LOCUS       sequence-295252-        8216 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Mammalian expression plasmid for SARS-CoV-2 3'-to-5' exonuclease.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8216)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 8216)
  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-m4134166
FEATURES             Location/Qualifiers
     source          1..8216
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1589..1609
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             1610..1687
                     /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    1756..1877
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1767..1786)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1821..1840
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      1992..2038
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      2068..2204
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    2249..2320
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(2546..3134)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2626..2645)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        3634..4843
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     4680..4699
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             4847..5875
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     polyA_signal    5902..5950
                     /note="synthetic polyadenylation signal"
     CDS             join(6216..6284,6285..7076)
                     /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(6434..6453)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      7089..7225
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    7370..7441
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(7446..7492)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    7510..7558
                     /note="synthetic polyadenylation signal"
     enhancer        7639..7942
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        7944..8146
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     8097..8117
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      join(8215..8216,1..8)
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
ORIGIN
        1 caccatggcc gaaaatgtca ctggcttatt taaggattgc agtaaagtga tcacagggtt
       61 gcacccgacc caagcgccta cgcatctttc agtagacacc aagttcaaaa ctgagggact
      121 ctgtgtggac ataccaggta ttcccaagga tatgacatac agacgcctga tctcgatgat
      181 gggcttcaag atgaactatc aggttaacgg ttaccccaat atgtttatta cccgagagga
      241 agctatccgg cacgtgcgtg cctggatagg gtttgacgtc gagggctgtc atgcaactag
      301 ggaagctgta ggaacaaatc tgccactgca actcggcttc tctaccggag tcaacctagt
      361 tgccgtgcct accgggtatg tggatacacc aaacaatact gattttagca gagtctccgc
      421 aaaaccccct cccggggacc agttcaaaca cttgatcccc ctgatgtaca agggtttacc
      481 atggaacgtg gtgcggatta aaatagttca gatgctcagc gacacactga agaatttaag
      541 tgatagagtt gtgtttgtac tttgggccca tggcttcgag ctgacctcta tgaagtattt
      601 cgttaaaatt ggacccgaac gaacgtgctg cctgtgtgac cggagggcga cgtgtttctc
      661 aactgcttcc gatacctatg cctgctggca ccatagcatc ggctttgatt acgtgtacaa
      721 cccttttatg attgacgtgc agcagtgggg attcactggg aacctacagt cgaatcacga
      781 tctttactgt caggtccatg ggaacgcaca cgttgctagc tgcgacgcaa tcatgacacg
      841 ttgccttgct gtccacgagt gttttgtgaa acgagtggac tggactattg agtatcctat
      901 cattggcgac gaactgaaga taaatgcggc ttgtaggaag gtgcagcata tggtcgtcaa
      961 agccgccctc ttagctgata agttcccggt cctgcacgat attggcaatc caaaggccat
     1021 caaatgcgta ccccaggcag atgttgagtg gaagttttac gacgcgcaac catgctccga
     1081 taaggcctat aaaatcgaag agctcttcta cagttatgcc acccattctg acaagtttac
     1141 cgacggtgtg tgtttgtttt ggaattgcaa cgtggatcgc taccctgcaa actctatcgt
     1201 ttgtcgcttc gacacacgcg tattgtccaa tctgaatcta cccggatgcg acggaggttc
     1261 actgtatgtg aacaagcacg ccttccacac ccccgcattc gataaatccg cctttgtcaa
     1321 ccttaaacaa ttacctttct tttactacag cgactctcct tgcgaatccc atggaaagca
     1381 agtagtgagt gatatcgatt acgtcccact caaaagtgca acatgtatta cgagatgcaa
     1441 tctcggtggt gcagtctgta gacaccatgc caatgaatat cggctgtatt tggacgccta
