LOCUS       sequence-295651-        7165 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 7165)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 7165)
  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-m4134542
FEATURES             Location/Qualifiers
     source          1..7165
                     /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             1040..1057
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     polyA_signal    1134..1255
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1145..1164)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1199..1218
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      1370..1416
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1446..1582
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1627..1698
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(1924..2512)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2004..2023)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        3012..4221
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     4058..4077
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             4225..4824
                     /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(4225..4244)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4721..4741
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4851..4899
                     /note="synthetic polyadenylation signal"
     CDS             join(5165..5233,5234..6025)
                     /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(5383..5402)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      6038..6174
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    6319..6390
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(6395..6441)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    6459..6507
                     /note="synthetic polyadenylation signal"
     enhancer        6588..6891
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6893..7095
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     7046..7066
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      join(7164..7165,1..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 gatgagtgac aaaatcattc atctaaccga
      721 tgactccttt gatacagacg tgttgaaggc cgatggtgcg atactggtag atttctgggc
      781 tgaatggtgt ggcccgtgca agatgatcgc acccattctg gacgagatcg ccgacgagta
      841 tcaaggaaaa ttaactgtcg ctaagcttaa tatagatcag aacccaggga cggcacctaa
      901 atacggcatc aggggaatac ctaccctcct gctctttaag aatggggaag ttgccgcgac
      961 aaaagtggga gccctgtcta agggccagct aaaagagttc ttggacgcta acctggcagg
     1021 ttcaggctcg gggcacatgc accatcacca tcaccatagc agcgggctcg tgccacgcgg
     1081 ctaagctagc ttgactgact gagatacagc gtaccttcag ctcacagaca tgataagata
     1141 cattgatgag tttggacaaa ccacaactag aatgcagtga aaaaaatgct ttatttgtga
     1201 aatttgtgat gctattgctt tatttgtaac cattataagc tgcaataaac aagttaacaa
     1261 caacaattgc attcatttta tgtttcaggt tcagggggag gtgtgggagg ttttttaaag
     1321 caagtaaaac ctctacaaat gtggtattgg cccatctcta tcggtatcgt agcataaccc
     1381 cttggggcct ctaaacgggt cttgaggggt tttttgtgcc cctcgggccg gattgctatc
     1441 taccggcatt ggcgcagaaa aaaatgcctg atgcgacgct gcgcgtctta tactcccaca
     1501 tatgccagat tcagcaacgg atacggcttc cccaacttgc ccacttccat acgtgtcctc
     1561 cttaccagaa atttatcctt aaggtcgtca gctatcctgc aggcgatctc tcgatttcga
     1621 tcaagacatt cctttaatgg tcttttctgg acaccactag gggtcagaag tagttcatca
     1681 aactttcttc cctccctaat ctcattggtt accttgggct atcgaaactt aattaaccag
     1741 tcaagtcagc tacttggcga gatcgacttg tctgggtttc gactacgctc agaattgcgt
     1801 cagtcaagtt cgatctggtc cttgctattg cacccgttct ccgattacga gtttcattta
     1861 aatcatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
     1921 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
     1981 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
     2041 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg
     2101 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg
     2161 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
     2221 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
     2281 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
     2341 gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt
     2401 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
     2461 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc
     2521 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt
     2581 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt
     2641 taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag
     2701 tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcat ttaaatttcc
     2761 gaactctcca aggccctcgt cggaaaatct tcaaaccttt cgtccgatcc atcttgcagg
     2821 ctacctctcg aacgaactat cgcaagtctc ttggccggcc ttgcgccttg gctattgctt
     2881 ggcagcgcct atcgccaggt attactccaa tcccgaatat ccgagatcgg gatcacccga
     2941 gagaagttca acctacatcc tcaatcccga tctatccgag atccgaggaa tatcgaaatc
     3001 ggggcgcgcc tggcctccgc gccgggtttt ggcgcctccc gcgggcgccc ccctcgtcac
     3061 ggcgagcgct gccacgtcag acgaagggcg caggagcgtc ctgatccttc cgcccggacg
