgg ccgc
//
