LOCUS       sequence-382967-        5182 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Production of SARS-CoV2 receptor binding domain in E. coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5182)
  AUTHORS   Gaynor KU, Vaysburd M, Harman MAJ, Albecka A, Jeffrey P, Beswick P, 
            Papa G, Chen L, Mallery D, McGuinness B, Van Rietschoten K, Stanway 
            S, Brear P, Lulla A, Ciazynska K, Chang VT, Sharp J, Neary M, Box H,
            Herriott J, Kijak E, Tatham L, Bentley EG, Sharma P, Kirby A, Han X,
            Stewart JP, Owen A, Briggs JAG, Hyvonen M, Skynner MJ, James LC
  TITLE     Multivalent bicyclic peptides are an effective antiviral modality 
            that can potently inhibit SARS-CoV-2.
  JOURNAL   Nat Commun. 2023 Jun 16;14(1):3583. doi: 10.1038/s41467-023-39158-1.
  PUBMED    37328472
REFERENCE   2  (bases 1 to 5182)
  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: "Nat 
            Commun."; date: "2023-06-16"; volume: "14(1):3583. doi"; pages: " 
            10.1038/s41467-023-39158-1"
COMMENT     Sequence Label: pExp-His-zBasic-RBD
FEATURES             Location/Qualifiers
     source          1..5182
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     14..33
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        14..32
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    32..56
                     /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             69..91
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             105..128
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     CDS             144..314
                     /codon_start=1
                     /product="Z domain of staphylococcal protein A, engineered 
                     to contain multiple positive charges (Hedhammar and Hober, 
                     2007)"
                     /label=Z-basic tag
                     /note="useful for purifying tagged proteins by 
                     cation-exchange chromatography"
                     /translation="VDNKFNKERRRARREIRHLPNLNREQRRAFIRSLRDDPSQSANLL
                     AEAKKLNDAQPK"
     CDS             324..344
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     promoter        complement(949..967)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(950..967)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(1025..1043)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1039..1086
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     complement(1322..1343)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1493..2575)
                     /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     2537..2556
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(2576..2653)
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     primer_bind     2792..2811
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(2965..2984)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3084..3106
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3144..3162)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3230..3334
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3335..3403,3404..4195)
                     /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(3553..3572)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4366..4954
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4855..4874
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5108..5125
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 taatgcagct gattaatacg actcactata gggaattgtg agcggataac aattcctcta
       61 gaaataattt tgtttaactt taagaaggag atatatccat gaatcaccat caccatcacc
      121 atcaccattc tggcactagt ggcgttgaca acaaattcaa caaagaacgt cgccgtgcgc
      181 gtcgcgaaat ccgtcacctg ccgaacctca accgtgaaca gcgtcgtgcg ttcatccgtt
      241 ctctgcgtga cgacccgtct cagtctgcga acctgctggc ggaagcgaag aaactgaacg
      301 acgcacaacc gaaaagcggt accgaaaacc tgtacttcca gggttctggt actaacctgt
      361 gcccgttcgg tgaagttttc aacgcgaccc gtttcgcgtc cgtctacgcg tggaaccgta
      421 aacgtatctc taactgcgtt gcggactact ctgtcctgta caactctgcg tctttctcta
      481 cctttaaatg ctacggtgtt tctccgacca aactgaacga cctctgcttt accaacgttt
      541 atgccgactc tttcgttatc cgtggtgacg aagttcgtca gatcgcgcca ggtcaaaccg
      601 gcaaaatcgc ggattacaat tacaaactgc cggacgattt caccggttgt gttattgctt
      661 ggaactctaa caacctggac tctaaagttg gtggtaacta caactacctg tatcgtctgt
      721 tccgtaaatc taacctcaaa ccatttgaac gtgacatcag caccgaaatc taccaggcgg
      781 gttctacccc gtgcaacggt gtcgagggtt tcaactgcta cttcccgctg cagtcttatg
      841 gtttccagcc tactaacggc gttggttacc agccgtaccg tgttgttgtt ctgtctttcg
      901 aactgctgca cgctcctgct accgtttgtg gtccgtaagc ttgagtattc tatagtgtca
      961 cctaaatccc agcttgatcc ggctgctaac aaagcccgaa aggaagctga gttggctgct
     1021 gccaccgctg agcaataact agcataaccc cttggggcct ctaaacgggt cttgaggggt
