LOCUS       sequence-435536-        8250 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  In mammalian cells expresses a Quartet of receptor-binding domains 
            (RBDs) from the sarbecoviruses SHC014, Rs4081, RaTG13, and 
            SARS-CoV-2 with no linkers between the RBDs and an N-terminal 
            SpyTag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8250)
  AUTHORS   Hills RA, Tan TK, Cohen AA, Keeffe JR, Keeble AH, Gnanapragasam PNP,
            Storm KN, Rorick AV, West AP Jr, Hill ML, Liu S, Gilbert-Jaramillo 
            J, Afzal M, Napier A, Admans G, James WS, Bjorkman PJ, Townsend AR, 
            Howarth MR
  TITLE     Proactive vaccination using multiviral Quartet Nanocages to elicit 
            broad anti-coronavirus responses.
  JOURNAL   Nat Nanotechnol. 2024 May 6. doi: 10.1038/s41565-024-01655-9.
  PUBMED    38710880
REFERENCE   2  (bases 1 to 8250)
  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 
            Nanotechnol. 2024 May 6. doi: 10.1038/s41565-024-01655-9."
COMMENT     Sequence Label: SpyTag-Quartet_NoLinker
FEATURES             Location/Qualifiers
     source          1..8250
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1010..1048
                     /codon_start=1
                     /product="peptide derived from the CnaB2 adhesin domain of 
                     Streptococcus pyogenes FbaB"
                     /label=SpyTag
                     /note="forms a covalent isopeptide bond with the SpyCatcher
                     protein (Zakeri et al., 2012)"
                     /translation="AHIVMVDAYKPTK"
     CDS             complement(3339..3356)
                     /codon_start=1
                     /product="tetracysteine peptide that binds biarsenical 
                     labeling reagents"
                     /label=tetracysteine tag
                     /translation="CCPGCC"
     primer_bind     complement(3841..3858)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3847..4071
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4117..4545
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4204..4223)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4414..4435
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4554..4574)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4559..4888
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4739..4874
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4801..4820
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4955..5749
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(5009..5028)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5619..5638
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5923..6044
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5960..5979)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6014..6033
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6093..6109)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6093..6109)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6106..6128)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6117..6133
                     /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)."
     promoter        complement(join(6141..6147,6148..6165,6166..6171))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6186..6207
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6324..6341)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6495..7083)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6575..6594)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7254..8045,8046..8114))
                     /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     7877..7896
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8115..8219)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gtttaaacgg gccctctaga ctcgagcggc cgccaccatg aacacccaga tcctggtgtt
      961 cgccctgatc gccatcattc ccaccaacgc cgacaagatc ggatctggcg cccacatcgt
     1021 gatggtggac gcctacaagc ctaccaaagg cagcggtgga agcggcggat ctggaaccgg
     1081 tagagtggcc cctagcaaag aagtcgtgcg gttccccaat atcaccaatc tgtgcccctt
     1141 cggcgaggtg ttcaacgcca ccacatttcc ctctgtgtac gcctgggaga gaaagcggat
     1201 cagcaactgc gtggccgact acagcgtgct gtacaacagc acctccttta gcaccttcaa
