LOCUS       Exported                7967 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Mammalian expression plasmid for SARS-CoV-2 helicase.
ACCESSION   .
VERSION     .
KEYWORDS    pGBW-m4133171
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7967)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 7967)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7967
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     CDS             1811..1831
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             1832..1876
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     polyA_signal    1929..2050
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1940..1959)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1994..2013
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      2165..2211
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      2241..2377
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    2422..2493
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(2719..3307)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2799..2818)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        3807..5016
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     4853..4872
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             5020..5619
                     /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(5020..5039)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5516..5536
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    5646..5694
                     /note="synthetic polyadenylation signal"
     CDS             5960..6820
                     /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(6178..6197)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      6833..6969
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    7121..7192
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      7197..7243
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    7261..7309
                     /note="synthetic polyadenylation signal"
     enhancer        7390..7693
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        7695..7897
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     7848..7868
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      7966..8
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
ORIGIN
        1 caccatggcg gtgggggctt gcgtactatg taacagccag acgtcattgc gctgcggtgc
       61 atgtataaga aggccgtttc tgtgttgcaa atgctgctac gatcatgtta tttcgacttc
      121 tcacaagctt gtgctcagtg tcaatcctta tgtgtgtaat gcccccggat gcgacgtcac
      181 agatgttacc caactgtacc tgggcggcat gtcctattat tgtaagagcc acaaaccacc
      241 aatctccttc cccctgtgcg ccaacggaca ggtgtttggg ctctacaaaa acacatgtgt
      301 gggatctgac aatgtcaccg acttcaacgc tatcgccaca tgcgattgga ctaatgcagg
      361 cgactacatt ttagccaaca cctgtaccga acggctgaag ttatttgcgg ctgagactct
      421 aaaggcaact gaggaaacct tcaaactttc atatggtatc gccacggtac gtgaggtgtt
      481 gtctgatcga gaactgcatc tcagttggga ggtcgggaag cctagacctc cccttaatcg
      541 aaattacgtg ttcacaggtt atagggttac caagaacagc aaagtccaga tcggcgaata
      601 tacttttgag aagggagatt acggtgacgc cgtagtgtat cgtggcacaa caacatacaa
      661 actgaatgtt ggggattact tcgtgctgac gtcccacacc gttatgccgc tctcagctcc
      721 aactttggtt ccccaggaac attacgtgcg gattactgga ctctatccaa cgctaaacat
      781 aagcgacgag ttttccagca acgtggctaa ttaccaaaag gtcggcatgc agaagtactc
      841 cacccttcag ggtcctcctg ggactggcaa aagtcacttc gccatcggac tggcacttta
      901 ctatcccagt gcgcgcattg tatataccgc ctgctcacat gcggcagtgg acgccctgtg
      961 tgagaaggct ttaaagtatc tgccaataga caaatgctct cgcattatcc ctgccagggc
     1021 cagagtggag tgctttgata agtttaaagt gaactcgacc ctcgaacagt atgtcttctg
     1081 cacagttaac gcattgcccg aaacgaccgc cgatatagtt gtgttcgacg aaatctcaat
     1141 ggctaccaat tatgatctct ccgtcgtcaa tgcacggctg cgggccaaac actacgtcta
     1201 catcggggac cccgcgcagt taccagcacc gcgaacccta ctgactaagg gcacacttga
     1261 gccagagtac ttcaacagcg tatgtagact gatgaaaacc attggccctg atatgttcct
     1321 gggaacatgt cggaggtgcc ccgctgaaat agtggacacc gtttcagcac ttgtgtacga
