LOCUS       Exported                8299 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  Mammalian expression plasmid for SARS-CoV-2 helicase.
ACCESSION   .
VERSION     .
KEYWORDS    pGBW-m4134102
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8299)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 8299)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 13, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..8299
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     CDS             1805..1825
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             2174..2191
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     polyA_signal    2268..2389
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2279..2298)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2333..2352
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      2504..2550
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      2580..2716
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    2761..2832
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(3058..3646)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3138..3157)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        4146..5355
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     5192..5211
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             5359..5958
                     /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(5359..5378)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5855..5875
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    5985..6033
                     /note="synthetic polyadenylation signal"
     CDS             6299..7159
                     /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(6517..6536)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      7172..7308
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    7453..7524
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      7529..7575
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    7593..7641
                     /note="synthetic polyadenylation signal"
     enhancer        7722..8025
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        8027..8229
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     8180..8200
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      8298..8
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
ORIGIN
        1 caccatggtt ggggcttgtg ttctttgcaa ttcacagact tcattaagat gtggtgcttg
       61 catacgtaga ccattcttat gttgtaaatg ctgttacgac catgtcatat caacatcaca
      121 taaattagtc ttgtctgtta atccgtatgt ttgcaatgct ccaggttgtg atgtcacaga
      181 tgtgactcaa ctttacttag gaggtatgag ctattattgt aaatcacata aaccacccat
      241 tagttttcca ttgtgtgcta atggacaagt ttttggttta tataaaaata catgtgttgg
      301 tagcgataat gttactgact ttaatgcaat tgcaacatgt gactggacaa atgctggtga
      361 ttacatttta gctaacacct gtactgaaag actcaagctt tttgcagcag aaacgctcaa
      421 agctactgag gagacattta aactgtctta tggtattgct actgtacgtg aagtgctgtc
      481 tgacagagaa ttacatcttt catgggaagt tggtaaacct agaccaccac ttaaccgaaa
      541 ttatgtcttt actggttatc gtgtaactaa aaacagtaaa gtacaaatag gagagtacac
      601 ctttgaaaaa ggtgactatg gtgatgctgt tgtttaccga ggtacaacaa cttacaaatt
      661 aaatgttggt gattattttg tgctgacatc acatacagta atgccattaa gtgcacctac
      721 actagtgcca caagagcact atgttagaat tactggctta tacccaacac tcaatatctc
      781 agatgagttt tctagcaatg ttgcaaatta tcaaaaggtt ggtatgcaaa agtattctac
      841 actccaggga ccacctggta ctggtaagag tcattttgct attggcctag ctctctacta
      901 cccttctgct cgcatagtgt atacagcttg ctctcatgcc gctgttgatg cactatgtga
      961 gaaggcatta aaatatttgc ctatagataa atgtagtaga attatacctg cacgtgctcg
     1021 tgtagagtgt tttgataaat tcaaagtgaa ttcaacatta gaacagtatg tcttttgtac
     1081 tgtaaatgca ttgcctgaga cgacagcaga tatagttgtc tttgatgaaa tttcaatggc
     1141 cacaaattat gatttgagtg ttgtcaatgc cagattacgt gctaagcact atgtgtacat
     1201 tggcgaccct gctcaattac ctgcaccacg cacattgcta actaagggca cactagaacc
     1261 agaatatttc aattcagtgt gtagacttat gaaaactata ggtccagaca tgttcctcgg
     1321 aacttgtcgg cgttgtcctg ctgaaattgt tgacactgtg agtgctttgg tttatgataa
     1381 taagcttaaa gcacataaag acaaatcagc tcaatgcttt aaaatgtttt ataagggtgt