     1261 gtgctacggc gtgtccgcca ccaagctgaa cgatctgtgc ttctccaacg tgtacgccga
     1321 cagcttcgtg gtcaagggcg acgatgtgcg gcagattgct cctggacaga caggcgtgat
     1381 cgccgattac aactacaagc tgcctgacga cttcctgggc tgtgtgctgg cctggaacac
     1441 caatagcaag gacagctcca ccagcggcaa ctacaactac ctgtacagat gggtccgacg
     1501 cagcaagctg aatccctacg agagggacct gagcaacgac atctatagcc ctggcggcca
     1561 gagctgttct gccgtgggac ccaattgcta caaccctctg aggccctacg gcttctttac
     1621 cacagctggc gtgggccacc agccttacag agtggtggtg ctgtcattcg agctgctgaa
     1681 tgcccctgcc acagtgtgtg gccctaagct gtctaccgac ctgatcaaga accagcgggt
     1741 gtcccctaca cacgaggtcg tgcggttccc caatatcacc aacagatgcc ccttcgacaa
     1801 ggtgttcaac gccagcagat tccccaacgt gtacgcctgg gagcgcacca agatcagcga
     1861 ttgcgtggcc gattacaccg tgctgtacaa ctccaccagc tttagcacct ttaagtgcta
     1921 cggcgtgtcc ccaagcaagc tgatcgacct gtgcttcacc tctgtgtacg ccgacacctt
     1981 cctgatccgg tctagcgaag tgcgacaagt ggcccctggc gaaacaggcg tgatcgccga
     2041 ctacaactac aagctgcccg acgacttcac cggctgtgtg atcgcctgga ataccgccaa
     2101 gcaggaccag ggccagtact actacagaag cagcagaaag accaagctca agcccttcga
     2161 gcgggacctg acctccgatg agaatggcgt cagaaccctg tccacctacg atttctaccc
     2221 caatgtgccc atcgagtacc aggccaccag agtggtggtg ctgtctttcg aactgctgaa
     2281 cgcccctgcc acagtgtgtg gccctaaact gtctacagcc ctggtcaaga accagtgcgt
     2341 gaacttccgg gtgcagccca ccgatagcat cgtgcggttc cccaatatca ccaatctgtg
     2401 ccccttcggc gaggtgttca atgccaccac attcgcctct gtgtacgcct ggaaccggaa
     2461 gcggatcagc aattgcgtgg ccgactactc cgtgctgtac aacagcacca gcttcagcac
     2521 cttcaagtgc tacggcgtgt cccctaccaa gctgaacgac ctgtgcttca caaacgtgta
     2581 cgccgacagc ttcgtgatca ccggggatga agtgcggcag attgcccctg gacagacagg
     2641 caagatcgcc gactacaact acaagctgcc cgacgacttc accggctgtg tgattgcctg
     2701 gaactccaag cacatcgacg ccaaagaagg cgggaacttc aactacctgt accggctgtt
     2761 ccggaaggcc aacctgaagc ctttcgagcg ggatatctcc accgagatct accaggccgg
     2821 cagcaagcct tgtaatggcc agacaggcct gaactgctac tacccactgt acagatacgg
     2881 cttctacccc acagacggcg tgggccacca gccttataga gtggtggtgc tgagcttcga
     2941 gctgctgaat gcccctgcca cagtgtgcgg ccctaagaaa agcaccaatc tcgtgaagaa
     3001 caaatgcgtg aacttcaaca tcaccaatct gtgccccttc ggcgaggtgt tcaatgccac
     3061 aagatttgcc agcgtgtacg cctggaaccg gaagagaatc agcaactgcg tggccgacta
     3121 cagcgtgctg tacaatagcg ccagcttcag caccttcaag tgctacggcg tgtcccctac
     3181 caagctgaac gacctgtgct tcaccaatgt gtacgccgac agcttcgtga tcagaggcga
     3241 cgaagttcgg cagatcgctc ctggacagac aggcaagatc gccgattaca actacaagct
     3301 gcccgacgac ttcaccggct gcgtgatcgc ctggaatagc aacaacctgg acagcaaagt
     3361 cggcggcaac tacaactacc tgtaccggct gttccggaag tccaacctga agcctttcga
     3421 gcgggacatc agcaccgaga tctatcaggc cggcagcacc ccttgtaatg gcgtggaagg
     3481 cttcaactgc tacttcccac tgcagtccta cggcttccag cctacaaatg gcgtcggcta
     3541 ccagccttac agagtggtgg tgctgtcctt cgaactgctg catgcccctg ctaccgtgtg
     3601 cggccctaaa aaaacctagt agaccggtaa gctgagcagc ggctacaagg acgtgatcct
     3661 gtggttcagc ttcggcgcca gctgcttcat cctgctggcc atcgtgatgg gcctggtgtt
     3721 catctgcgtg aagaacggca acatgcggtg caccatctgc atctgatgat atcgaattcc
     3781 accacactgg actagtggat ccgagctcgg taccaagctt aagtttaaac cgctgatcag
     3841 cctcgactgt gccttctagt tgccagccat ctgttgtttg cccctccccc gtgccttcct
     3901 tgaccctgga aggtgccact cccactgtcc tttcctaata aaatgaggaa attgcatcgc
     3961 attgtctgag taggtgtcat tctattctgg ggggtggggt ggggcaggac agcaaggggg
     4021 aggattggga agacaatagc aggcatgctg gggatgcggt gggctctatg gcttctgagg
     4081 cggaaagaac cagctggggc tctagggggt atccccacgc gccctgtagc ggcgcattaa
     4141 gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc
     4201 ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag
     4261 ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca
     4321 aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc
     4381 gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa
     4441 cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct
     4501 attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattaattc tgtggaatgt
     4561 gtgtcagtta gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat
     4621 gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag
