LOCUS       sequence-295985-        7020 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Mammalian expression plasmid for SARS-CoV-2 RNA-dependent RNA 
            polymerase.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7020)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 7020)
  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-m4133484
FEATURES             Location/Qualifiers
     source          1..7020
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             2798..2818
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             2819..2836
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     polyA_signal    2889..3010
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2900..2919)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2954..2973
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      3125..3171
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      3201..3337
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    3382..3453
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(3679..4267)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3759..3778)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             join(5013..5081,5082..5873)
                     /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(5231..5250)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      5886..6022
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    6174..6245
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(6250..6296)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    6314..6362
                     /note="synthetic polyadenylation signal"
     enhancer        6443..6746
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6748..6950
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     6901..6921
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      join(7019..7020,1..8)
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
ORIGIN
        1 caccatggct gatgcacaat cgtttttaaa ccgggtttgc ggtgtaagtg cagcccgtct
       61 tacaccgtgc ggcacaggca ctagtactga tgtcgtatac agggcttttg acatctacaa
      121 tgataaagta gctggttttg ctaaattcct aaaaactaat tgttgtcgct tccaagaaaa
      181 ggacgaagat gacaatttaa ttgattctta ctttgtagtt aagagacaca ctttctctaa
      241 ctaccaacat gaagaaacaa tttataattt acttaaggat tgtccagctg ttgctaaaca
      301 tgacttcttt aagtttagaa tagacggtga catggtacca catatatcac gtcaacgtct
      361 tactaaatac acaatggcag acctcgtcta tgctttaagg cattttgatg aaggtaattg
      421 tgacacatta aaagaaatac ttgtcacata caattgttgt gatgatgatt atttcaataa
      481 aaaggactgg tatgattttg tagaaaaccc agatatatta cgcgtatacg ccaacttagg
      541 tgaacgtgta cgccaagctt tgttaaaaac agtacaattc tgtgatgcca tgcgaaatgc
      601 tggtattgtt ggtgtactga cattagataa tcaagatctc aatggtaact ggtatgattt
      661 cggtgatttc atacaaacca cgccaggtag tggagttcct gttgtagatt cttattattc
      721 attgttaatg cctatattaa ccttgaccag ggctttaact gcagagtcac atgttgacac
      781 tgacttaaca aagccttaca ttaagtggga tttgttaaaa tatgacttca cggaagagag
      841 gttaaaactc tttgaccgtt attttaaata ttgggatcag acataccacc caaattgtgt
      901 taactgtttg gatgacagat gcattctgca ttgtgcaaac tttaatgttt tattctctac
      961 agtgttccca cctacaagtt ttggaccact agtgagaaaa atatttgttg atggtgttcc
     1021 atttgtagtt tcaactggat accacttcag agagctaggt gttgtacata atcaggatgt
     1081 aaacttacat agctctagac ttagttttaa ggaattactt gtgtatgctg ctgaccctgc
     1141 tatgcacgct gcttctggta atctattact agataaacgc actacgtgct tttcagtagc
     1201 tgcacttact aacaatgttg cttttcaaac tgtcaaaccc ggtaatttta acaaagactt
     1261 ctatgacttt gctgtgtcta agggtttctt taaggaagga agttctgttg aattaaaaca
     1321 cttcttcttt gctcaggatg gtaatgctgc tatcagcgat tatgactact atcgttataa
     1381 tctaccaaca atgtgtgata tcagacaact actatttgta gttgaagttg ttgataagta
     1441 ctttgattgt tacgatggtg gctgtattaa tgctaaccaa gtcatcgtca acaacctaga
     1501 caaatcagct ggttttccat ttaataaatg gggtaaggct agactttatt atgattcaat
     1561 gagttatgag gatcaagatg cacttttcgc atatacaaaa cgtaatgtca tccctactat
     1621 aactcaaatg aatcttaagt atgccattag tgcaaagaat agagctcgca ccgtagctgg
     1681 tgtctctatc tgtagtacta tgaccaatag acagtttcat caaaaattat tgaaatcaat
     1741 agccgccact agaggagcta ctgtagtaat tggaacaagc aaattctatg gtggttggca
     1801 caacatgtta aaaactgttt atagtgatgt agaaaaccct caccttatgg gttgggatta
     1861 tcctaaatgt gatagagcca tgcctaacat gcttagaatt atggcctcac ttgttcttgc
     1921 tcgcaaacat acaacgtgtt gtagcttgtc acaccgtttc tatagattag ctaatgagtg
     1981 tgctcaagta ttgagtgaaa tggtcatgtg tggcggttca ctatatgtta aaccaggtgg
     2041 aacctcatca ggagatgcca caactgctta tgctaatagt gtttttaaca tttgtcaagc
     2101 tgtcacggcc aatgttaatg cacttttatc tactgatggt aacaaaattg ccgataagta
     2161 tgtccgcaat ttacaacaca gactttatga gtgtctctat agaaatagag atgttgacac
     2221 agactttgtg aatgagtttt acgcatattt gcgtaaacat ttctcaatga tgatactctc
     2281 tgacgatgct gttgtgtgtt tcaatagcac ttatgcatct caaggtctag tggctagcat
     2341 aaagaacttt aagtcagttc tttattatca aaacaatgtt tttatgtctg aagcaaaatg
     2401 ttggactgag actgacctta ctaaaggacc tcatgaattt tgctctcaac atacaatgct
     2461 agttaaacag ggtgatgatt atgtgtacct tccttaccca gatccatcaa gaatcctagg
     2521 ggccggctgt tttgtagatg atatcgtaaa aacagatggt acacttatga ttgaacggtt
     2581 cgtgtcttta gctatagatg cttacccact tactaaacat cctaatcagg agtatgctga
