LOCUS       sequence-295626-        7298 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression plasmid for SARS-CoV-2 M (membrane).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7298)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 7298)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: pGBW-m4134499
FEATURES             Location/Qualifiers
     source          1..7298
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             671..691
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             692..769
                     /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    838..959
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(849..868)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     903..922
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      1074..1120
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1150..1286
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1331..1402
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(1628..2216)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1708..1727)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2716..3925
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     3762..3781
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             3929..4957
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     polyA_signal    4984..5032
                     /note="synthetic polyadenylation signal"
     CDS             join(5298..5366,5367..6158)
                     /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(5516..5535)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      6171..6307
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    6452..6523
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(6528..6574)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    6592..6640
                     /note="synthetic polyadenylation signal"
     enhancer        6721..7024
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        7026..7228
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     7179..7199
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      join(7297..7298,1..8)
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
ORIGIN
        1 caccatggcg gacagcaacg gcactattac agtcgaagag cttaagaaat tactcgagca
       61 gtggaatttg gtaataggat tcctgtttct aacgtggatc tgtctgctgc aattcgcata
      121 tgccaaccgg aatagatttc tctacatcat taagttgatc ttcctgtggc tgctttggcc
      181 tgtgaccttg gcttgctttg ttctggccgc cgtgtaccgc ataaactgga ttaccggtgg
      241 gatcgctatc gcaatggcgt gcctcgtggg ccttatgtgg ctgtcctatt ttatagcctc
      301 attccgactc tttgctcgta ctaggagtat gtggtctttc aatcccgaaa caaatatttt
      361 actaaacgtc ccgctgcatg ggaccatttt aacaagacca ttgctagaga gcgagctcgt
      421 gatcggcgca gttatactgc ggggacacct caggatcgcc ggacaccatc tgggtcgctg
      481 tgatattaaa gacctgccca aggaaataac agtggctacc tcgcgtactc tttcatacta
      541 taaactggga gccagccaga gagtggccgg ggattctggc ttcgcagcat atagtcgata
      601 caggatcggg aactacaagc tgaatacgga ccactcctcc tctagtgata atattgcatt
      661 gctcgtacag gaaaaccttt actttcagtc gaagaggcga tggaaaaaga attttatcgc
      721 ggtgagtgca gccaacagat tcaaaaagat atcctcgtct ggtgccctct aaggatccgc
      781 aggcctctgc tagcttgact gactgagata cagcgtacct tcagctcaca gacatgataa
      841 gatacattga tgagtttgga caaaccacaa ctagaatgca gtgaaaaaaa tgctttattt
      901 gtgaaatttg tgatgctatt gctttatttg taaccattat aagctgcaat aaacaagtta
      961 acaacaacaa ttgcattcat tttatgtttc aggttcaggg ggaggtgtgg gaggtttttt
     1021 aaagcaagta aaacctctac aaatgtggta ttggcccatc tctatcggta tcgtagcata
     1081 accccttggg gcctctaaac gggtcttgag gggttttttg tgcccctcgg gccggattgc
     1141 tatctaccgg cattggcgca gaaaaaaatg cctgatgcga cgctgcgcgt cttatactcc
