LOCUS       sequence-331342-        5157 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of Nsp1 residues 13-127.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5157)
  AUTHORS   Semper C, Watanabe N, Savchenko A
  TITLE     Structural characterization of nonstructural protein 1 from 
            SARS-CoV-2.
  JOURNAL   iScience. 2021 Jan 22;24(1):101903. doi: 10.1016/j.isci.2020.101903.
            Epub 2020 Dec 7.
  PUBMED    33319167
REFERENCE   2  (bases 1 to 5157)
  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: "iScience.";
            date: "2021-01-22"; volume: "24(1):101903. doi"; pages: " 
            10.1016/j.isci.2020.101903. Epub 2020 Dec 7"
COMMENT     Sequence Label: pMCSG53_Nsp1
FEATURES             Location/Qualifiers
     source          1..5157
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     970..989
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(1009..1086)
                     /gene="lacI"
                     /label=lacI promoter
     tRNA            1226..1301
                     /gene="ileX"
                     /product="E. coli tRNA-Ile recognizing AUA"
                     /label=ileX
     primer_bind     1450..1469
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     tRNA            1571..1647
                     /gene="argU"
                     /product="E. coli tRNA-Arg recognizing AGA and AGG"
                     /label=argU
     primer_bind     1922..1941
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1922..1940
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1941..1965
                     /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)."
     RBS             1980..2002
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             2013..2030
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             2055..2075
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             2493..2510
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2563..2581)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2577..2624
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     complement(2748..2767)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2934..3037
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3038..3106,3107..3898)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(3256..3275)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4069..4657
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4558..4577
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4811..4828
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(4908..4927)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     CDS             complement(join(5083..5157,1..1008))
                     /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 ggtttgcgta ttgggcgcca gggtggtttt tcttttcacc agtgagacgg gcaacagctg
       61 attgcccttc accgcctggc cctgagagag ttgcagcaag cggtccacgc tggtttgccc
      121 cagcaggcga aaatcctgtt tgatggtggt taacggcggg atataacatg agctgtcttc
      181 ggtatcgtcg tatcccacta ccgagatatc cgcaccaacg cgcagcccgg actcggtaat
      241 ggcgcgcatt gcgcccagcg ccatctgatc gttggcaacc agcatcgcag tgggaacgat
      301 gccctcattc agcatttgca tggtttgttg aaaaccggac atggcactcc agtcgccttc
      361 ccgttccgct atcggctgaa tttgattgcg agtgagatat ttatgccagc cagccagacg
      421 cagacgcgcc gagacagaac ttaatgggcc cgctaacagc gcgatttgct ggtgacccaa
      481 tgcgaccaga tgctccacgc ccagtcgcgt accgtcttca tgggagaaaa taatactgtt
      541 gatgggtgtc tggtcagaga catcaagaaa taacgccgga acattagtgc aggcagcttc
      601 cacagcaatg gcatcctggt catccagcgg atagttaatg atcagcccac tgacgcgttg
      661 cgcgagaaga ttgtgcaccg ccgctttaca ggcttcgacg ccgcttcgtt ctaccatcga
      721 caccaccacg ctggcaccca gttgatcggc gcgagattta atcgccgcga caatttgcga
      781 cggcgcgtgc agggccagac tggaggtggc aacgccaatc agcaacgact gtttgcccgc
      841 cagttgttgt gccacgcggt tgggaatgta attcagctcc gccatcgccg cttccacttt
      901 ttcccgcgtt ttcgcagaaa cgtggctggc ctggttcacc acgcgggaaa cggtctgata
      961 agagacaccg gcatactctg cgacatcgta taacgttact ggtttcacat tcaccaccct
     1021 gaattgactc tcttccgggc gctatcatgc cataccgcga aaggttttgc gccattcgat
     1081 ggtgtccggg atctcgacgc tctcccttat gcgactcctg cattaggaag cagcccagta
     1141 gtaggttgag gccgttgagc accgccgccg caaggaatgg tgcatgcgga ttgcgacacg
     1201 gagttacttt ataatccgct accatggccc cttagctcag tggttagagc aggcgactca
     1261 taatcgcttg gtcgctggtt caagtccagc aggggccacc agatatagca aaggctgacg
     1321 agaaatcgtc agcctttttg catgcaagga gatggcgccc aacagtcccc cggccacggg
     1381 gcctgccacc atacccacgc cgaaacaagc gctcatgagc ccgaagtggc gagcccgatc
