LOCUS       Exported                8454 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  Mammalian expression plasmid for SARS-CoV-2 nsp4.
ACCESSION   .
VERSION     .
KEYWORDS    pGBW-m4134486
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8454)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 8454)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 13, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..8454
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        1..304
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        306..508
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     459..479
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             2086..2106
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             2455..2472
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     polyA_signal    2565..2686
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2576..2595)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2630..2649
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      2801..2847
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      2877..3013
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    3058..3129
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(3355..3943)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3435..3454)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        4443..5652
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     5489..5508
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             5656..6684
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     polyA_signal    6711..6759
                     /note="synthetic polyadenylation signal"
     CDS             7025..7885
                     /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(7243..7262)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      7898..8034
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    8186..8257
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      8262..8308
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    8326..8374
                     /note="synthetic polyadenylation signal"
ORIGIN
        1 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
       61 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      121 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      181 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      241 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      301 catgctgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      361 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      421 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      481 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatca gatctttgtc
      541 gatcctacca tccactcgac acacccgcca gcggccgcca ccatgaagat tgtaaataac
      601 tggttgaaac aactaataaa ggttacttta gtgttcctgt ttgtcgccgc aatcttctat
      661 cttatcaccc cggtgcacgt gatgagtaaa catacagact tttcttcaga aattatcggg
      721 tacaaggcta tcgatggcgg tgtaacgcgc gacattgcct ccacagatac ctgcttcgcg
      781 aacaaacacg ctgactttga tacttggttc agccagcgag gaggctcgta caccaatgac
      841 aaagcctgtc cactgatcgc agccgtgata actagagagg ttggattcgt cgtccctggg
      901 ctccccggga caattctccg tacaaccaac ggcgactttc tgcattttct gccccgggtg
      961 ttctccgcag tcggtaatat ctgttatact ccatctaagc tcattgagta cacggatttt
     1021 gctaccagcg cgtgcgtgtt ggccgcagaa tgcaccatat ttaaggacgc ttctggaaaa
     1081 cctgtgccct attgttacga taccaatgtg cttgagggct cagttgccta tgaaagtctg
     1141 aggccagaca ctagatacgt tttaatggat ggaagcatca tccagttccc aaacacgtat
     1201 ctggaggggt ccgtgcgggt cgtgaccaca ttcgattcag aatactgccg ccacggtaca
     1261 tgcgagaggt ctgaggctgg cgtatgtgtg tcaactagcg gtcgatgggt tctaaataac
     1321 gattattatc gttctctgcc cggcgttttt tgtggagtgg acgccgttaa ccttttgacc
     1381 aacatgttta cacctctcat tcagcccatt ggtgcactgg atatctccgc gagtattgtg
     1441 gctgggggaa tagtcgccat tgtcgtaacc tgcttggcat actatttcat gcgtttcagg
     1501 cgagcttttg gcgaatactc acatgtcgtg gccttcaata ctcttctgtt tctgatgagc
     1561 ttcacggtac tctgtctgac cccagtatac agtttcctac ctggggtgta ttctgtgatc
     1621 tacttatacc tcacatttta cctgaccaat gacgtgtcct ttctggccca catccaatgg
