LOCUS       Exported                7765 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  Mammalian expression plasmid for SARS-CoV-2 3'-to-5' exonuclease.
ACCESSION   .
VERSION     .
KEYWORDS    pGBW-m4134334
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7765)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 7765)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 13, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7765
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     CDS             1583..1603
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             1604..1681
                     /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    1734..1855
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1745..1764)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1799..1818
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      1970..2016
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      2046..2182
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    2227..2298
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(2524..3112)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2604..2623)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        3612..4821
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     4658..4677
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             4825..5424
                     /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(4825..4844)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5321..5341
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    5451..5499
                     /note="synthetic polyadenylation signal"
     CDS             5765..6625
                     /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(5983..6002)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      6638..6774
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    6919..6990
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      6995..7041
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    7059..7107
                     /note="synthetic polyadenylation signal"
     enhancer        7188..7491
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        7493..7695
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     7646..7666
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      7764..8
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
ORIGIN
        1 caccatggaa aatgtaacag gactctttaa agattgtagt aaggtaatca ctgggttaca
       61 tcctacacag gcacctacac acctcagtgt tgacactaaa ttcaaaactg aaggtttatg
      121 tgttgacata cctggcatac ctaaggacat gacctataga agactcatct ctatgatggg
      181 ttttaaaatg aattatcaag ttaatggtta ccctaacatg tttatcaccc gcgaagaagc
      241 tataagacat gtacgtgcat ggattggctt cgatgtcgag gggtgtcatg ctactagaga
      301 agctgttggt accaatttac ctttacagct aggtttttct acaggtgtta acctagttgc
      361 tgtacctaca ggttatgttg atacacctaa taatacagat ttttccagag ttagtgctaa
      421 accaccgcct ggagatcaat ttaaacacct cataccactt atgtacaaag gacttccttg
      481 gaatgtagtg cgtataaaga ttgtacaaat gttaagtgac acacttaaaa atctctctga
      541 cagagtcgta tttgtcttat gggcacatgg ctttgagttg acatctatga agtattttgt
      601 gaaaatagga cctgagcgca cctgttgtct atgtgataga cgtgccacat gcttttccac
      661 tgcttcagac acttatgcct gttggcatca ttctattgga tttgattacg tctataatcc
      721 gtttatgatt gatgttcaac aatggggttt tacaggtaac ctacaaagca accatgatct
      781 gtattgtcaa gtccatggta atgcacatgt agctagttgt gatgcaatca tgactaggtg
      841 tctagctgtc cacgagtgct ttgttaagcg tgttgactgg actattgaat atcctataat
      901 tggtgatgaa ctgaagatta atgcggcttg tagaaaggtt caacacatgg ttgttaaagc
      961 tgcattatta gcagacaaat tcccagttct tcacgacatt ggtaacccta aagctattaa
     1021 gtgtgtacct caagctgatg tagaatggaa gttctatgat gcacagcctt gtagtgacaa
     1081 agcttataaa atagaagaat tattctattc ttatgccaca cattctgaca aattcacaga
     1141 tggtgtatgc ctattttgga attgcaatgt cgatagatat cctgctaatt ccattgtttg
     1201 tagatttgac actagagtgc tatctaacct taacttgcct ggttgtgatg gtggcagttt