     1441 ttccccatcg gtgatgtcgg cgatataggc gccagcaacc gcacctgtgg cgccggcgag
     1501 atcaaaagcc attgactcag caagggttga ccgtataatt cacgcgatta caccgcattg
     1561 cggtatcaac gcgcccttag ctcagttgga tagagcaacg accttctaag tcgtgggccg
     1621 caggttcgaa tcctgcaggg cgcgccatta caattcaatc agttacgcct tctttatatc
     1681 ctccataatt ccagagtggg acatatttgg gacattatca ccaaaaatgt cgtctatttt
     1741 cctcgcatgc tctgtcaaat gattaggcgc aaggtgagca tacctacgaa ccatttctat
     1801 ggactcccat ccgcccattt cctgaagcac tgataatggg acgcctgact gaatccaccg
     1861 gtgatgccgg ccacgatgcg tccggcgtag aggatcgaga tcgatctcga tcccgcgaaa
     1921 ttaatacgac tcactatagg ggaattgtga gcggataaca attcccctct agaaataatt
     1981 ttgtttaact ttaagaagga gatatacata tgcaccatca tcatcatcat tcttctggtg
     2041 tagatctggg taccgagaac ctgtacttcc aatccaatgc ccatgttcag ctgagtttac
     2101 cggttctgca agttcgtgat gttctggttc gcggttttgg tgatagcgtt gaagaagttc
     2161 tgagtgaagc acgtcaacac ctgaaagatg gtacatgcgg cttagtggaa gtggaaaaag
     2221 gtgttttacc tcagctggaa cagccgtacg tgttcattaa acgcagcgat gcaagaacag
     2281 cacctcatgg tcatgttatg gttgaactgg tggcagaact ggaaggtatc cagtatggta
     2341 gatctggtga aacactgggt gttttagttc cgcatgtggg cgaaattcct gtggcatacc
     2401 gtaaagtgct gctgcgtaaa aatggctaac attggaagtg gataacggat ccgaattcga
     2461 gctccgtcga caagcttgcg gccgcactcg agcaccacca ccaccaccac tgagatccgg
     2521 ctgctaacaa agcccgaaag gaagctgagt tggctgctgc caccgctgag caataactag
     2581 cataacccct tggggcctct aaacgggtct tgaggggttt tttgctgaaa ggaggaacta
     2641 tatccggatt ggcgaatggg acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt
     2701 ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt
     2761 cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct
     2821 ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg
     2881 tgatggttca cattaacgct tacaatttag gtggcacttt tcggggaaat gtgcgcggaa
     2941 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac
     3001 cctgataaat gcttcaataa tttgaaaaag gaagagtatg agtattcaac atttccgtgt
     3061 cgcccttatt cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct
     3121 ggtgaaagta aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga
     3181 tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag
     3241 cacttttaaa gttctgctat gtggcgcggt attatcccgt attgacgccg ggcaagagca
     3301 actcggtcgc cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga
     3361 aaagcatctt acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag
     3421 tgataacact gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc
     3481 ttttttgcac aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa
     3541 tgaagccata ccaaacgacg agcgtgacac cacgatgcct gcagcaatgg caacaacgtt
     3601 gcgcaaacta ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg
     3661 gatggaggcg gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt
     3721 tattgctgat aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg
     3781 gccagatggt aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat
     3841 ggatgaacga aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact
     3901 gtcagaccaa gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa
     3961 aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt
     4021 ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt
     4081 ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg
     4141 tttgccggat caagagctac caactctttt tccgaaggta actggcttca gcagagcgca
     4201 gataccaaat actgtccttc tagtgtagcc gtagttaggc caccacttca agaactctgt
     4261 agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga
     4321 taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc
     4381 gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact
     4441 gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga
     4501 caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg
     4561 aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt
     4621 tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt
     4681 acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga
     4741 ttctgtggat aaccgtatta ccgcctttga gtgagctgat accgctcgcc gcagccgaac
     4801 gaccgagcgc agcgagtcag tgagcgagga agcggaagag cgcctgatgc ggtattttct
     4861 ccttacgcat ctgtgcggta tttcacaccg caatggtgca ctctcagtac aatctgctct
     4921 gatgccgcat agttaagcca gtatacactc cgctatcgct acgtgactgt agtcatgccc
     4981 cgcgcccacc ggaaggagct gactgggttg aaggctctca agggcatcgg tcgagatccc
     5041 ggtgcctaat gagtgagcta acttacatta attgcgttgc gctcactgcc cgctttccag
     5101 tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggc
//