     1681 atggttatgt tcacgccgct cgtgcctttc tggattacaa tcgcttatat aatctgtatt
     1741 agcacgaagc atttctattg gttcttttcc aactacttaa agcggcgcgt cgtatttaat
     1801 ggggtatcgt ttagcacttt tgaggaagcc gcgctctgca ctttcttgct taataaggag
     1861 atgtatttga aattaagatc tgacgttctg cttcctttaa cacagtacaa cagatacctg
     1921 gcgttgtata acaaatacaa atatttctcg ggagccatgg atacaacttc ctatagagag
     1981 gctgcttgtt gccaccttgc aaaggcactc aacgacttca gtaacagcgg cagtgatgtg
     2041 ctttatcagc cccctcagac ttcaataaca tccgccgtcc tccaagaaaa cctttacttt
     2101 cagtcgatga gtgacaaaat cattcatcta accgatgact cctttgatac agacgtgttg
     2161 aaggccgatg gtgcgatact ggtagatttc tgggctgaat ggtgtggccc gtgcaagatg
     2221 atcgcaccca ttctggacga gatcgccgac gagtatcaag gaaaattaac tgtcgctaag
     2281 cttaatatag atcagaaccc agggacggca cctaaatacg gcatcagggg aatacctacc
     2341 ctcctgctct ttaagaatgg ggaagttgcc gcgacaaaag tgggagccct gtctaagggc
     2401 cagctaaaag agttcttgga cgctaacctg gcaggttcag gctcggggca catgcaccat
     2461 caccatcacc atagcagcgg gctcgtgcca cgcggctaag gatccgcagg cctctgctag
     2521 cttgactgac tgagatacag cgtaccttca gctcacagac atgataagat acattgatga
     2581 gtttggacaa accacaacta gaatgcagtg aaaaaaatgc tttatttgtg aaatttgtga
     2641 tgctattgct ttatttgtaa ccattataag ctgcaataaa caagttaaca acaacaattg
     2701 cattcatttt atgtttcagg ttcaggggga ggtgtgggag gttttttaaa gcaagtaaaa
     2761 cctctacaaa tgtggtattg gcccatctct atcggtatcg tagcataacc ccttggggcc
     2821 tctaaacggg tcttgagggg ttttttgtgc ccctcgggcc ggattgctat ctaccggcat
     2881 tggcgcagaa aaaaatgcct gatgcgacgc tgcgcgtctt atactcccac atatgccaga
     2941 ttcagcaacg gatacggctt ccccaacttg cccacttcca tacgtgtcct ccttaccaga
     3001 aatttatcct taaggtcgtc agctatcctg caggcgatct ctcgatttcg atcaagacat
     3061 tcctttaatg gtcttttctg gacaccacta ggggtcagaa gtagttcatc aaactttctt
     3121 ccctccctaa tctcattggt taccttgggc tatcgaaact taattaacca gtcaagtcag
     3181 ctacttggcg agatcgactt gtctgggttt cgactacgct cagaattgcg tcagtcaagt
     3241 tcgatctggt ccttgctatt gcacccgttc tccgattacg agtttcattt aaatcatgtg
     3301 agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca
     3361 taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa
     3421 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc
     3481 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc
     3541 gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct
     3601 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg
     3661 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag
     3721 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta
     3781 cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg
     3841 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt
     3901 tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt
     3961 ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag
     4021 attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat
     4081 ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc
     4141 tatctcagcg atctgtctat ttcgttcatc catagttgca tttaaatttc cgaactctcc
     4201 aaggccctcg tcggaaaatc ttcaaacctt tcgtccgatc catcttgcag gctacctctc
     4261 gaacgaacta tcgcaagtct cttggccggc cttgcgcctt ggctattgct tggcagcgcc
     4321 tatcgccagg tattactcca atcccgaata tccgagatcg ggatcacccg agagaagttc
     4381 aacctacatc ctcaatcccg atctatccga gatccgagga atatcgaaat cggggcgcgc
     4441 ctggcctccg cgccgggttt tggcgcctcc cgcgggcgcc cccctcgtca cggcgagcgc
     4501 tgccacgtca gacgaagggc gcaggagcgt cctgatcctt ccgcccggac gctcaggaca
     4561 gcggcccgct gctcataaga ctcggcctta gaaccccagt atcagcagaa ggacatttta
     4621 ggacgggact tgggtgactc tagggcactg gttttctttc cagagagcgg aacaggcgag
     4681 gaaaagtagt cccttctcgg cgattctgcg gagggatctc cgtggggcgg tgaacgccga
     4741 tgattatata aggacgcgcc gggtgtggca cagctagttc cgtcgcagcc gggatttggg
     4801 tcgcggttct tgtttgtgga tcgctgtgat cgtcacttgg tgagtagcgg gctgctgggc
     4861 tggccggggc tttcgtggcc gccgggccgc tcggtgggac ggaagcgtgt ggagagaccg
     4921 ccaagggctg tagtctgggt ccgcgagcaa ggttgccctg aactgggggt tggggggagc
     4981 gcagcaaaat ggcggctgtt cccgagtctt gaatggaaga cgcttgtgag gcgggctgtg
     5041 aggtcgttga aacaaggtgg ggggcatggt gggcggcaag aacccaaggt cttgagccct