     3121 ctcaggacag cggcccgctg ctcataagac tcggccttag aaccccagta tcagcagaag
     3181 gacattttag gacgggactt gggtgactct agggcactgg ttttctttcc agagagcgga
     3241 acaggcgagg aaaagtagtc ccttctcggc gattctgcgg agggatctcc gtggggcggt
     3301 gaacgccgat gattatataa ggacgcgccg ggtgtggcac agctagttcc gtcgcagccg
     3361 ggatttgggt cgcggttctt gtttgtggat cgctgtgatc gtcacttggt gagtagcggg
     3421 ctgctgggct ggccggggct ttcgtggccg ccgggccgct cggtgggacg gaagcgtgtg
     3481 gagagaccgc caagggctgt agtctgggtc cgcgagcaag gttgccctga actgggggtt
     3541 ggggggagcg cagcaaaatg gcggctgttc ccgagtcttg aatggaagac gcttgtgagg
     3601 cgggctgtga ggtcgttgaa acaaggtggg gggcatggtg ggcggcaaga acccaaggtc
     3661 ttgagccctt cgctaatgcg ggaaagctct tattcgggtg agatgggctg ggcaccatct
     3721 ggggaccctg acgtgaagtt tgtcactgac tggagaactc ggtttgtcgt ctgttgcggg
     3781 ggcggcagtt atggcggtgc cgttgggcag tgcacccgta cctttgggag cgcgcgccct
     3841 cgtcgtgtcg tgacgtcacc cgttctgttg gcttataatg cagggtgggg ccacctgccg
     3901 gtaggtgtgc ggtaggcttt tctccgtcgc aggacgcagg gttcgggcct agggtaggct
     3961 ctcctgaatc gacaggcgcc ggacctctgg tgaggggagg gataagtgag gcgtcagttt
     4021 ctttggtcgg ttttatgtac ctatcttctt aagtagctga agctccggtt ttgaactatg
     4081 cgctcggggt tggcgagtgt gttttgtgaa gttttttagg caccttttga aatgtaatca
     4141 tttgggtcaa tatgtaattt tcagtgttag acttgtaaat tgtccgctaa attctggccg
     4201 tttttggctt ttttgttaga caacatgacc gagtacaagc ccacggtgcg cctcgccacc
     4261 cgcgacgacg tccccagggc cgtacgcacc ctcgccgccg cgttcgccga ctaccccgcc
     4321 acgcgccaca ccgtcgatcc ggaccgccac atcgagcggg tcaccgagct gcaagaactc
     4381 ttcctcacgc gcgtcgggct cgacatcggc aaggtgtggg tcgcggacga cggcgccgcg
     4441 gtggcggtct ggaccacgcc ggagagcgtc gaagcggggg cggtgttcgc cgagatcggc
     4501 ccgcgcatgg ccgagttgag cggttcccgg ctggccgcgc agcaacagat ggagggcctc
     4561 ctggcgccgc accggcccaa ggagcccgcg tggttcctgg ccaccgtcgg cgtctcgccc
     4621 gaccaccagg gcaagggtct gggcagcgcc gtcgtgctcc ccggagtgga ggccgccgag
     4681 cgcgccgggg tgcccgcctt cctggagacc tccgcgcccc gcaacctccc cttctacgag
     4741 cggctcggct tcaccgtcac cgccgacgtc gaggtgcccg aaggaccgcg cacctggtgc
     4801 atgacccgca agcccggtgc ctaggctagt atgtaagcct agtcttagat aataaaatcg
     4861 ctatccatcg aagatggatg tgtgttggtt ttttgtgtgt gtaacgctag gcgcgcctgg
     4921 tgtaccgaga acgatcctct cagtgcgagt ctcgacgatc catatcgttg cttggcagtc
     4981 agccagtcgg aatccagctt gggacccagg aagtccaatc gtcagatatt gtactcaagc
     5041 ctggtcacgg cagcgtaccg atctgtttaa acctagatat tgatagtctg atcggtcaac
     5101 gtataatcga gtcctagctt ttgcaaacat ctatcaagag acaggatcag caggaggctt
     5161 tcgcatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct
     5221 tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg
     5281 tgcgcgagtg ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg
     5341 ccccgaagaa cgctttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt
     5401 atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga
     5461 cttggttgag tattcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga
     5521 attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac
     5581 gattggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg
     5641 ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac
     5701 gatgcctgta gcaatggcaa caaccttgcg taaactatta actggcgaac tacttactct
     5761 agcttcccgg caacagttga tagactggat ggaggcggat aaagttgcag gaccacttct
     5821 gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg
     5881 gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat
     5941 ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg
     6001 tgcctcactg attaagcatt ggtaaccgat tctaggtgca ttggcgcaga aaaaaatgcc
     6061 tgatgcgacg ctgcgcgtct tatactccca catatgccag attcagcaac ggatacggct
     6121 tccccaactt gcccacttcc atacgtgtcc tccttaccag aaatttatcc ttaagatcgt
     6181 ttaaactcga ctctggctct atcgaatctc cgtcgtttcg agcttacgcg aacagccgtg
     6241 gcgctcattt gctcgtcggg catcgaatct cgtcagctat cgtcagctta cctttttggc
     6301 agcgatcgcg gctcccgaca tcttggacca ttagctccac aggtatcttc ttccctctag
     6361 tggtcataac agcagcttca gctacctctc aattcaaaaa acccctcaag acccgtttag
     6421 aggccccaag gggttatgct atcaatcgtt gcgttacaca cacaaaaaac caacacacat
     6481 ccatcttcga tggatagcga ttttattatc taactgctga tcgagtgtag ccagatctag
     6541 taatcaatta cggggtcatt agttcatagc ccatatatgg agttccgcgt tacataactt
     6601 acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg
     6661 acgtatgttc ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat
     6721 ttacggtaaa ctgcccactt ggcagtacat caagtgtatc atatgccaag tacgccccct
     6781 attgacgtca atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttatgg
     6841 gactttccta cttggcagta catctacgta ttagtcatcg ctattaccat gctgatgcgg
     6901 ttttggcagt acatcaatgg gcgtggatag cggtttgact cacggggatt tccaagtctc
     6961 caccccattg acgtcaatgg gagtttgttt tggcaccaaa atcaacggga ctttccaaaa
     7021 tgtcgtaaca actccgcccc attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc
     7081 tatataagca gagctggttt agtgaaccgt cagatcagat ctttgtcgat cctaccatcc
     7141 actcgacaca cccgccagcg gccgc
//
