LOCUS       sequence-295922-        7330 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 7330)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 7330)
  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-m4133798
FEATURES             Location/Qualifiers
     source          1..7330
                     /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             725..802
                     /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"
     CDS             830..850
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     polyA_signal    1299..1420
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1310..1329)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1364..1383
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      1535..1581
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1611..1747
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1792..1863
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(2089..2677)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2169..2188)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        3177..4386
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     4223..4242
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             4390..4989
                     /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(4390..4409)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4886..4906
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    5016..5064
                     /note="synthetic polyadenylation signal"
     CDS             join(5330..5398,5399..6190)
                     /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(5548..5567)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      6203..6339
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    6484..6555
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(6560..6606)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    6624..6672
                     /note="synthetic polyadenylation signal"
     enhancer        6753..7056
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        7058..7260
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     7211..7231
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      join(7329..7330,1..8)
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
ORIGIN
        1 caccatggca gattccaacg gtactattac cgttgaagag cttaaaaagc tccttgaaca
       61 atggaaccta gtaataggtt tcctattcct tacatggatt tgtcttctac aatttgccta
      121 tgccaacagg aataggtttt tgtatataat taagttaatt ttcctctggc tgttatggcc
      181 agtaacttta gcttgttttg tgcttgctgc tgtttacaga ataaattgga tcaccggtgg
      241 aattgctatc gcaatggctt gtcttgtagg cttgatgtgg ctcagctact tcattgcttc
      301 tttcagactg tttgcgcgta cgcgttccat gtggtcattc aatccagaaa ctaacattct
      361 tctcaacgtg ccactccatg gcactattct gaccagaccg cttctagaaa gtgaactcgt
      421 aatcggagct gtgatccttc gtggacatct tcgtattgct ggacaccatc taggacgctg
      481 tgacatcaag gacctgccta aagaaatcac tgttgctaca tcacgaacgc tttcttatta
      541 caaattggga gcttcgcagc gtgtagcagg tgactcaggt tttgctgcat acagtcgcta
      601 caggattggc aactataaat taaacacaga ccattccagt agcagtgaca atattgcttt
      661 gcttgtacag gaaaaccttt actttcagtc gggacggcga atcccggggc tgataaatcc
      721 ctggaaaagg agatggaaga agaactttat tgcagtgtct gcggctaacc ggttcaaaaa
      781 gatcagttca agcggcgccc tagattacga cattcctacg accgcctccg agaatctcta
      841 tttccagggt gaatttggct tggcacaaca tgacgaggcc gtcgataaca agtttaataa
      901 agaacagcag aacgctttct atgagatcct tcacttacca aatctgaacg aagagcaacg
      961 caacgctttc attcagagcc tgaaagacga tccctcgcag tctgcaaatc tcctggccga
     1021 agccaagaag ctgaatgacg cacaggctcc aaaagttgat aacaagttta acaaggagca
     1081 gcagaatgct ttctacgaga tattgcacct tcctaatctg aatgaggaac aaagaaatgc