     1441 tatcacgcat gatgtttcat ctgcaattaa caggccacaa ataggcgtgg taagagaatt
     1501 ccttacacgt aaccctgctt ggagaaaagc tgtctttatt tcaccttata attcacagaa
     1561 tgctgtagcc tcaaagattt tgggactacc aactcaaact gttgattcat cacagggctc
     1621 agaatatgac tatgtcatat tcactcaaac cactgaaaca gctcactctt gtaatgtaaa
     1681 cagatttaat gttgctatta ccagagcaaa agtaggcata ctttgcataa tgtctgatag
     1741 agacctttat gacaagttgc aatttacaag tcttgaaatt ccacgtagga atgtggcaac
     1801 tttagaaaac ctttactttc agtcgatgag tgacaaaatc attcatctaa ccgatgactc
     1861 ctttgataca gacgtgttga aggccgatgg tgcgatactg gtagatttct gggctgaatg
     1921 gtgtggcccg tgcaagatga tcgcacccat tctggacgag atcgccgacg agtatcaagg
     1981 aaaattaact gtcgctaagc ttaatataga tcagaaccca gggacggcac ctaaatacgg
     2041 catcagggga atacctaccc tcctgctctt taagaatggg gaagttgccg cgacaaaagt
     2101 gggagccctg tctaagggcc agctaaaaga gttcttggac gctaacctgg caggttcagg
     2161 ctcggggcac atgcaccatc accatcacca tagcagcggg ctcgtgccac gcggctaagc
     2221 tagcttgact gactgagata cagcgtacct tcagctcaca gacatgataa gatacattga
     2281 tgagtttgga caaaccacaa ctagaatgca gtgaaaaaaa tgctttattt gtgaaatttg
     2341 tgatgctatt gctttatttg taaccattat aagctgcaat aaacaagtta acaacaacaa
     2401 ttgcattcat tttatgtttc aggttcaggg ggaggtgtgg gaggtttttt aaagcaagta
     2461 aaacctctac aaatgtggta ttggcccatc tctatcggta tcgtagcata accccttggg
     2521 gcctctaaac gggtcttgag gggttttttg tgcccctcgg gccggattgc tatctaccgg
     2581 cattggcgca gaaaaaaatg cctgatgcga cgctgcgcgt cttatactcc cacatatgcc
     2641 agattcagca acggatacgg cttccccaac ttgcccactt ccatacgtgt cctccttacc
     2701 agaaatttat ccttaaggtc gtcagctatc ctgcaggcga tctctcgatt tcgatcaaga
     2761 cattccttta atggtctttt ctggacacca ctaggggtca gaagtagttc atcaaacttt
     2821 cttccctccc taatctcatt ggttaccttg ggctatcgaa acttaattaa ccagtcaagt
     2881 cagctacttg gcgagatcga cttgtctggg tttcgactac gctcagaatt gcgtcagtca
     2941 agttcgatct ggtccttgct attgcacccg ttctccgatt acgagtttca tttaaatcat
     3001 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
     3061 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
     3121 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
     3181 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     3241 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     3301 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     3361 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     3421 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     3481 ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     3541 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     3601 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     3661 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     3721 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc
     3781 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc
     3841 acctatctca gcgatctgtc tatttcgttc atccatagtt gcatttaaat ttccgaactc
     3901 tccaaggccc tcgtcggaaa atcttcaaac ctttcgtccg atccatcttg caggctacct
     3961 ctcgaacgaa ctatcgcaag tctcttggcc ggccttgcgc cttggctatt gcttggcagc
     4021 gcctatcgcc aggtattact ccaatcccga atatccgaga tcgggatcac ccgagagaag
     4081 ttcaacctac atcctcaatc ccgatctatc cgagatccga ggaatatcga aatcggggcg
     4141 cgcctggcct ccgcgccggg ttttggcgcc tcccgcgggc gcccccctcg tcacggcgag
     4201 cgctgccacg tcagacgaag ggcgcaggag cgtcctgatc cttccgcccg gacgctcagg
     4261 acagcggccc gctgctcata agactcggcc ttagaacccc agtatcagca gaaggacatt
     4321 ttaggacggg acttgggtga ctctagggca ctggttttct ttccagagag cggaacaggc
     4381 gaggaaaagt agtcccttct cggcgattct gcggagggat ctccgtgggg cggtgaacgc
     4441 cgatgattat ataaggacgc gccgggtgtg gcacagctag ttccgtcgca gccgggattt
     4501 gggtcgcggt tcttgtttgt ggatcgctgt gatcgtcact tggtgagtag cgggctgctg
     4561 ggctggccgg ggctttcgtg gccgccgggc cgctcggtgg gacggaagcg tgtggagaga
     4621 ccgccaaggg ctgtagtctg ggtccgcgag caaggttgcc ctgaactggg ggttgggggg
     4681 agcgcagcaa aatggcggct gttcccgagt cttgaatgga agacgcttgt gaggcgggct
     4741 gtgaggtcgt tgaaacaagg tggggggcat ggtgggcggc aagaacccaa ggtcttgagc
     4801 ccttcgctaa tgcgggaaag ctcttattcg ggtgagatgg gctgggcacc atctggggac
     4861 cctgacgtga agtttgtcac tgactggaga actcggtttg tcgtctgttg cgggggcggc