     2641 tgtctttcat ttgtacttac aatacataag aaagctacat gatgagttaa caggacacat
     2701 gttagacatg tattctgtta tgcttactaa tgataacact tcaaggtatt gggaacctga
     2761 gttttatgag gctatgtaca caccgcatac agtcttagaa aacctttact ttcagtcgca
     2821 ccatcaccat caccattaag ctagcttgac tgactgagat acagcgtacc ttcagctcac
     2881 agacatgata agatacattg atgagtttgg acaaaccaca actagaatgc agtgaaaaaa
     2941 atgctttatt tgtgaaattt gtgatgctat tgctttattt gtaaccatta taagctgcaa
     3001 taaacaagtt aacaacaaca attgcattca ttttatgttt caggttcagg gggaggtgtg
     3061 ggaggttttt taaagcaagt aaaacctcta caaatgtggt attggcccat ctctatcggt
     3121 atcgtagcat aaccccttgg ggcctctaaa cgggtcttga ggggtttttt gtgcccctcg
     3181 ggccggattg ctatctaccg gcattggcgc agaaaaaaat gcctgatgcg acgctgcgcg
     3241 tcttatactc ccacatatgc cagattcagc aacggatacg gcttccccaa cttgcccact
     3301 tccatacgtg tcctccttac cagaaattta tccttaaggt cgtcagctat cctgcaggcg
     3361 atctctcgat ttcgatcaag acattccttt aatggtcttt tctggacacc actaggggtc
     3421 agaagtagtt catcaaactt tcttccctcc ctaatctcat tggttacctt gggctatcga
     3481 aacttaatta accagtcaag tcagctactt ggcgagatcg acttgtctgg gtttcgacta
     3541 cgctcagaat tgcgtcagtc aagttcgatc tggtccttgc tattgcaccc gttctccgat
     3601 tacgagtttc atttaaatca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     3661 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     3721 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     3781 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     3841 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
     3901 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     3961 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     4021 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     4081 gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc
     4141 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     4201 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     4261 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     4321 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
     4381 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta
     4441 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     4501 tgcatttaaa tttccgaact ctccaaggcc ctcgtcggaa aatcttcaaa cctttcgtcc
     4561 gatccatctt gcaggctacc tctcgaacga actatcgcaa gtctcttggc cggccttgcg
     4621 ccttggctat tgcttggcag cgcctatcgc caggtattac tccaatcccg aatatccgag
     4681 atcgggatca cccgagagaa gttcaaccta catcctcaat cccgatctat ccgagatccg
     4741 aggaatatcg aaatcggggc gcgcctggtg taccgagaac gatcctctca gtgcgagtct
     4801 cgacgatcca tatcgttgct tggcagtcag ccagtcggaa tccagcttgg gacccaggaa
     4861 gtccaatcgt cagatattgt actcaagcct ggtcacggca gcgtaccgat ctgtttaaac
     4921 ctagatattg atagtctgat cggtcaacgt ataatcgagt cctagctttt gcaaacatct
     4981 atcaagagac aggatcagca ggaggctttc gcatgagtat tcaacatttc cgtgtcgccc
     5041 ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga
     5101 aagtaaaaga tgctgaagat cagttgggtg cgcgagtggg ttacatcgaa ctggatctca
     5161 acagcggtaa gatccttgag agttttcgcc ccgaagaacg ctttccaatg atgagcactt
     5221 ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg
     5281 gtcgccgcat acactattct cagaatgact tggttgagta ttcaccagtc acagaaaagc
     5341 atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata
     5401 acactgcggc caacttactt ctgacaacga ttggaggacc gaaggagcta accgcttttt
     5461 tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag
     5521 ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca accttgcgta
     5581 aactattaac tggcgaacta cttactctag cttcccggca acagttgata gactggatgg
     5641 aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg
     5701 ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag
     5761 atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg
     5821 aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg taaccgattc
     5881 taggtgcatt ggcgcagaaa aaaatgcctg atgcgacgct gcgcgtctta tactcccaca
     5941 tatgccagat tcagcaacgg atacggcttc cccaacttgc ccacttccat acgtgtcctc
     6001 cttaccagaa atttatcctt aagatcccga atcgtttaaa ctcgactctg gctctatcga
     6061 atctccgtcg tttcgagctt acgcgaacag ccgtggcgct catttgctcg tcgggcatcg
     6121 aatctcgtca gctatcgtca gcttaccttt ttggcagcga tcgcggctcc cgacatcttg
     6181 gaccattagc tccacaggta tcttcttccc tctagtggtc ataacagcag cttcagctac
     6241 ctctcaattc aaaaaacccc tcaagacccg tttagaggcc ccaaggggtt atgctatcaa
     6301 tcgttgcgtt acacacacaa aaaaccaaca cacatccatc ttcgatggat agcgatttta
     6361 ttatctaact gctgatcgag tgtagccaga tctagtaatc aattacgggg tcattagttc
     6421 atagcccata tatggagttc cgcgttacat aacttacggt aaatggcccg cctggctgac
     6481 cgcccaacga cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa
     6541 tagggacttt ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag
     6601 tacatcaagt gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc
     6661 ccgcctggca ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct
     6721 acgtattagt catcgctatt accatgctga tgcggttttg gcagtacatc aatgggcgtg
     6781 gatagcggtt tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt
     6841 tgttttggca ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga
     6901 cgcaaatggg cggtaggcgt gtacggtggg aggtctatat aagcagagct ggtttagtga
     6961 accgtcagat cagatctttg tcgatcctac catccactcg acacacccgc cagcggccgc
//
