LOCUS       sequence-435532-        8331 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 each separated by GS-linkers with an N-terminal SpyTag 
            fusion.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8331)
  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 8331)
  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
FEATURES             Location/Qualifiers
     source          1..8331
                     /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(3420..3437)
                     /codon_start=1
                     /product="tetracysteine peptide that binds biarsenical 
                     labeling reagents"
                     /label=tetracysteine tag
                     /translation="CCPGCC"
     primer_bind     complement(3922..3939)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3928..4152
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4198..4626
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4285..4304)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4495..4516
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4635..4655)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4640..4969
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4820..4955
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4882..4901
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             5036..5830
                     /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(5090..5109)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5700..5719
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    6004..6125
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6041..6060)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6095..6114
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6174..6190)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6174..6190)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6187..6209)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6198..6214
                     /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(6222..6228,6229..6246,6247..6252))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6267..6288
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6405..6422)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6576..7164)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6656..6675)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7335..8126,8127..8195))
                     /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     7958..7977
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8196..8300)
                     /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 accaggggag
     1741 tggtgggtcc gggggttcgg gacgggtgtc ccctacacac gaggtcgtgc ggttccccaa
     1801 tatcaccaac agatgcccct tcgacaaggt gttcaacgcc agcagattcc ccaacgtgta
     1861 cgcctgggag cgcaccaaga tcagcgattg cgtggccgat tacaccgtgc tgtacaactc
     1921 caccagcttt agcaccttta agtgctacgg cgtgtcccca agcaagctga tcgacctgtg
     1981 cttcacctct gtgtacgccg acaccttcct gatccggtct agcgaagtgc gacaagtggc
     2041 ccctggcgaa acaggcgtga tcgccgacta caactacaag ctgcccgacg acttcaccgg
     2101 ctgtgtgatc gcctggaata ccgccaagca ggaccagggc cagtactact acagaagcag
     2161 cagaaagacc aagctcaagc ccttcgagcg ggacctgacc tccgatgaga atggcgtcag
     2221 aaccctgtcc acctacgatt tctaccccaa tgtgcccatc gagtaccagg ccaccagagt
     2281 ggtggtgctg tctttcgaac tgctgaacgc ccctgccaca gtgtgtggcc ctaaactgtc
     2341 tacagccctg gtcaagaacc agtgcgtgaa cttcggcagc ggcggaacgg gaggtagtgg
     2401 ccgggtgcag cccaccgata gcatcgtgcg gttccccaat atcaccaatc tgtgcccctt
     2461 cggcgaggtg ttcaatgcca ccacattcgc ctctgtgtac gcctggaacc ggaagcggat
     2521 cagcaattgc gtggccgact actccgtgct gtacaacagc accagcttca gcaccttcaa
     2581 gtgctacggc gtgtccccta ccaagctgaa cgacctgtgc ttcacaaacg tgtacgccga
     2641 cagcttcgtg atcaccgggg atgaagtgcg gcagattgcc cctggacaga caggcaagat
     2701 cgccgactac aactacaagc tgcccgacga cttcaccggc tgtgtgattg cctggaactc
     2761 caagcacatc gacgccaaag aaggcgggaa cttcaactac ctgtaccggc tgttccggaa
     2821 ggccaacctg aagcctttcg agcgggatat ctccaccgag atctaccagg ccggcagcaa
     2881 gccttgtaat ggccagacag gcctgaactg ctactaccca ctgtacagat acggcttcta
     2941 ccccacagac ggcgtgggcc accagcctta tagagtggtg gtgctgagct tcgagctgct
     3001 gaatgcccct gccacagtgt gcggccctaa gaaaagcacc aatctcgtga agaacaaatg
     3061 cgtgaacttc ggtacgggtg gcagtggagg gagcgggaac atcaccaatc tgtgcccctt
     3121 cggcgaggtg ttcaatgcca caagatttgc cagcgtgtac gcctggaacc ggaagagaat
     3181 cagcaactgc gtggccgact acagcgtgct gtacaatagc gccagcttca gcaccttcaa
     3241 gtgctacggc gtgtccccta ccaagctgaa cgacctgtgc ttcaccaatg tgtacgccga
     3301 cagcttcgtg atcagaggcg acgaagttcg gcagatcgct cctggacaga caggcaagat
     3361 cgccgattac aactacaagc tgcccgacga cttcaccggc tgcgtgatcg cctggaatag
     3421 caacaacctg gacagcaaag tcggcggcaa ctacaactac ctgtaccggc tgttccggaa
     3481 gtccaacctg aagcctttcg agcgggacat cagcaccgag atctatcagg ccggcagcac
     3541 cccttgtaat ggcgtggaag gcttcaactg ctacttccca ctgcagtcct acggcttcca
     3601 gcctacaaat ggcgtcggct accagcctta cagagtggtg gtgctgtcct tcgaactgct
     3661 gcatgcccct gctaccgtgt gcggccctaa aaaaacctag tagaccggta agctgagcag
     3721 cggctacaag gacgtgatcc tgtggttcag cttcggcgcc agctgcttca tcctgctggc