     1501 taacatgatg atttcagctg gcttctctct ttgggtttac aagcagtttg acacgtataa
     1561 tctgtggaac acctttacaa ggctgcaaga aaacctttac tttcagtcga agaggcgatg
     1621 gaaaaagaat tttatcgcgg tgagtgcagc caacagattc aaaaagatat cctcgtctgg
     1681 tgccctctaa ggatccgcag gcctctgcta gcttgactga ctgagataca gcgtaccttc
     1741 agctcacaga catgataaga tacattgatg agtttggaca aaccacaact agaatgcagt
     1801 gaaaaaaatg ctttatttgt gaaatttgtg atgctattgc tttatttgta accattataa
     1861 gctgcaataa acaagttaac aacaacaatt gcattcattt tatgtttcag gttcaggggg
     1921 aggtgtggga ggttttttaa agcaagtaaa acctctacaa atgtggtatt ggcccatctc
     1981 tatcggtatc gtagcataac cccttggggc ctctaaacgg gtcttgaggg gttttttgtg
     2041 cccctcgggc cggattgcta tctaccggca ttggcgcaga aaaaaatgcc tgatgcgacg
     2101 ctgcgcgtct tatactccca catatgccag attcagcaac ggatacggct tccccaactt
     2161 gcccacttcc atacgtgtcc tccttaccag aaatttatcc ttaaggtcgt cagctatcct
     2221 gcaggcgatc tctcgatttc gatcaagaca ttcctttaat ggtcttttct ggacaccact
     2281 aggggtcaga agtagttcat caaactttct tccctcccta atctcattgg ttaccttggg
     2341 ctatcgaaac ttaattaacc agtcaagtca gctacttggc gagatcgact tgtctgggtt
     2401 tcgactacgc tcagaattgc gtcagtcaag ttcgatctgg tccttgctat tgcacccgtt
     2461 ctccgattac gagtttcatt taaatcatgt gagcaaaagg ccagcaaaag gccaggaacc
     2521 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
     2581 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
     2641 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     2701 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
     2761 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     2821 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     2881 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     2941 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta
     3001 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     3061 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
     3121 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
     3181 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc
     3241 ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg
     3301 acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat
     3361 ccatagttgc atttaaattt ccgaactctc caaggccctc gtcggaaaat cttcaaacct
     3421 ttcgtccgat ccatcttgca ggctacctct cgaacgaact atcgcaagtc tcttggccgg
     3481 ccttgcgcct tggctattgc ttggcagcgc ctatcgccag gtattactcc aatcccgaat
     3541 atccgagatc gggatcaccc gagagaagtt caacctacat cctcaatccc gatctatccg
     3601 agatccgagg aatatcgaaa tcggggcgcg cctggcctcc gcgccgggtt ttggcgcctc
     3661 ccgcgggcgc ccccctcgtc acggcgagcg ctgccacgtc agacgaaggg cgcaggagcg
     3721 tcctgatcct tccgcccgga cgctcaggac agcggcccgc tgctcataag actcggcctt
     3781 agaaccccag tatcagcaga aggacatttt aggacgggac ttgggtgact ctagggcact
     3841 ggttttcttt ccagagagcg gaacaggcga ggaaaagtag tcccttctcg gcgattctgc
     3901 ggagggatct ccgtggggcg gtgaacgccg atgattatat aaggacgcgc cgggtgtggc
     3961 acagctagtt ccgtcgcagc cgggatttgg gtcgcggttc ttgtttgtgg atcgctgtga
     4021 tcgtcacttg gtgagtagcg ggctgctggg ctggccgggg ctttcgtggc cgccgggccg
     4081 ctcggtggga cggaagcgtg tggagagacc gccaagggct gtagtctggg tccgcgagca
     4141 aggttgccct gaactggggg ttggggggag cgcagcaaaa tggcggctgt tcccgagtct
     4201 tgaatggaag acgcttgtga ggcgggctgt gaggtcgttg aaacaaggtg gggggcatgg
     4261 tgggcggcaa gaacccaagg tcttgagccc ttcgctaatg cgggaaagct cttattcggg
     4321 tgagatgggc tgggcaccat ctggggaccc tgacgtgaag tttgtcactg actggagaac
     4381 tcggtttgtc gtctgttgcg ggggcggcag ttatggcggt gccgttgggc agtgcacccg
     4441 tacctttggg agcgcgcgcc ctcgtcgtgt cgtgacgtca cccgttctgt tggcttataa
     4501 tgcagggtgg ggccacctgc cggtaggtgt gcggtaggct tttctccgtc gcaggacgca
     4561 gggttcgggc ctagggtagg ctctcctgaa tcgacaggcg ccggacctct ggtgagggga
     4621 gggataagtg aggcgtcagt ttctttggtc ggttttatgt acctatcttc ttaagtagct
     4681 gaagctccgg ttttgaacta tgcgctcggg gttggcgagt gtgttttgtg aagtttttta
     4741 ggcacctttt gaaatgtaat catttgggtc aatatgtaat tttcagtgtt agacttgtaa
     4801 attgtccgct aaattctggc cgtttttggc ttttttgtta gacaacatgg gtaaaaagcc
     4861 tgaactcacc gcgacgtctg tcgagaagtt tctgatcgaa aagttcgaca gcgtctccga