     5101 tcgctaatgc gggaaagctc ttattcgggt gagatgggct gggcaccatc tggggaccct
     5161 gacgtgaagt ttgtcactga ctggagaact cggtttgtcg tctgttgcgg gggcggcagt
     5221 tatggcggtg ccgttgggca gtgcacccgt acctttggga gcgcgcgccc tcgtcgtgtc
     5281 gtgacgtcac ccgttctgtt ggcttataat gcagggtggg gccacctgcc ggtaggtgtg
     5341 cggtaggctt ttctccgtcg caggacgcag ggttcgggcc tagggtaggc tctcctgaat
     5401 cgacaggcgc cggacctctg gtgaggggag ggataagtga ggcgtcagtt tctttggtcg
     5461 gttttatgta cctatcttct taagtagctg aagctccggt tttgaactat gcgctcgggg
     5521 ttggcgagtg tgttttgtga agttttttag gcaccttttg aaatgtaatc atttgggtca
     5581 atatgtaatt ttcagtgtta gacttgtaaa ttgtccgcta aattctggcc gtttttggct
     5641 tttttgttag acaacatggg taaaaagcct gaactcaccg cgacgtctgt cgagaagttt
     5701 ctgatcgaaa agttcgacag cgtctccgac ctgatgcagc tctcggaggg cgaagaatct
     5761 cgtgctttca gcttcgatgt aggagggcgt ggatatgtcc tgcgggtaaa tagctgcgcc
     5821 gatggtttct acaaagatcg ttatgtttat cggcactttg catccgccgc gctcccgatt
     5881 ccggaagtgc ttgacattgg ggagttcagc gagagcctga cctattgcat ctcccgccgt
     5941 gcacagggtg tcacgttgca agacctgcct gaaaccgaac tgcccgctgt tctgcagccg
     6001 gtcgcggagg caatggatgc catcgctgcc gccgatctta gccagacgag cgggttcggc
     6061 ccattcggac cgcaaggaat cggtcaatac actacatggc gtgatttcat atgcgcgatt
     6121 gctgatcccc atgtgtatca ctggcaaact gtgatggacg acaccgtcag tgcgtccgtc
     6181 gcgcaggctc tcgatgagct gatgctttgg gccgaggact gccccgaagt ccggcacctc
     6241 gtgcacgcgg atttcggctc caacaatgtc ctgacggaca atggccgcat aacagcggtc
     6301 attgactgga gcgaggcgat gttcggggat tcccaatacg aggtcgccaa catcttcttc
     6361 tggaggccgt ggttggcttg tatggagcag cagacgcgct acttcgagcg gaggcatccg
     6421 gagcttgcag gatcgccccg gctccgggcg tatatgctcc gcattggtct tgaccaactc
     6481 tatcagagct tggttgacgg caatttcgat gatgcagctt gggcgcaggg tcgatgcgac
     6541 gcaatcgtcc gatccggagc cgggactgtc gggcgtacac aaatcgcccg cagaagcgcc
     6601 gccgtctgga ccgatggctg tgtagaagta ctcgccgata gtggaaaccg acgccccagc
     6661 actcgtccga gggcaaagga ataagctagt atgtaagcct agtcttagat aataaaatcg
     6721 ctatccatcg aagatggatg tgtgttggtt ttttgtgtgt gtaacgctag gcgcgcctgg
     6781 tgtaccgaga acgatcctct cagtgcgagt ctcgacgatc catatcgttg cttggcagtc
     6841 agccagtcgg aatccagctt gggacccagg aagtccaatc gtcagatatt gtactcaagc
     6901 ctggtcacgg cagcgtaccg atctgtttaa acctagatat tgatagtctg atcggtcaac
     6961 gtataatcga gtcctagctt ttgcaaacat ctatcaagag acaggatcag caggaggctt
     7021 tcgcatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct
     7081 tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg
     7141 tgcgcgagtg ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg
     7201 ccccgaagaa cgctttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt
     7261 atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga
     7321 cttggttgag tattcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga
     7381 attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac
     7441 gattggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg
     7501 ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac
     7561 gatgcctgta gcaatggcaa caaccttgcg taaactatta actggcgaac tacttactct
     7621 agcttcccgg caacagttga tagactggat ggaggcggat aaagttgcag gaccacttct
     7681 gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg
     7741 gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat
     7801 ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg
     7861 tgcctcactg attaagcatt ggtaaccgat tctaggtgca ttggcgcaga aaaaaatgcc
     7921 tgatgcgacg ctgcgcgtct tatactccca catatgccag attcagcaac ggatacggct
     7981 tccccaactt gcccacttcc atacgtgtcc tccttaccag aaatttatcc ttaagatccc
     8041 gaatcgttta aactcgactc tggctctatc gaatctccgt cgtttcgagc ttacgcgaac
     8101 agccgtggcg ctcatttgct cgtcgggcat cgaatctcgt cagctatcgt cagcttacct
     8161 ttttggcagc gatcgcggct cccgacatct tggaccatta gctccacagg tatcttcttc
     8221 cctctagtgg tcataacagc agcttcagct acctctcaat tcaaaaaacc cctcaagacc
     8281 cgtttagagg ccccaagggg ttatgctatc aatcgttgcg ttacacacac aaaaaaccaa
     8341 cacacatcca tcttcgatgg atagcgattt tattatctaa ctgctgatcg agtgtagcca
     8401 gatctagtaa tcaattacgg ggtcattagt tcatagccca tatatggagt tccg
//