     1261 gtatgtaaat aaacatgcat tccacacacc agcttttgat aaaagtgctt ttgttaattt
     1321 aaaacaatta ccatttttct attactctga cagtccatgt gagtctcatg gaaaacaagt
     1381 agtgtcagat atagattatg taccactaaa gtctgctacg tgtataacac gttgcaattt
     1441 aggtggtgct gtctgtagac atcatgctaa tgagtacaga ttgtatctcg atgcttataa
     1501 catgatgatc tcagctggct ttagcttgtg ggtttacaaa caatttgata cttataacct
     1561 ctggaacact tttacaagac ttgaaaacct ttactttcag tcgaagaggc gatggaaaaa
     1621 gaattttatc gcggtgagtg cagccaacag attcaaaaag atatcctcgt ctggtgccct
     1681 ctaagctagc ttgactgact gagatacagc gtaccttcag ctcacagaca tgataagata
     1741 cattgatgag tttggacaaa ccacaactag aatgcagtga aaaaaatgct ttatttgtga
     1801 aatttgtgat gctattgctt tatttgtaac cattataagc tgcaataaac aagttaacaa
     1861 caacaattgc attcatttta tgtttcaggt tcagggggag gtgtgggagg ttttttaaag
     1921 caagtaaaac ctctacaaat gtggtattgg cccatctcta tcggtatcgt agcataaccc
     1981 cttggggcct ctaaacgggt cttgaggggt tttttgtgcc cctcgggccg gattgctatc
     2041 taccggcatt ggcgcagaaa aaaatgcctg atgcgacgct gcgcgtctta tactcccaca
     2101 tatgccagat tcagcaacgg atacggcttc cccaacttgc ccacttccat acgtgtcctc
     2161 cttaccagaa atttatcctt aaggtcgtca gctatcctgc aggcgatctc tcgatttcga
     2221 tcaagacatt cctttaatgg tcttttctgg acaccactag gggtcagaag tagttcatca
     2281 aactttcttc cctccctaat ctcattggtt accttgggct atcgaaactt aattaaccag
     2341 tcaagtcagc tacttggcga gatcgacttg tctgggtttc gactacgctc agaattgcgt
     2401 cagtcaagtt cgatctggtc cttgctattg cacccgttct ccgattacga gtttcattta
     2461 aatcatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
     2521 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
     2581 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
     2641 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg
     2701 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg
     2761 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
     2821 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
     2881 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
     2941 gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt
     3001 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
     3061 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc
     3121 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt
     3181 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt
     3241 taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag
     3301 tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcat ttaaatttcc
     3361 gaactctcca aggccctcgt cggaaaatct tcaaaccttt cgtccgatcc atcttgcagg
     3421 ctacctctcg aacgaactat cgcaagtctc ttggccggcc ttgcgccttg gctattgctt
     3481 ggcagcgcct atcgccaggt attactccaa tcccgaatat ccgagatcgg gatcacccga
     3541 gagaagttca acctacatcc tcaatcccga tctatccgag atccgaggaa tatcgaaatc
     3601 ggggcgcgcc tggcctccgc gccgggtttt ggcgcctccc gcgggcgccc ccctcgtcac
     3661 ggcgagcgct gccacgtcag acgaagggcg caggagcgtc ctgatccttc cgcccggacg
     3721 ctcaggacag cggcccgctg ctcataagac tcggccttag aaccccagta tcagcagaag
     3781 gacattttag gacgggactt gggtgactct agggcactgg ttttctttcc agagagcgga
     3841 acaggcgagg aaaagtagtc ccttctcggc gattctgcgg agggatctcc gtggggcggt
     3901 gaacgccgat gattatataa ggacgcgccg ggtgtggcac agctagttcc gtcgcagccg
     3961 ggatttgggt cgcggttctt gtttgtggat cgctgtgatc gtcacttggt gagtagcggg
     4021 ctgctgggct ggccggggct ttcgtggccg ccgggccgct cggtgggacg gaagcgtgtg
     4081 gagagaccgc caagggctgt agtctgggtc cgcgagcaag gttgccctga actgggggtt
     4141 ggggggagcg cagcaaaatg gcggctgttc ccgagtcttg aatggaagac gcttgtgagg
     4201 cgggctgtga ggtcgttgaa acaaggtggg gggcatggtg ggcggcaaga acccaaggtc
     4261 ttgagccctt cgctaatgcg ggaaagctct tattcgggtg agatgggctg ggcaccatct
     4321 ggggaccctg acgtgaagtt tgtcactgac tggagaactc ggtttgtcgt ctgttgcggg
     4381 ggcggcagtt atggcggtgc cgttgggcag tgcacccgta cctttgggag cgcgcgccct
     4441 cgtcgtgtcg tgacgtcacc cgttctgttg gcttataatg cagggtgggg ccacctgccg
     4501 gtaggtgtgc ggtaggcttt tctccgtcgc aggacgcagg gttcgggcct agggtaggct