     1381 caataagctc aaagcacaca aggataaatc cgcacaatgt tttaagatgt tttacaaagg
     1441 ggttattacc catgatgtgt ctagcgctat caatcgccct cagattggcg tggtgagaga
     1501 gtttttgacg cgaaatcccg cctggcgaaa ggccgtattc atcagtcctt acaactcgca
     1561 aaatgccgtc gctagtaaga ttttaggact ccctactcag acagttgata gctctcaagg
     1621 ttctgaatac gattatgtga tttttactca gactaccgaa acggcacatt cttgtaacgt
     1681 caaccgtttt aacgtggcca ttacaagggc caaggttggc atcttgtgta tcatgtcaga
     1741 ccgggacctg tatgataaac tgcaattcac atctttggag ataccgcgcc gcaatgtcgc
     1801 tacccttcag gaaaaccttt actttcagtc gaaagaaacc gcggcagcta agttcgagag
     1861 gcagcacatg gacagttaag ctagcttgac tgactgagat acagcgtacc ttcagctcac
     1921 agacatgata agatacattg atgagtttgg acaaaccaca actagaatgc agtgaaaaaa
     1981 atgctttatt tgtgaaattt gtgatgctat tgctttattt gtaaccatta taagctgcaa
     2041 taaacaagtt aacaacaaca attgcattca ttttatgttt caggttcagg gggaggtgtg
     2101 ggaggttttt taaagcaagt aaaacctcta caaatgtggt attggcccat ctctatcggt
     2161 atcgtagcat aaccccttgg ggcctctaaa cgggtcttga ggggtttttt gtgcccctcg
     2221 ggccggattg ctatctaccg gcattggcgc agaaaaaaat gcctgatgcg acgctgcgcg
     2281 tcttatactc ccacatatgc cagattcagc aacggatacg gcttccccaa cttgcccact
     2341 tccatacgtg tcctccttac cagaaattta tccttaaggt cgtcagctat cctgcaggcg
     2401 atctctcgat ttcgatcaag acattccttt aatggtcttt tctggacacc actaggggtc
     2461 agaagtagtt catcaaactt tcttccctcc ctaatctcat tggttacctt gggctatcga
     2521 aacttaatta accagtcaag tcagctactt ggcgagatcg acttgtctgg gtttcgacta
     2581 cgctcagaat tgcgtcagtc aagttcgatc tggtccttgc tattgcaccc gttctccgat
     2641 tacgagtttc atttaaatca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     2701 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     2761 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     2821 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     2881 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
     2941 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     3001 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     3061 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     3121 gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc
     3181 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     3241 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     3301 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     3361 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
     3421 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta
     3481 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     3541 tgcatttaaa tttccgaact ctccaaggcc ctcgtcggaa aatcttcaaa cctttcgtcc
     3601 gatccatctt gcaggctacc tctcgaacga actatcgcaa gtctcttggc cggccttgcg
     3661 ccttggctat tgcttggcag cgcctatcgc caggtattac tccaatcccg aatatccgag
     3721 atcgggatca cccgagagaa gttcaaccta catcctcaat cccgatctat ccgagatccg
     3781 aggaatatcg aaatcggggc gcgcctggcc tccgcgccgg gttttggcgc ctcccgcggg
     3841 cgcccccctc gtcacggcga gcgctgccac gtcagacgaa gggcgcagga gcgtcctgat
     3901 ccttccgccc ggacgctcag gacagcggcc cgctgctcat aagactcggc cttagaaccc
     3961 cagtatcagc agaaggacat tttaggacgg gacttgggtg actctagggc actggttttc
     4021 tttccagaga gcggaacagg cgaggaaaag tagtcccttc tcggcgattc tgcggaggga
     4081 tctccgtggg gcggtgaacg ccgatgatta tataaggacg cgccgggtgt ggcacagcta
     4141 gttccgtcgc agccgggatt tgggtcgcgg ttcttgtttg tggatcgctg tgatcgtcac
     4201 ttggtgagta gcgggctgct gggctggccg gggctttcgt ggccgccggg ccgctcggtg
     4261 ggacggaagc gtgtggagag accgccaagg gctgtagtct gggtccgcga gcaaggttgc
     4321 cctgaactgg gggttggggg gagcgcagca aaatggcggc tgttcccgag tcttgaatgg
     4381 aagacgcttg tgaggcgggc tgtgaggtcg ttgaaacaag gtggggggca tggtgggcgg
     4441 caagaaccca aggtcttgag cccttcgcta atgcgggaaa gctcttattc gggtgagatg
     4501 ggctgggcac catctgggga ccctgacgtg aagtttgtca ctgactggag aactcggttt
     4561 gtcgtctgtt gcgggggcgg cagttatggc ggtgccgttg ggcagtgcac ccgtaccttt
     4621 gggagcgcgc gccctcgtcg tgtcgtgacg tcacccgttc tgttggctta taatgcaggg