     4921 agttatggcg gtgccgttgg gcagtgcacc cgtacctttg ggagcgcgcg ccctcgtcgt
     4981 gtcgtgacgt cacccgttct gttggcttat aatgcagggt ggggccacct gccggtaggt
     5041 gtgcggtagg cttttctccg tcgcaggacg cagggttcgg gcctagggta ggctctcctg
     5101 aatcgacagg cgccggacct ctggtgaggg gagggataag tgaggcgtca gtttctttgg
     5161 tcggttttat gtacctatct tcttaagtag ctgaagctcc ggttttgaac tatgcgctcg
     5221 gggttggcga gtgtgttttg tgaagttttt taggcacctt ttgaaatgta atcatttggg
     5281 tcaatatgta attttcagtg ttagacttgt aaattgtccg ctaaattctg gccgtttttg
     5341 gcttttttgt tagacaacat gaccgagtac aagcccacgg tgcgcctcgc cacccgcgac
     5401 gacgtcccca gggccgtacg caccctcgcc gccgcgttcg ccgactaccc cgccacgcgc
     5461 cacaccgtcg atccggaccg ccacatcgag cgggtcaccg agctgcaaga actcttcctc
     5521 acgcgcgtcg ggctcgacat cggcaaggtg tgggtcgcgg acgacggcgc cgcggtggcg
     5581 gtctggacca cgccggagag cgtcgaagcg ggggcggtgt tcgccgagat cggcccgcgc
     5641 atggccgagt tgagcggttc ccggctggcc gcgcagcaac agatggaggg cctcctggcg
     5701 ccgcaccggc ccaaggagcc cgcgtggttc ctggccaccg tcggcgtctc gcccgaccac
     5761 cagggcaagg gtctgggcag cgccgtcgtg ctccccggag tggaggccgc cgagcgcgcc
     5821 ggggtgcccg ccttcctgga gacctccgcg ccccgcaacc tccccttcta cgagcggctc
     5881 ggcttcaccg tcaccgccga cgtcgaggtg cccgaaggac cgcgcacctg gtgcatgacc
     5941 cgcaagcccg gtgcctaggc tagtatgtaa gcctagtctt agataataaa atcgctatcc
     6001 atcgaagatg gatgtgtgtt ggttttttgt gtgtgtaacg ctaggcgcgc ctggtgtacc
     6061 gagaacgatc ctctcagtgc gagtctcgac gatccatatc gttgcttggc agtcagccag
     6121 tcggaatcca gcttgggacc caggaagtcc aatcgtcaga tattgtactc aagcctggtc
     6181 acggcagcgt accgatctgt ttaaacctag atattgatag tctgatcggt caacgtataa
     6241 tcgagtccta gcttttgcaa acatctatca agagacagga tcagcaggag gctttcgcat
     6301 gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt
     6361 ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcgcg
     6421 agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga
     6481 agaacgcttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg
     6541 tattgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt
     6601 tgagtattca ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg
     6661 cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga caacgattgg
     6721 aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga
     6781 tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc
     6841 tgtagcaatg gcaacaacct tgcgtaaact attaactggc gaactactta ctctagcttc
     6901 ccggcaacag ttgatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc
     6961 ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc gtgggtctcg
     7021 cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac
     7081 gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc
     7141 actgattaag cattggtaac cgattctagg tgcattggcg cagaaaaaaa tgcctgatgc
     7201 gacgctgcgc gtcttatact cccacatatg ccagattcag caacggatac ggcttcccca
     7261 acttgcccac ttccatacgt gtcctcctta ccagaaattt atccttaaga tcgtttaaac
     7321 tcgactctgg ctctatcgaa tctccgtcgt ttcgagctta cgcgaacagc cgtggcgctc
     7381 atttgctcgt cgggcatcga atctcgtcag ctatcgtcag cttacctttt tggcagcgat
     7441 cgcggctccc gacatcttgg accattagct ccacaggtat cttcttccct ctagtggtca
     7501 taacagcagc ttcagctacc tctcaattca aaaaacccct caagacccgt ttagaggccc
     7561 caaggggtta tgctatcaat cgttgcgtta cacacacaaa aaaccaacac acatccatct
     7621 tcgatggata gcgattttat tatctaactg ctgatcgagt gtagccagat ctagtaatca
     7681 attacggggt cattagttca tagcccatat atggagttcc gcgttacata acttacggta
     7741 aatggcccgc ctggctgacc gcccaacgac ccccgcccat tgacgtcaat aatgacgtat
     7801 gttcccatag taacgccaat agggactttc cattgacgtc aatgggtgga gtatttacgg
     7861 taaactgccc acttggcagt acatcaagtg tatcatatgc caagtacgcc ccctattgac
     7921 gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt acatgacctt atgggacttt
     7981 cctacttggc agtacatcta cgtattagtc atcgctatta ccatgctgat gcggttttgg
     8041 cagtacatca atgggcgtgg atagcggttt gactcacggg gatttccaag tctccacccc
     8101 attgacgtca atgggagttt gttttggcac caaaatcaac gggactttcc aaaatgtcgt
     8161 aacaactccg ccccattgac gcaaatgggc ggtaggcgtg tacggtggga ggtctatata
     8221 agcagagctg gtttagtgaa ccgtcagatc agatctttgt cgatcctacc atccactcga
     8281 cacacccgcc agcggccgc
//
