LOCUS       sequence-473709-        5724 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  bacterial protein expression of NSP10.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5724)
  AUTHORS   Rona G, Zeke A, Miwatani-Minter B, de Vries M, Kaur R, Schinlever A,
            Garcia SF, Goldberg HV, Wang H, Hinds TR, Bailly F, Zheng N, Cotelle
            P, Desmaele D, Landau NR, Dittmann M, Pagano M
  TITLE     The NSP14/NSP10 RNA repair complex as a Pan-coronavirus therapeutic 
            target.
  JOURNAL   Cell Death Differ. 2022 Feb;29(2):285-292. doi: 
            10.1038/s41418-021-00900-1. Epub 2021 Dec 3.
  PUBMED    34862481
REFERENCE   2  (bases 1 to 5724)
  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"; doi: 
            "10.1038/s41418-021-00900-1"; journalName: "Cell Death Differ"; 
            date: "2022-02"; volume: "29"; issue: "2"; pages: "285-292"
COMMENT     Sequence Label: pET-30a-NSP10-MERS-CoV
FEATURES             Location/Qualifiers
     source          1..5724
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(142..597)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(279..300)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     491..510
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     terminator      complement(634..681)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     677..695
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(748..765)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(793..810)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             1265..1287
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    1302..1326
                     /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)."
     primer_bind     complement(1326..1345)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1327..1345)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1433..1452)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        1658..1735
                     /gene="lacI"
                     /label=lacI promoter
     CDS             1736..2818
                     /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"
     primer_bind     complement(1755..1774)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             3627..3818
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(3812..3834)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3920..4062
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3975..3994
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4077..4094)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4248..4836)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4328..4347)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(4947..4966)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             join(4958..5724,1..49)
                     /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             join(4958..5724,1..49)
                     /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     complement(5031..5050)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
ORIGIN
        1 tgatatgaat aaattgcagt ttcatttgat gctcgatgag tttttctaag aattaattca
       61 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
      121 ttccccgaaa agtgccacct gaaattgtaa acgttaatat tttgttaaaa ttcgcgttaa
      181 atttttgtta aatcagctca ttttttaacc aataggccga aatcggcaaa atcccttata
      241 aatcaaaaga atagaccgag atagggttga gtgttgttcc agtttggaac aagagtccac
      301 tattaaagaa cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc
      361 cactacgtga accatcaccc taatcaagtt ttttggggtc gaggtgccgt aaagcactaa
      421 atcggaaccc taaagggagc ccccgattta gagcttgacg gggaaagccg gcgaacgtgg
      481 cgagaaagga agggaagaaa gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg
      541 tcacgctgcg cgtaaccacc acacccgccg cgcttaatgc gccgctacag ggcgcgtccc
      601 attcgccaat ccggatatag ttcctccttt cagcaaaaaa cccctcaaga cccgtttaga
      661 ggccccaagg ggttatgcta gttattgctc agcggtggca gcagccaact cagcttcctt
      721 tcgggctttg ttagcagccg gatctcagtg gtggtggtgg tggtgctcga gtgcggccgc
      781 aagctttcat tagtggtggt ggtggtgatg ggagccaccg ccggagccgc caccttgcgg
      841 cagtgccgcc tggcgcaaag agtcgcagtt acaaccataa ccaatccaat actggcaaac
      901 attacacgga gtattgctca ggcaaaaacc aacaggatca cgaacacact gcgccgggat
      961 ctggacaaat ttacctttgt atttgcacac cccgctaaca tccggatgct cgatatgtgc
     1021 acggcagtac aggcagacgc tcgcgccgcc gtacgtctct tggtcagcgg tgctttccgg
     1081 cttcacgctg atcgcaatac cggtgccggt ctttggcgtc agcatcttca cgcaattggt
     1141 cagcggagca ccaccggcgt tcacgaagtc cagatacgct ttttgcggat ctacggtgaa
     1201 gttaaccaaa gaaagcaccg agctgttact tgcaaattct gtatttgatc cagccatatg
     1261 tatatctcct tcttaaagtt aaacaaaatt atttctagag gggaattgtt atccgctcac
     1321 aattccccta tagtgagtcg tattaatttc gcgggatcga gatcgatctc gatcctctac
     1381 gccggacgca tcgtggccgg catcaccggc gccacaggtg cggttgctgg cgcctatatc
     1441 gccgacatca ccgatgggga agatcgggct cgccacttcg ggctcatgag cgcttgtttc
     1501 ggcgtgggta tggtggcagg ccccgtggcc gggggactgt tgggcgccat ctccttgcat
     1561 gcaccattcc ttgcggcggc ggtgctcaac ggcctcaacc tactactggg ctgcttccta