     4681 tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat
     4741 cccgccccta actccgccca gttccgccca ttctccgccc catggctgac taattttttt
     4801 tatttatgca gaggccgagg ccgcctctgc ctctgagcta ttccagaagt agtgaggagg
     4861 cttttttgga ggcctaggct tttgcaaaaa gctcccggga gcttgtatat ccattttcgg
     4921 atctgatcaa gagacaggat gaggatcgtt tcgcatgatt gaacaagatg gattgcacgc
     4981 aggttctccg gccgcttggg tggagaggct attcggctat gactgggcac aacagacaat
     5041 cggctgctct gatgccgccg tgttccggct gtcagcgcag gggcgcccgg ttctttttgt
     5101 caagaccgac ctgtccggtg ccctgaatga actgcaggac gaggcagcgc ggctatcgtg
     5161 gctggccacg acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag
     5221 ggactggctg ctattgggcg aagtgccggg gcaggatctc ctgtcatctc accttgctcc
     5281 tgccgagaaa gtatccatca tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc
     5341 tacctgccca ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga
     5401 agccggtctt gtcgatcagg atgatctgga cgaagagcat caggggctcg cgccagccga
     5461 actgttcgcc aggctcaagg cgcgcatgcc cgacggcgag gatctcgtcg tgacccatgg
     5521 cgatgcctgc ttgccgaata tcatggtgga aaatggccgc ttttctggat tcatcgactg
     5581 tggccggctg ggtgtggcgg accgctatca ggacatagcg ttggctaccc gtgatattgc
     5641 tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg ctttacggta tcgccgctcc
     5701 cgattcgcag cgcatcgcct tctatcgcct tcttgacgag ttcttctgag cgggactctg
     5761 gggttcgaaa tgaccgacca agcgacgccc aacctgccat cacgagattt cgattccacc
     5821 gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc
     5881 ctccagcgcg gggatctcat gctggagttc ttcgcccacc ccaacttgtt tattgcagct
     5941 tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc atttttttca
     6001 ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctgtataccg
     6061 tcgacctcta gctagagctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt
     6121 tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt
     6181 gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg
     6241 ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg
     6301 cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg
     6361 cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat
     6421 aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc
     6481 gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc
     6541 tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga
     6601 agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt
     6661 ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg
     6721 taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc
     6781 gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg
     6841 gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc
     6901 ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg
     6961 ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc
     7021 gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct
     7081 caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt
     7141 taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa
     7201 aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa
     7261 tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc
     7321 tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct
     7381 gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca
     7441 gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt
     7501 aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt
     7561 gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc
     7621 ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc
     7681 tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt
     7741 atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact
     7801 ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc
     7861 ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt
     7921 ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg
     7981 atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct
     8041 gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa
     8101 tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt
     8161 ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc
     8221 acatttcccc gaaaagtgcc acctgacgtc
//