     4921 cctgatgcag ctctcggagg gcgaagaatc tcgtgctttc agcttcgatg taggagggcg
     4981 tggatatgtc ctgcgggtaa atagctgcgc cgatggtttc tacaaagatc gttatgttta
     5041 tcggcacttt gcatccgccg cgctcccgat tccggaagtg cttgacattg gggagttcag
     5101 cgagagcctg acctattgca tctcccgccg tgcacagggt gtcacgttgc aagacctgcc
     5161 tgaaaccgaa ctgcccgctg ttctgcagcc ggtcgcggag gcaatggatg ccatcgctgc
     5221 cgccgatctt agccagacga gcgggttcgg cccattcgga ccgcaaggaa tcggtcaata
     5281 cactacatgg cgtgatttca tatgcgcgat tgctgatccc catgtgtatc actggcaaac
     5341 tgtgatggac gacaccgtca gtgcgtccgt cgcgcaggct ctcgatgagc tgatgctttg
     5401 ggccgaggac tgccccgaag tccggcacct cgtgcacgcg gatttcggct ccaacaatgt
     5461 cctgacggac aatggccgca taacagcggt cattgactgg agcgaggcga tgttcgggga
     5521 ttcccaatac gaggtcgcca acatcttctt ctggaggccg tggttggctt gtatggagca
     5581 gcagacgcgc tacttcgagc ggaggcatcc ggagcttgca ggatcgcccc ggctccgggc
     5641 gtatatgctc cgcattggtc ttgaccaact ctatcagagc ttggttgacg gcaatttcga
     5701 tgatgcagct tgggcgcagg gtcgatgcga cgcaatcgtc cgatccggag ccgggactgt
     5761 cgggcgtaca caaatcgccc gcagaagcgc cgccgtctgg accgatggct gtgtagaagt
     5821 actcgccgat agtggaaacc gacgccccag cactcgtccg agggcaaagg aataagctag
     5881 tatgtaagcc tagtcttaga taataaaatc gctatccatc gaagatggat gtgtgttggt
     5941 tttttgtgtg tgtaacgcta ggcgcgcctg gtgtaccgag aacgatcctc tcagtgcgag
     6001 tctcgacgat ccatatcgtt gcttggcagt cagccagtcg gaatccagct tgggacccag
     6061 gaagtccaat cgtcagatat tgtactcaag cctggtcacg gcagcgtacc gatctgttta
     6121 aacctagata ttgatagtct gatcggtcaa cgtataatcg agtcctagct tttgcaaaca
     6181 tctatcaaga gacaggatca gcaggaggct ttcgcatgag tattcaacat ttccgtgtcg
     6241 cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg
     6301 tgaaagtaaa agatgctgaa gatcagttgg gtgcgcgagt gggttacatc gaactggatc
     6361 tcaacagcgg taagatcctt gagagttttc gccccgaaga acgctttcca atgatgagca
     6421 cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac
     6481 tcggtcgccg catacactat tctcagaatg acttggttga gtattcacca gtcacagaaa
     6541 agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg
     6601 ataacactgc ggccaactta cttctgacaa cgattggagg accgaaggag ctaaccgctt
     6661 ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg
     6721 aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaaccttgc
     6781 gtaaactatt aactggcgaa ctacttactc tagcttcccg gcaacagttg atagactgga
     6841 tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta
     6901 ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc
     6961 cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg
     7021 atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat tggtaaccga
     7081 ttctaggtgc attggcgcag aaaaaaatgc ctgatgcgac gctgcgcgtc ttatactccc
     7141 acatatgcca gattcagcaa cggatacggc ttccccaact tgcccacttc catacgtgtc
     7201 ctccttacca gaaatttatc cttaagatcg tttaaactcg actctggctc tatcgaatct
     7261 ccgtcgtttc gagcttacgc gaacagccgt ggcgctcatt tgctcgtcgg gcatcgaatc
     7321 tcgtcagcta tcgtcagctt acctttttgg cagcgatcgc ggctcccgac atcttggacc
     7381 attagctcca caggtatctt cttccctcta gtggtcataa cagcagcttc agctacctct
     7441 caattcaaaa aacccctcaa gacccgttta gaggccccaa ggggttatgc tatcaatcgt
     7501 tgcgttacac acacaaaaaa ccaacacaca tccatcttcg atggatagcg attttattat
     7561 ctaactgctg atcgagtgta gccagatcta gtaatcaatt acggggtcat tagttcatag
     7621 cccatatatg gagttccgcg ttacataact tacggtaaat ggcccgcctg gctgaccgcc
     7681 caacgacccc cgcccattga cgtcaataat gacgtatgtt cccatagtaa cgccaatagg
     7741 gactttccat tgacgtcaat gggtggagta tttacggtaa actgcccact tggcagtaca
     7801 tcaagtgtat catatgccaa gtacgccccc tattgacgtc aatgacggta aatggcccgc
     7861 ctggcattat gcccagtaca tgaccttatg ggactttcct acttggcagt acatctacgt
     7921 attagtcatc gctattacca tgctgatgcg gttttggcag tacatcaatg ggcgtggata
     7981 gcggtttgac tcacggggat ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt
     8041 ttggcaccaa aatcaacggg actttccaaa atgtcgtaac aactccgccc cattgacgca
     8101 aatgggcggt aggcgtgtac ggtgggaggt ctatataagc agagctggtt tagtgaaccg
     8161 tcagatcaga tctttgtcga tcctaccatc cactcgacac acccgccagc ggccgc
//