     4561 ctcctgaatc gacaggcgcc ggacctctgg tgaggggagg gataagtgag gcgtcagttt
     4621 ctttggtcgg ttttatgtac ctatcttctt aagtagctga agctccggtt ttgaactatg
     4681 cgctcggggt tggcgagtgt gttttgtgaa gttttttagg caccttttga aatgtaatca
     4741 tttgggtcaa tatgtaattt tcagtgttag acttgtaaat tgtccgctaa attctggccg
     4801 tttttggctt ttttgttaga caacatgacc gagtacaagc ccacggtgcg cctcgccacc
     4861 cgcgacgacg tccccagggc cgtacgcacc ctcgccgccg cgttcgccga ctaccccgcc
     4921 acgcgccaca ccgtcgatcc ggaccgccac atcgagcggg tcaccgagct gcaagaactc
     4981 ttcctcacgc gcgtcgggct cgacatcggc aaggtgtggg tcgcggacga cggcgccgcg
     5041 gtggcggtct ggaccacgcc ggagagcgtc gaagcggggg cggtgttcgc cgagatcggc
     5101 ccgcgcatgg ccgagttgag cggttcccgg ctggccgcgc agcaacagat ggagggcctc
     5161 ctggcgccgc accggcccaa ggagcccgcg tggttcctgg ccaccgtcgg cgtctcgccc
     5221 gaccaccagg gcaagggtct gggcagcgcc gtcgtgctcc ccggagtgga ggccgccgag
     5281 cgcgccgggg tgcccgcctt cctggagacc tccgcgcccc gcaacctccc cttctacgag
     5341 cggctcggct tcaccgtcac cgccgacgtc gaggtgcccg aaggaccgcg cacctggtgc
     5401 atgacccgca agcccggtgc ctaggctagt atgtaagcct agtcttagat aataaaatcg
     5461 ctatccatcg aagatggatg tgtgttggtt ttttgtgtgt gtaacgctag gcgcgcctgg
     5521 tgtaccgaga acgatcctct cagtgcgagt ctcgacgatc catatcgttg cttggcagtc
     5581 agccagtcgg aatccagctt gggacccagg aagtccaatc gtcagatatt gtactcaagc
     5641 ctggtcacgg cagcgtaccg atctgtttaa acctagatat tgatagtctg atcggtcaac
     5701 gtataatcga gtcctagctt ttgcaaacat ctatcaagag acaggatcag caggaggctt
     5761 tcgcatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct
     5821 tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg
     5881 tgcgcgagtg ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg
     5941 ccccgaagaa cgctttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt
     6001 atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga
     6061 cttggttgag tattcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga
     6121 attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac
     6181 gattggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg
     6241 ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac
     6301 gatgcctgta gcaatggcaa caaccttgcg taaactatta actggcgaac tacttactct
     6361 agcttcccgg caacagttga tagactggat ggaggcggat aaagttgcag gaccacttct
     6421 gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg
     6481 gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat
     6541 ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg
     6601 tgcctcactg attaagcatt ggtaaccgat tctaggtgca ttggcgcaga aaaaaatgcc
     6661 tgatgcgacg ctgcgcgtct tatactccca catatgccag attcagcaac ggatacggct
     6721 tccccaactt gcccacttcc atacgtgtcc tccttaccag aaatttatcc ttaagatcgt
     6781 ttaaactcga ctctggctct atcgaatctc cgtcgtttcg agcttacgcg aacagccgtg
     6841 gcgctcattt gctcgtcggg catcgaatct cgtcagctat cgtcagctta cctttttggc
     6901 agcgatcgcg gctcccgaca tcttggacca ttagctccac aggtatcttc ttccctctag
     6961 tggtcataac agcagcttca gctacctctc aattcaaaaa acccctcaag acccgtttag
     7021 aggccccaag gggttatgct atcaatcgtt gcgttacaca cacaaaaaac caacacacat
     7081 ccatcttcga tggatagcga ttttattatc taactgctga tcgagtgtag ccagatctag
     7141 taatcaatta cggggtcatt agttcatagc ccatatatgg agttccgcgt tacataactt
     7201 acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg
     7261 acgtatgttc ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat
     7321 ttacggtaaa ctgcccactt ggcagtacat caagtgtatc atatgccaag tacgccccct
     7381 attgacgtca atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttatgg
     7441 gactttccta cttggcagta catctacgta ttagtcatcg ctattaccat gctgatgcgg
     7501 ttttggcagt acatcaatgg gcgtggatag cggtttgact cacggggatt tccaagtctc
     7561 caccccattg acgtcaatgg gagtttgttt tggcaccaaa atcaacggga ctttccaaaa
     7621 tgtcgtaaca actccgcccc attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc
     7681 tatataagca gagctggttt agtgaaccgt cagatcagat ctttgtcgat cctaccatcc
     7741 actcgacaca cccgccagcg gccgc
//