     3781 catcgtgatg ggcctggtgt tcatctgcgt gaagaacggc aacatgcggt gcaccatctg
     3841 catctgatga tatcgaattc caccacactg gactagtgga tccgagctcg gtaccaagct
     3901 taagtttaaa ccgctgatca gcctcgactg tgccttctag ttgccagcca tctgttgttt
     3961 gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc ctttcctaat
     4021 aaaatgagga aattgcatcg cattgtctga gtaggtgtca ttctattctg gggggtgggg
     4081 tggggcagga cagcaagggg gaggattggg aagacaatag caggcatgct ggggatgcgg
     4141 tgggctctat ggcttctgag gcggaaagaa ccagctgggg ctctaggggg tatccccacg
     4201 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
     4261 cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
     4321 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
     4381 ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat
     4441 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
     4501 tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag
     4561 ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     4621 cgaattaatt ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc
     4681 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc
     4741 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt
     4801 cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc
     4861 ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctctg cctctgagct
     4921 attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa agctcccggg
     4981 agcttgtata tccattttcg gatctgatca agagacagga tgaggatcgt ttcgcatgat
     5041 tgaacaagat ggattgcacg caggttctcc ggccgcttgg gtggagaggc tattcggcta
     5101 tgactgggca caacagacaa tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca
     5161 ggggcgcccg gttctttttg tcaagaccga cctgtccggt gccctgaatg aactgcagga
     5221 cgaggcagcg cggctatcgt ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga
     5281 cgttgtcact gaagcgggaa gggactggct gctattgggc gaagtgccgg ggcaggatct
     5341 cctgtcatct caccttgctc ctgccgagaa agtatccatc atggctgatg caatgcggcg
     5401 gctgcatacg cttgatccgg ctacctgccc attcgaccac caagcgaaac atcgcatcga
     5461 gcgagcacgt actcggatgg aagccggtct tgtcgatcag gatgatctgg acgaagagca
     5521 tcaggggctc gcgccagccg aactgttcgc caggctcaag gcgcgcatgc ccgacggcga
     5581 ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat atcatggtgg aaaatggccg
     5641 cttttctgga ttcatcgact gtggccggct gggtgtggcg gaccgctatc aggacatagc
     5701 gttggctacc cgtgatattg ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt
     5761 gctttacggt atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga
     5821 gttcttctga gcgggactct ggggttcgaa atgaccgacc aagcgacgcc caacctgcca
     5881 tcacgagatt tcgattccac cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc
     5941 cgggacgccg gctggatgat cctccagcgc ggggatctca tgctggagtt cttcgcccac
     6001 cccaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc
     6061 acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta
     6121 tcttatcatg tctgtatacc gtcgacctct agctagagct tggcgtaatc atggtcatag
     6181 ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc
     6241 ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc
     6301 tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa
     6361 cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg
     6421 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
     6481 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
     6541 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     6601 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     6661 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     6721 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     6781 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     6841 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     6901 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     6961 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
     7021 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     7081 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     7141 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     7201 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     7261 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     7321 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     7381 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     7441 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
     7501 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     7561 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     7621 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
     7681 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     7741 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     7801 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     7861 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     7921 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
     7981 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     8041 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     8101 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     8161 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     8221 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
     8281 aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt c
//
