LOCUS       sequence-421875-        4713 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses the MTase domain of NS5 of ZIKV.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4713)
  AUTHORS   Meyer C, Garzia A, Miller MW, Huggins DJ, Myers RW, Hoffmann HH, 
            Ashbrook AW, Jannath SY, Liverton N, Kargman S, Zimmerman M, Nelson 
            AM, Sharma V, Dolgov E, Cangialosi J, Penalva-Lopez S, Alvarez N, 
            Chang CW, Oswal N, Gonzalez I, Rasheed R, Goldgirsh K, Davis JA, 
            Ramos-Espiritu L, Menezes MR, Larson C, Nitsche J, Ganichkin O, 
            Alwaseem H, Molina H, Steinbacher S, Glickman JF, Perlin DS, Rice 
            CM, Meinke PT, Tuschl T
  TITLE     Small-molecule inhibition of SARS-CoV-2 NSP14 RNA cap 
            methyltransferase.
  JOURNAL   Nature. 2024 Dec 11. doi: 10.1038/s41586-024-08320-0.
  PUBMED    39663451
REFERENCE   2  (bases 1 to 4713)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nature. 
            2024 Dec 11. doi: 10.1038/s41586-024-08320-0."
COMMENT     Sequence Label: pRSF_his6_sumo_ZIKV_NS5_MTase
FEATURES             Location/Qualifiers
     source          1..4713
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     812..831
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(851..928)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     974..993
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        974..992
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    993..1017
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     CDS             1073..1090
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1100..1117
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             1130..1423
                     /codon_start=1
                     /gene="S. cerevisiae SMT3 (truncated)"
                     /product="cleavable ubiquitin-like protein tag"
                     /label=SUMO
                     /translation="MSDSEVNQEAKPEVKPEVKPETHINLKVSDGSSEIFFKIKKTTPL
                     RRLMEAFAKRQGKEMDSLRFLYDGIRIQADQTPEDLDMEDNDIIEAHREQIGG"
     CDS             2240..2284
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(2322..2340)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2336..2383
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(2613..3428)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(2613..3428)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     3336..3355
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(3429..3520)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(3536..4285)
                     /direction=LEFT
                     /label=RSF ori
                     /note="Plasmids containing the RSF 1030 origin of 
                     replication can be propagated in E. coli cells that contain
                     additional plasmids with compatible origins."
     primer_bind     complement(3546..3563)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(3786..3805)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4481..4713,1..850))
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=lacI
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
ORIGIN
        1 ggatataaca tgagctgtct tcggtatcgt cgtatcccac taccgagatg tccgcaccaa
       61 cgcgcagccc ggactcggta atggcgcgca ttgcgcccag cgccatctga tcgttggcaa
      121 ccagcatcgc agtgggaacg atgccctcat tcagcatttg catggtttgt tgaaaaccgg
      181 acatggcact ccagtcgcct tcccgttccg ctatcggctg aatttgattg cgagtgagat
      241 atttatgcca gccagccaga cgcagacgcg ccgagacaga acttaatggg cccgctaaca
      301 gcgcgatttg ctggtgaccc aatgcgacca gatgctccac gcccagtcgc gtaccgtctt
      361 catgggagaa aataatactg ttgatgggtg tctggtcaga gacatcaaga aataacgccg
      421 gaacattagt gcaggcagct tccacagcaa tggcatcctg gtcatccagc ggatagttaa
      481 tgatcagccc actgacgcgt tgcgcgagaa gattgtgcac cgccgcttta caggcttcga
      541 cgccgcttcg ttctaccatc gacaccacca cgctggcacc cagttgatcg gcgcgagatt
      601 taatcgccgc gacaatttgc gacggcgcgt gcagggccag actggaggtg gcaacgccaa
      661 tcagcaacga ctgtttgccc gccagttgtt gtgccacgcg gttgggaatg taattcagct
      721 ccgccatcgc cgcttccact ttttcccgcg ttttcgcaga aacgtggctg gcctggttca
      781 ccacgcggga aacggtctga taagagacac cggcatactc tgcgacatcg tataacgtta
      841 ctggtttcac attcaccacc ctgaattgac tctcttccgg gcgctatcat gccataccgc
      901 gaaaggtttt gcgccattcg atggtgtccg ggatctcgac gctctccctt atgcgactcc
      961 tgcattagga aattaatacg actcactata ggggaattgt gagcggataa caattcccct
     1021 gtagaaataa ttttgtttaa ctttaataag gagatatacc atgggcagca gccatcatca