     1201 cacatatgcc agattcagca acggatacgg cttccccaac ttgcccactt ccatacgtgt
     1261 cctccttacc agaaatttat ccttaaggtc gtcagctatc ctgcaggcga tctctcgatt
     1321 tcgatcaaga cattccttta atggtctttt ctggacacca ctaggggtca gaagtagttc
     1381 atcaaacttt cttccctccc taatctcatt ggttaccttg ggctatcgaa acttaattaa
     1441 ccagtcaagt cagctacttg gcgagatcga cttgtctggg tttcgactac gctcagaatt
     1501 gcgtcagtca agttcgatct ggtccttgct attgcacccg ttctccgatt acgagtttca
     1561 tttaaatcat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
     1621 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
     1681 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
     1741 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
     1801 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg
     1861 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
     1921 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
     1981 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
     2041 ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc
     2101 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta
     2161 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
     2221 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga
     2281 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa
     2341 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa
     2401 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcatttaaat
     2461 ttccgaactc tccaaggccc tcgtcggaaa atcttcaaac ctttcgtccg atccatcttg
     2521 caggctacct ctcgaacgaa ctatcgcaag tctcttggcc ggccttgcgc cttggctatt
     2581 gcttggcagc gcctatcgcc aggtattact ccaatcccga atatccgaga tcgggatcac
     2641 ccgagagaag ttcaacctac atcctcaatc ccgatctatc cgagatccga ggaatatcga
     2701 aatcggggcg cgcctggcct ccgcgccggg ttttggcgcc tcccgcgggc gcccccctcg
     2761 tcacggcgag cgctgccacg tcagacgaag ggcgcaggag cgtcctgatc cttccgcccg
     2821 gacgctcagg acagcggccc gctgctcata agactcggcc ttagaacccc agtatcagca
     2881 gaaggacatt ttaggacggg acttgggtga ctctagggca ctggttttct ttccagagag
     2941 cggaacaggc gaggaaaagt agtcccttct cggcgattct gcggagggat ctccgtgggg
     3001 cggtgaacgc cgatgattat ataaggacgc gccgggtgtg gcacagctag ttccgtcgca
     3061 gccgggattt gggtcgcggt tcttgtttgt ggatcgctgt gatcgtcact tggtgagtag
     3121 cgggctgctg ggctggccgg ggctttcgtg gccgccgggc cgctcggtgg gacggaagcg
     3181 tgtggagaga ccgccaaggg ctgtagtctg ggtccgcgag caaggttgcc ctgaactggg
     3241 ggttgggggg agcgcagcaa aatggcggct gttcccgagt cttgaatgga agacgcttgt
     3301 gaggcgggct gtgaggtcgt tgaaacaagg tggggggcat ggtgggcggc aagaacccaa
     3361 ggtcttgagc ccttcgctaa tgcgggaaag ctcttattcg ggtgagatgg gctgggcacc
     3421 atctggggac cctgacgtga agtttgtcac tgactggaga actcggtttg tcgtctgttg
     3481 cgggggcggc agttatggcg gtgccgttgg gcagtgcacc cgtacctttg ggagcgcgcg
     3541 ccctcgtcgt gtcgtgacgt cacccgttct gttggcttat aatgcagggt ggggccacct
     3601 gccggtaggt gtgcggtagg cttttctccg tcgcaggacg cagggttcgg gcctagggta
     3661 ggctctcctg aatcgacagg cgccggacct ctggtgaggg gagggataag tgaggcgtca
     3721 gtttctttgg tcggttttat gtacctatct tcttaagtag ctgaagctcc ggttttgaac
     3781 tatgcgctcg gggttggcga gtgtgttttg tgaagttttt taggcacctt ttgaaatgta
     3841 atcatttggg tcaatatgta attttcagtg ttagacttgt aaattgtccg ctaaattctg
     3901 gccgtttttg gcttttttgt tagacaacat gggtaaaaag cctgaactca ccgcgacgtc
     3961 tgtcgagaag tttctgatcg aaaagttcga cagcgtctcc gacctgatgc agctctcgga
     4021 gggcgaagaa tctcgtgctt tcagcttcga tgtaggaggg cgtggatatg tcctgcgggt
     4081 aaatagctgc gccgatggtt tctacaaaga tcgttatgtt tatcggcact ttgcatccgc
     4141 cgcgctcccg attccggaag tgcttgacat tggggagttc agcgagagcc tgacctattg
     4201 catctcccgc cgtgcacagg gtgtcacgtt gcaagacctg cctgaaaccg aactgcccgc