     1141 atttatccag tccctaaaag acgatccatc acagagtgcc aacctgcttg cggaagccaa
     1201 aaagctcaat gatgcccagg ccccgaaagt agacgcgaac catcaataag ctagcttgac
     1261 tgactgagat acagcgtacc ttcagctcac agacatgata agatacattg atgagtttgg
     1321 acaaaccaca actagaatgc agtgaaaaaa atgctttatt tgtgaaattt gtgatgctat
     1381 tgctttattt gtaaccatta taagctgcaa taaacaagtt aacaacaaca attgcattca
     1441 ttttatgttt caggttcagg gggaggtgtg ggaggttttt taaagcaagt aaaacctcta
     1501 caaatgtggt attggcccat ctctatcggt atcgtagcat aaccccttgg ggcctctaaa
     1561 cgggtcttga ggggtttttt gtgcccctcg ggccggattg ctatctaccg gcattggcgc
     1621 agaaaaaaat gcctgatgcg acgctgcgcg tcttatactc ccacatatgc cagattcagc
     1681 aacggatacg gcttccccaa cttgcccact tccatacgtg tcctccttac cagaaattta
     1741 tccttaaggt cgtcagctat cctgcaggcg atctctcgat ttcgatcaag acattccttt
     1801 aatggtcttt tctggacacc actaggggtc agaagtagtt catcaaactt tcttccctcc
     1861 ctaatctcat tggttacctt gggctatcga aacttaatta accagtcaag tcagctactt
     1921 ggcgagatcg acttgtctgg gtttcgacta cgctcagaat tgcgtcagtc aagttcgatc
     1981 tggtccttgc tattgcaccc gttctccgat tacgagtttc atttaaatca tgtgagcaaa
     2041 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     2101 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     2161 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     2221 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     2281 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     2341 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     2401 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     2461 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     2521 cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
     2581 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
     2641 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
     2701 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
     2761 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag
     2821 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc
     2881 agcgatctgt ctatttcgtt catccatagt tgcatttaaa tttccgaact ctccaaggcc
     2941 ctcgtcggaa aatcttcaaa cctttcgtcc gatccatctt gcaggctacc tctcgaacga
     3001 actatcgcaa gtctcttggc cggccttgcg ccttggctat tgcttggcag cgcctatcgc
     3061 caggtattac tccaatcccg aatatccgag atcgggatca cccgagagaa gttcaaccta
     3121 catcctcaat cccgatctat ccgagatccg aggaatatcg aaatcggggc gcgcctggcc
     3181 tccgcgccgg gttttggcgc ctcccgcggg cgcccccctc gtcacggcga gcgctgccac
     3241 gtcagacgaa gggcgcagga gcgtcctgat ccttccgccc ggacgctcag gacagcggcc
     3301 cgctgctcat aagactcggc cttagaaccc cagtatcagc agaaggacat tttaggacgg
     3361 gacttgggtg actctagggc actggttttc tttccagaga gcggaacagg cgaggaaaag
     3421 tagtcccttc tcggcgattc tgcggaggga tctccgtggg gcggtgaacg ccgatgatta
     3481 tataaggacg cgccgggtgt ggcacagcta gttccgtcgc agccgggatt tgggtcgcgg
     3541 ttcttgtttg tggatcgctg tgatcgtcac ttggtgagta gcgggctgct gggctggccg
     3601 gggctttcgt ggccgccggg ccgctcggtg ggacggaagc gtgtggagag accgccaagg
     3661 gctgtagtct gggtccgcga gcaaggttgc cctgaactgg gggttggggg gagcgcagca
     3721 aaatggcggc tgttcccgag tcttgaatgg aagacgcttg tgaggcgggc tgtgaggtcg
     3781 ttgaaacaag gtggggggca tggtgggcgg caagaaccca aggtcttgag cccttcgcta
     3841 atgcgggaaa gctcttattc gggtgagatg ggctgggcac catctgggga ccctgacgtg
     3901 aagtttgtca ctgactggag aactcggttt gtcgtctgtt gcgggggcgg cagttatggc
     3961 ggtgccgttg ggcagtgcac ccgtaccttt gggagcgcgc gccctcgtcg tgtcgtgacg
     4021 tcacccgttc tgttggctta taatgcaggg tggggccacc tgccggtagg tgtgcggtag
     4081 gcttttctcc gtcgcaggac gcagggttcg ggcctagggt aggctctcct gaatcgacag
     4141 gcgccggacc tctggtgagg ggagggataa gtgaggcgtc agtttctttg gtcggtttta
     4201 tgtacctatc ttcttaagta gctgaagctc cggttttgaa ctatgcgctc ggggttggcg
     4261 agtgtgtttt gtgaagtttt ttaggcacct tttgaaatgt aatcatttgg gtcaatatgt