     1081 tcatcatcac agcagcggcc tggtgccgcg cggcagccac atggcaagca tgtcggactc
     1141 agaagtcaat caagaagcta agccagaggt caagccagaa gtcaagcctg agactcacat
     1201 caatttaaag gtgtccgatg gatcttcaga gatcttcttc aagatcaaaa agaccactcc
     1261 tttaagaagg ctgatggaag cgttcgctaa aagacagggt aaggaaatgg actccttaag
     1321 attcttgtac gacggtatta gaatccaagc tgatcagacc cctgaagatt tggacatgga
     1381 ggataacgat attattgagg ctcacagaga acagattggt ggatccgggg gtggaacggg
     1441 agagaccctg ggagagaaat ggaaggcccg cctgaaccag atgtcggccc tggagttcta
     1501 ctcctacaaa aagtcaggca tcaccgaggt gtgcagagaa gaggcccgcc gcgccctcaa
     1561 ggacggtgtg gcaacgggag gccacgctgt gtcccgagga agtgcaaagc tgagatggtt
     1621 ggtggagagg ggatacctgc agccctatgg aaaggtcatt gatcttggat gtggcagagg
     1681 gggctggagt tactatgccg ccaccatccg caaagttcaa gaagtgaaag gatacacaaa
     1741 aggaggccct ggtcatgaag aacccatgtt ggtgcaaagc tatgggtgga acatagtccg
     1801 tcttaagagt ggggtggacg tctttcatat ggcggctgag ccgtgtgaca cgttgctgtg
     1861 tgatataggt gagtcatcat ctagtcctga agtggaagaa gcacggacgc tcagagtcct
     1921 ctccatggtg ggggattggc ttgaaaaaag accaggagcc ttttgtataa aagtgttgtg
     1981 cccatacacc agcactatga tggaaaccct ggagcgactg cagcgtaggt atgggggagg
     2041 actggtcaga gtgccactct cccgcaactc tacacatgag atgtactggg tctctggagc
     2101 gaaaagcaac accataaaaa gtgtgtccac cacgagccag ctccttttgg ggcgcatgga
     2161 cgggcccagg aggccagtga aatatgaaga ggatgtgaat ctcggctctg gcacgcgggc
     2221 ttgaccgctc gagtctggta aagaaaccgc tgctgcgaaa tttgaacgcc agcacatgga
     2281 ctcgtctact agcgcagctt aattaaccta ggctgctgcc accgctgagc aataactagc
     2341 ataacccctt ggggcctcta aacgggtctt gaggggtttt ttgctgaaac ctcaggcatt
     2401 tgagaagcac acggtcacac tgcttccggt agtcaataaa ccggtaaacc agcaatagac
     2461 ataagcggct atttaacgac cctgccctga accgacgaca agctgacgac cgggtctccg
     2521 caagtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac
     2581 attcaaatat gtatccgctc atgaattaat tcttagaaaa actcatcgag catcaaatga
     2641 aactgcaatt tattcatatc aggattatca ataccatatt tttgaaaaag ccgtttctgt
     2701 aatgaaggag aaaactcacc gaggcagttc cataggatgg caagatcctg gtatcggtct
     2761 gcgattccga ctcgtccaac atcaatacaa cctattaatt tcccctcgtc aaaaataagg
     2821 ttatcaagtg agaaatcacc atgagtgacg actgaatccg gtgagaatgg caaaagttta
     2881 tgcatttctt tccagacttg ttcaacaggc cagccattac gctcgtcatc aaaatcactc
     2941 gcatcaacca aaccgttatt cattcgtgat tgcgcctgag cgagacgaaa tacgcggtcg
     3001 ctgttaaaag gacaattaca aacaggaatc gaatgcaacc ggcgcaggaa cactgccagc
     3061 gcatcaacaa tattttcacc tgaatcagga tattcttcta atacctggaa tgctgttttc
     3121 ccggggatcg cagtggtgag taaccatgca tcatcaggag tacggataaa atgcttgatg
     3181 gtcggaagag gcataaattc cgtcagccag tttagtctga ccatctcatc tgtaacatca
     3241 ttggcaacgc tacctttgcc atgtttcaga aacaactctg gcgcatcggg cttcccatac
     3301 aatcgataga ttgtcgcacc tgattgcccg acattatcgc gagcccattt atacccatat
     3361 aaatcagcat ccatgttgga atttaatcgc ggcctagagc aagacgtttc ccgttgaata
     3421 tggctcatac tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg
     3481 agcggataca tatttgaatg tatttagaaa aataaacaaa taggcatgca gcgctcttcc
     3541 gcttcctcgc tcactgactc gctacgctcg gtcgttcgac tgcggcgagc ggtgtcagct
     3601 cactcaaaag cggtaatacg gttatccaca gaatcagggg ataaagccgg aaagaacatg
     3661 tgagcaaaaa gcaaagcacc ggaagaagcc aacgccgcag gcgtttttcc ataggctccg
     3721 cccccctgac gagcatcaca aaaatcgacg ctcaagccag aggtggcgaa acccgacagg
     3781 actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac
     3841 cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca
     3901 tagctcacgc tgttggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt
     3961 gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc
     4021 caacccggta agacacgact tatcgccact ggcagcagcc attggtaact gatttagagg
     4081 actttgtctt gaagttatgc acctgttaag gctaaactga aagaacagat tttggtgagt
     4141 gcggtcctcc aacccactta ccttggttca aagagttggt agctcagcga accttgagaa
     4201 aaccaccgtt ggtagcggtg gtttttcttt atttatgaga tgatgaatca atcggtctat
     4261 caagtcaacg aacagctatt ccgttactct agatttcagt gcaatttatc tcttcaaatg
     4321 tagcacctga agtcagcccc atacgatata agttgtaatt ctcatgttag tcatgccccg
     4381 cgcccaccgg aaggagctga ctgggttgaa ggctctcaag ggcatcggtc gagatcccgg
     4441 tgcctaatga gtgagctaac ttacattaat tgcgttgcgc tcactgcccg ctttccagtc
     4501 gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt
     4561 gcgtattggg cgccagggtg gtttttcttt tcaccagtga gacgggcaac agctgattgc
     4621 ccttcaccgc ctggccctga gagagttgca gcaagcggtc cacgctggtt tgccccagca
     4681 ggcgaaaatc ctgtttgatg gtggttaacg gcg
//