     4261 tgttctgcag ccggtcgcgg aggcaatgga tgccatcgct gccgccgatc ttagccagac
     4321 gagcgggttc ggcccattcg gaccgcaagg aatcggtcaa tacactacat ggcgtgattt
     4381 catatgcgcg attgctgatc cccatgtgta tcactggcaa actgtgatgg acgacaccgt
     4441 cagtgcgtcc gtcgcgcagg ctctcgatga gctgatgctt tgggccgagg actgccccga
     4501 agtccggcac ctcgtgcacg cggatttcgg ctccaacaat gtcctgacgg acaatggccg
     4561 cataacagcg gtcattgact ggagcgaggc gatgttcggg gattcccaat acgaggtcgc
     4621 caacatcttc ttctggaggc cgtggttggc ttgtatggag cagcagacgc gctacttcga
     4681 gcggaggcat ccggagcttg caggatcgcc ccggctccgg gcgtatatgc tccgcattgg
     4741 tcttgaccaa ctctatcaga gcttggttga cggcaatttc gatgatgcag cttgggcgca
     4801 gggtcgatgc gacgcaatcg tccgatccgg agccgggact gtcgggcgta cacaaatcgc
     4861 ccgcagaagc gccgccgtct ggaccgatgg ctgtgtagaa gtactcgccg atagtggaaa
     4921 ccgacgcccc agcactcgtc cgagggcaaa ggaataagct agtatgtaag cctagtctta
     4981 gataataaaa tcgctatcca tcgaagatgg atgtgtgttg gttttttgtg tgtgtaacgc
     5041 taggcgcgcc tggtgtaccg agaacgatcc tctcagtgcg agtctcgacg atccatatcg
     5101 ttgcttggca gtcagccagt cggaatccag cttgggaccc aggaagtcca atcgtcagat
     5161 attgtactca agcctggtca cggcagcgta ccgatctgtt taaacctaga tattgatagt
     5221 ctgatcggtc aacgtataat cgagtcctag cttttgcaaa catctatcaa gagacaggat
     5281 cagcaggagg ctttcgcatg agtattcaac atttccgtgt cgcccttatt cccttttttg
     5341 cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg
     5401 aagatcagtt gggtgcgcga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc
     5461 ttgagagttt tcgccccgaa gaacgctttc caatgatgag cacttttaaa gttctgctat
     5521 gtggcgcggt attatcccgt attgacgccg ggcaagagca actcggtcgc cgcatacact
     5581 attctcagaa tgacttggtt gagtattcac cagtcacaga aaagcatctt acggatggca
     5641 tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact
     5701 tacttctgac aacgattgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg
     5761 atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg
     5821 agcgtgacac cacgatgcct gtagcaatgg caacaacctt gcgtaaacta ttaactggcg
     5881 aactacttac tctagcttcc cggcaacagt tgatagactg gatggaggcg gataaagttg
     5941 caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag
     6001 ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc
     6061 gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga
     6121 tcgctgagat aggtgcctca ctgattaagc attggtaacc gattctaggt gcattggcgc
     6181 agaaaaaaat gcctgatgcg acgctgcgcg tcttatactc ccacatatgc cagattcagc
     6241 aacggatacg gcttccccaa cttgcccact tccatacgtg tcctccttac cagaaattta
     6301 tccttaagat cgtttaaact cgactctggc tctatcgaat ctccgtcgtt tcgagcttac
     6361 gcgaacagcc gtggcgctca tttgctcgtc gggcatcgaa tctcgtcagc tatcgtcagc
     6421 ttaccttttt ggcagcgatc gcggctcccg acatcttgga ccattagctc cacaggtatc
     6481 ttcttccctc tagtggtcat aacagcagct tcagctacct ctcaattcaa aaaacccctc
     6541 aagacccgtt tagaggcccc aaggggttat gctatcaatc gttgcgttac acacacaaaa
     6601 aaccaacaca catccatctt cgatggatag cgattttatt atctaactgc tgatcgagtg
     6661 tagccagatc tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
     6721 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
     6781 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
     6841 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
     6901 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
     6961 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
     7021 catgctgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
     7081 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
     7141 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
     7201 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatca gatctttgtc
     7261 gatcctacca tccactcgac acacccgcca gcggccgc
//