     4321 aattttcagt gttagacttg taaattgtcc gctaaattct ggccgttttt ggcttttttg
     4381 ttagacaaca tgaccgagta caagcccacg gtgcgcctcg ccacccgcga cgacgtcccc
     4441 agggccgtac gcaccctcgc cgccgcgttc gccgactacc ccgccacgcg ccacaccgtc
     4501 gatccggacc gccacatcga gcgggtcacc gagctgcaag aactcttcct cacgcgcgtc
     4561 gggctcgaca tcggcaaggt gtgggtcgcg gacgacggcg ccgcggtggc ggtctggacc
     4621 acgccggaga gcgtcgaagc gggggcggtg ttcgccgaga tcggcccgcg catggccgag
     4681 ttgagcggtt cccggctggc cgcgcagcaa cagatggagg gcctcctggc gccgcaccgg
     4741 cccaaggagc ccgcgtggtt cctggccacc gtcggcgtct cgcccgacca ccagggcaag
     4801 ggtctgggca gcgccgtcgt gctccccgga gtggaggccg ccgagcgcgc cggggtgccc
     4861 gccttcctgg agacctccgc gccccgcaac ctccccttct acgagcggct cggcttcacc
     4921 gtcaccgccg acgtcgaggt gcccgaagga ccgcgcacct ggtgcatgac ccgcaagccc
     4981 ggtgcctagg ctagtatgta agcctagtct tagataataa aatcgctatc catcgaagat
     5041 ggatgtgtgt tggttttttg tgtgtgtaac gctaggcgcg cctggtgtac cgagaacgat
     5101 cctctcagtg cgagtctcga cgatccatat cgttgcttgg cagtcagcca gtcggaatcc
     5161 agcttgggac ccaggaagtc caatcgtcag atattgtact caagcctggt cacggcagcg
     5221 taccgatctg tttaaaccta gatattgata gtctgatcgg tcaacgtata atcgagtcct
     5281 agcttttgca aacatctatc aagagacagg atcagcagga ggctttcgca tgagtattca
     5341 acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca
     5401 cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcgc gagtgggtta
     5461 catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgctt
     5521 tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc
     5581 cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtattc
     5641 accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc
     5701 cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgattg gaggaccgaa
     5761 ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga
     5821 accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat
     5881 ggcaacaacc ttgcgtaaac tattaactgg cgaactactt actctagctt cccggcaaca
     5941 gttgatagac tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc
     6001 ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat
     6061 tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag
     6121 tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa
     6181 gcattggtaa ccgattctag gtgcattggc gcagaaaaaa atgcctgatg cgacgctgcg
     6241 cgtcttatac tcccacatat gccagattca gcaacggata cggcttcccc aacttgccca
     6301 cttccatacg tgtcctcctt accagaaatt tatccttaag atcgtttaaa ctcgactctg
     6361 gctctatcga atctccgtcg tttcgagctt acgcgaacag ccgtggcgct catttgctcg
     6421 tcgggcatcg aatctcgtca gctatcgtca gcttaccttt ttggcagcga tcgcggctcc
     6481 cgacatcttg gaccattagc tccacaggta tcttcttccc tctagtggtc ataacagcag
     6541 cttcagctac ctctcaattc aaaaaacccc tcaagacccg tttagaggcc ccaaggggtt
     6601 atgctatcaa tcgttgcgtt acacacacaa aaaaccaaca cacatccatc ttcgatggat
     6661 agcgatttta ttatctaact gctgatcgag tgtagccaga tctagtaatc aattacgggg
     6721 tcattagttc atagcccata tatggagttc cgcgttacat aacttacggt aaatggcccg
     6781 cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta tgttcccata
     6841 gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg gtaaactgcc
     6901 cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga cgtcaatgac
     6961 ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt tcctacttgg
     7021 cagtacatct acgtattagt catcgctatt accatgctga tgcggttttg gcagtacatc
     7081 aatgggcgtg gatagcggtt tgactcacgg ggatttccaa gtctccaccc cattgacgtc
     7141 aatgggagtt tgttttggca ccaaaatcaa cgggactttc caaaatgtcg taacaactcc
     7201 gccccattga cgcaaatggg cggtaggcgt gtacggtggg aggtctatat aagcagagct
     7261 ggtttagtga accgtcagat cagatctttg tcgatcctac catccactcg acacacccgc
     7321 cagcggccgc
//