     1081 tttttgctga aaggaggaac tatatccgga taacctggcg taatagcgaa gaggcccgca
     1141 ccgatcgccc ttcccaacag ttgcgcagcc tgaatggcga atggaaattg taagcgttaa
     1201 tattttgtta aaattcgcgt taaatttttg ttaaatcagc tcatttttta accaataggc
     1261 cgaaatcggc aaaatccctt ataaatcaaa agaatagacc gagatagggt tgagtgttgt
     1321 tccagtttgg aacaagagtc cactattaaa gaacgtggac tccaacgtca aagggcgaaa
     1381 aaccgtctat cagggcgatg gcccacagct tgcatgcctg caggtcggaa gcataaagtg
     1441 taaagcctgg ggtgcctaat gagtgagcta acttacatta attgcgttgc gctcactgcc
     1501 cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg
     1561 gagaggcggt ttgcgtattg ggcgccaggg tggtttttct tttcaccagt gagacgggca
     1621 acagctgatt gcccttcacc gcctggccct gagagagttg cagcaagcgg tccacgctgg
     1681 tttgccccag caggcgaaaa tcctgtttga tggtggttaa cggcgggata taacatgagc
     1741 tgtcttcggt atcgtcgtat cccactaccg agatatccgc accaacgcgc agcccggact
     1801 cggtaatggc gcgcattgcg cccagcgcca tctgatcgtt ggcaaccagc atcgcagtgg
     1861 gaacgatgcc ctcattcagc atttgcatgg tttgttgaaa accggacatg gcactccagt
     1921 cgccttcccg ttccgctatc ggctgaattt gattgcgagt gagatattta tgccagccag
     1981 ccagacgcag acgcgccgag acagaactta atgggcccgc taacagcgcg atttgctggt
     2041 gacccaatgc gaccagatgc tccacgccca gtcgcgtacc gtcttcatgg gagaaaataa
     2101 tactgttgat gggtgtctgg tcagagacat caagaaataa cgccggaaca ttagtgcagg
     2161 cagcttccac agcaatggca tcctggtcat ccagcggata gttaatgatc agcccactga
     2221 cgcgttgcgc gagaagattg tgcaccgccg ctttacaggc ttcgacgccg cttcgttcta
     2281 ccatcgacac caccacgctg gcacccagtt gatcggcgcg agatttaatc gccgcgacaa
     2341 tttgcgacgg cgcgtgcagg gccagactgg aggtggcaac gccaatcagc aacgactgtt
     2401 tgcccgccag ttgttgtgcc acgcggttgg gaatgtaatt cagctccgcc atcgccgctt
     2461 ccactttttc ccgcgttttc gcagaaacgt ggctggcctg gttcaccacg cgggaaacgg
     2521 tctgataaga gacaccggca tactctgcga catcgtataa cgttactggt ttcacattca
     2581 ccaccctgaa ttgactctct tccgggcgct atcatgccat accgcgaaag gttttgcacc
     2641 attcgatggt gtcggcgagc tcgaattgtg aaccatcacc ctaatcaagt tttttggggt
     2701 cgaggtgccg taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac
     2761 ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta
     2821 gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg
     2881 cgccgctaca gggcgcgtcc tgatgcggta ttttctcctt acgcatctgt gcggtatttc
     2941 acaccgcata tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagcc
     3001 ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc
     3061 ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt caccgtcatc
     3121 accgaaacgc gcgagacgaa agggcctcgt gatacgccta tttttatagg ttaatgtcat
     3181 gataataatg gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc
     3241 tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg
     3301 ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc
     3361 ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt
     3421 gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct
     3481 caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac
     3541 ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact
     3601 cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa
     3661 gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga
     3721 taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt
     3781 tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga
     3841 agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg
     3901 caaactatta actggcgaac tacttactct agcttcccgg caacaattaa tagactggat
     3961 ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat
     4021 tgctgataaa tctggagccg gtgagcgtgg ttctcgcggt atcattgcag cactggggcc
     4081 agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga
     4141 tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc
     4201 agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag
     4261 gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc
     4321 gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt
     4381 tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt
     4441 gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat
     4501 accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc
     4561 accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa
     4621 gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg
     4681 ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag
     4741 atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag
     4801 gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa
     4861 cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt
     4921 gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg
     4981 gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc
     5041 tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac
     5101 cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct
     5161 ccccgcgcgt tggccgattc at
//