     1621 atgcaggagt cgcataaggg agagcgtcga gatcccggac accatcgaat ggcgcaaaac
     1681 ctttcgcggt atggcatgat agcgcccgga agagagtcaa ttcagggtgg tgaatgtgaa
     1741 accagtaacg ttatacgatg tcgcagagta tgccggtgtc tcttatcaga ccgtttcccg
     1801 cgtggtgaac caggccagcc acgtttctgc gaaaacgcgg gaaaaagtgg aagcggcgat
     1861 ggcggagctg aattacattc ccaaccgcgt ggcacaacaa ctggcgggca aacagtcgtt
     1921 gctgattggc gttgccacct ccagtctggc cctgcacgcg ccgtcgcaaa ttgtcgcggc
     1981 gattaaatct cgcgccgatc aactgggtgc cagcgtggtg gtgtcgatgg tagaacgaag
     2041 cggcgtcgaa gcctgtaaag cggcggtgca caatcttctc gcgcaacgcg tcagtgggct
     2101 gatcattaac tatccgctgg atgaccagga tgccattgct gtggaagctg cctgcactaa
     2161 tgttccggcg ttatttcttg atgtctctga ccagacaccc atcaacagta ttattttctc
     2221 ccatgaagac ggtacgcgac tgggcgtgga gcatctggtc gcattgggtc accagcaaat
     2281 cgcgctgtta gcgggcccat taagttctgt ctcggcgcgt ctgcgtctgg ctggctggca
     2341 taaatatctc actcgcaatc aaattcagcc gatagcggaa cgggaaggcg actggagtgc
     2401 catgtccggt tttcaacaaa ccatgcaaat gctgaatgag ggcatcgttc ccactgcgat
     2461 gctggttgcc aacgatcaga tggcgctggg cgcaatgcgc gccattaccg agtccgggct
     2521 gcgcgttggt gcggacatct cggtagtggg atacgacgat accgaagaca gctcatgtta
     2581 tatcccgccg ttaaccacca tcaaacagga ttttcgcctg ctggggcaaa ccagcgtgga
     2641 ccgcttgctg caactctctc agggccaggc ggtgaagggc aatcagctgt tgcccgtctc
     2701 actggtgaaa agaaaaacca ccctggcgcc caatacgcaa accgcctctc cccgcgcgtt
     2761 ggccgattca ttaatgcagc tggcacgaca ggtttcccga ctggaaagcg ggcagtgagc
     2821 gcaacgcaat taatgtaagt tagctcactc attaggcacc gggatctcga ccgatgccct
     2881 tgagagcctt caacccagtc agctccttcc ggtgggcgcg gggcatgact atcgtcgccg
     2941 cacttatgac tgtcttcttt atcatgcaac tcgtaggaca ggtgccggca gcgctctggg
     3001 tcattttcgg cgaggaccgc tttcgctgga gcgcgacgat gatcggcctg tcgcttgcgg
     3061 tattcggaat cttgcacgcc ctcgctcaag ccttcgtcac tggtcccgcc accaaacgtt
     3121 tcggcgagaa gcaggccatt atcgccggca tggcggcccc acgggtgcgc atgatcgtgc
     3181 tcctgtcgtt gaggacccgg ctaggctggc ggggttgcct tactggttag cagaatgaat
     3241 caccgatacg cgagcgaacg tgaagcgact gctgctgcaa aacgtctgcg acctgagcaa
     3301 caacatgaat ggtcttcggt ttccgtgttt cgtaaagtct ggaaacgcgg aagtcagcgc
     3361 cctgcaccat tatgttccgg atctgcatcg caggatgctg ctggctaccc tgtggaacac
     3421 ctacatctgt attaacgaag cgctggcatt gaccctgagt gatttttctc tggtcccgcc
     3481 gcatccatac cgccagttgt ttaccctcac aacgttccag taaccgggca tgttcatcat
     3541 cagtaacccg tatcgtgagc atcctctctc gtttcatcgg tatcattacc cccatgaaca
     3601 gaaatccccc ttacacggag gcatcagtga ccaaacagga aaaaaccgcc cttaacatgg
     3661 cccgctttat cagaagccag acattaacgc ttctggagaa actcaacgag ctggacgcgg
     3721 atgaacaggc agacatctgt gaatcgcttc acgaccacgc tgatgagctt taccgcagct
     3781 gcctcgcgcg tttcggtgat gacggtgaaa acctctgaca catgcagctc ccggagacgg
     3841 tcacagcttg tctgtaagcg gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg
     3901 gtgttggcgg gtgtcggggc gcagccatga cccagtcacg tagcgatagc ggagtgtata
     3961 ctggcttaac tatgcggcat cagagcagat tgtactgaga gtgcaccata tatgcggtgt
     4021 gaaataccgc acagatgcgt aaggagaaaa taccgcatca ggcgctcttc cgcttcctcg
     4081 ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag
     4141 gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa
     4201 ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc
     4261 cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca
     4321 ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg
     4381 accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct
     4441 catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt
     4501 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag
     4561 tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc
     4621 agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac
     4681 actagaagga cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga
     4741 gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc
     4801 aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg
     4861 gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gaacaataaa
     4921 actgtctgct tacataaaca gtaatacaag gggtgttatg agccatattc aacgggaaac
     4981 gtcttgctct aggccgcgat taaattccaa catggatgct gatttatatg ggtataaatg
     5041 ggctcgcgat aatgtcgggc aatcaggtgc gacaatctat cgattgtatg ggaagcccga
     5101 tgcgccagag ttgtttctga aacatggcaa aggtagcgtt gccaatgatg ttacagatga
     5161 gatggtcaga ctaaactggc tgacggaatt tatgcctctt ccgaccatca agcattttat
     5221 ccgtactcct gatgatgcat ggttactcac cactgcgatc cccgggaaaa cagcattcca
     5281 ggtattagaa gaatatcctg attcaggtga aaatattgtt gatgcgctgg cagtgttcct
     5341 gcgccggttg cattcgattc ctgtttgtaa ttgtcctttt aacagcgatc gcgtatttcg
     5401 tctcgctcag gcgcaatcac gaatgaataa cggtttggtt gatgcgagtg attttgatga
     5461 cgagcgtaat ggctggcctg ttgaacaagt ctggaaagaa atgcataaac ttttgccatt
     5521 ctcaccggat tcagtcgtca ctcatggtga tttctcactt gataacctta tttttgacga
     5581 ggggaaatta ataggttgta ttgatgttgg acgagtcgga atcgcagacc gataccagga
     5641 tcttgccatc ctatggaact gcctcggtga gttttctcct tcattacaga aacggctttt
     5701 tcaaaaatat ggtattgata atcc
//