     4681 tggggccacc tgccggtagg tgtgcggtag gcttttctcc gtcgcaggac gcagggttcg
     4741 ggcctagggt aggctctcct gaatcgacag gcgccggacc tctggtgagg ggagggataa
     4801 gtgaggcgtc agtttctttg gtcggtttta tgtacctatc ttcttaagta gctgaagctc
     4861 cggttttgaa ctatgcgctc ggggttggcg agtgtgtttt gtgaagtttt ttaggcacct
     4921 tttgaaatgt aatcatttgg gtcaatatgt aattttcagt gttagacttg taaattgtcc
     4981 gctaaattct ggccgttttt ggcttttttg ttagacaaca tgaccgagta caagcccacg
     5041 gtgcgcctcg ccacccgcga cgacgtcccc agggccgtac gcaccctcgc cgccgcgttc
     5101 gccgactacc ccgccacgcg ccacaccgtc gatccggacc gccacatcga gcgggtcacc
     5161 gagctgcaag aactcttcct cacgcgcgtc gggctcgaca tcggcaaggt gtgggtcgcg
     5221 gacgacggcg ccgcggtggc ggtctggacc acgccggaga gcgtcgaagc gggggcggtg
     5281 ttcgccgaga tcggcccgcg catggccgag ttgagcggtt cccggctggc cgcgcagcaa
     5341 cagatggagg gcctcctggc gccgcaccgg cccaaggagc ccgcgtggtt cctggccacc
     5401 gtcggcgtct cgcccgacca ccagggcaag ggtctgggca gcgccgtcgt gctccccgga
     5461 gtggaggccg ccgagcgcgc cggggtgccc gccttcctgg agacctccgc gccccgcaac
     5521 ctccccttct acgagcggct cggcttcacc gtcaccgccg acgtcgaggt gcccgaagga
     5581 ccgcgcacct ggtgcatgac ccgcaagccc ggtgcctagg ctagtatgta agcctagtct
     5641 tagataataa aatcgctatc catcgaagat ggatgtgtgt tggttttttg tgtgtgtaac
     5701 gctaggcgcg cctggtgtac cgagaacgat cctctcagtg cgagtctcga cgatccatat
     5761 cgttgcttgg cagtcagcca gtcggaatcc agcttgggac ccaggaagtc caatcgtcag
     5821 atattgtact caagcctggt cacggcagcg taccgatctg tttaaaccta gatattgata
     5881 gtctgatcgg tcaacgtata atcgagtcct agcttttgca aacatctatc aagagacagg
     5941 atcagcagga ggctttcgca tgagtattca acatttccgt gtcgccctta ttcccttttt
     6001 tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc
     6061 tgaagatcag ttgggtgcgc gagtgggtta catcgaactg gatctcaaca gcggtaagat
     6121 ccttgagagt tttcgccccg aagaacgctt tccaatgatg agcactttta aagttctgct
     6181 atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca
     6241 ctattctcag aatgacttgg ttgagtattc accagtcaca gaaaagcatc ttacggatgg
     6301 catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa
     6361 cttacttctg acaacgattg gaggaccgaa ggagctaacc gcttttttgc acaacatggg
     6421 ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga
     6481 cgagcgtgac accacgatgc ctgtagcaat ggcaacaacc ttgcgtaaac tattaactgg
     6541 cgaactactt actctagctt cccggcaaca gttgatagac tggatggagg cggataaagt
     6601 tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg
     6661 agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc
     6721 ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca
     6781 gatcgctgag ataggtgcct cactgattaa gcattggtaa ccgattctag gtgcattggc
     6841 gcagaaaaaa atgcctgatg cgacgctgcg cgtcttatac tcccacatat gccagattca
     6901 gcaacggata cggcttcccc aacttgccca cttccatacg tgtcctcctt accagaaatt
     6961 tatccttaag atcccgaatc gtttaaactc gactctggct ctatcgaatc tccgtcgttt
     7021 cgagcttacg cgaacagccg tggcgctcat ttgctcgtcg ggcatcgaat ctcgtcagct
     7081 atcgtcagct tacctttttg gcagcgatcg cggctcccga catcttggac cattagctcc
     7141 acaggtatct tcttccctct agtggtcata acagcagctt cagctacctc tcaattcaaa
     7201 aaacccctca agacccgttt agaggcccca aggggttatg ctatcaatcg ttgcgttaca
     7261 cacacaaaaa accaacacac atccatcttc gatggatagc gattttatta tctaactgct
     7321 gatcgagtgt agccagatct agtaatcaat tacggggtca ttagttcata gcccatatat
     7381 ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc
     7441 ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag ggactttcca
     7501 ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac atcaagtgta
     7561 tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg cctggcatta
     7621 tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg tattagtcat
     7681 cgctattacc atgctgatgc ggttttggca gtacatcaat gggcgtggat agcggtttga
     7741 ctcacgggga tttccaagtc tccaccccat tgacgtcaat gggagtttgt tttggcacca
     7801 aaatcaacgg gactttccaa aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg
     7861 taggcgtgta cggtgggagg tctatataag cagagctggt ttagtgaacc gtcagatcag
     7921 atctttgtcg atcctaccat ccactcgaca cacccgccag cggccgc
//
