LOCUS       sequence-362477-        6775 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  yeast surface display of the SARS-CoV-2 Eta variant RBD.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6775)
  AUTHORS   Starr TN, Greaney AJ, Hannon WW, Loes AN, Hauser K, Dillen JR, Ferri
            E, Farrell AG, Dadonaite B, McCallum M, Matreyek KA, Corti D, 
            Veesler D, Snell G, Bloom JD
  TITLE     Shifting mutational constraints in the SARS-CoV-2 receptor-binding 
            domain during viral evolution.
  JOURNAL   Science. 2022 Jun 28:eabo7896. doi: 10.1126/science.abo7896.
  PUBMED    35762884
REFERENCE   2  (bases 1 to 6775)
  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: "Science. 
            2022 Jun 28:eabo7896. doi: 10.1126/science.abo7896."
COMMENT     Sequence Label: 3019_pETcon-SARS-CoV-2-RBD_E484K
FEATURES             Location/Qualifiers
     source          1..6775
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    21..42
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(57..62,63..80,81..87)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    95..111
                     /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     100..122
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     119..135
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     119..135
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        156..174
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        204..856
                     /gene="GAL1 and GAL10 genes of S. cerevisiae"
                     /label=GAL1,10 promoter
                     /note="divergent inducible promoter, regulated by Gal4"
     protein_bind    408..525
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     primer_bind     820..843
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     CDS             1175..1186
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             1187..1213
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1904..1933
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     primer_bind     complement(2262..2281)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2263..2281)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     rep_origin      2472..2927
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2559..2578)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2769..2790
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(3030..3704)
                     /codon_start=1
                     /gene="S. cerevisiae TRP1"
                     /product="phosphoribosylanthranilate isomerase, required 
                     for tryptophan biosynthesis"
                     /label=TRP1
                     /note="yeast auxotrophic marker"
                     /translation="MSVINFTGSSGPLVKVCGLQSTEAAECALDSDADLLGIICVPNRK
                     RTIDPVIARKISSLVKAYKNSSGTPKYLVGVFRNQPKEDVLALVNDYGIDIVQLHGDES
                     WQEYQEFLGLPVIKRLVFPKDCNILLSAASQKPHSFIPLFDSEAGGTGELLDWNSISDW
                     VGRQESPESLHFMLAGGLTPENVGDALRLNGVIGVDVSGGVETNGVKDSNKIANFVKNA
                     KK"
     promoter        complement(3705..3985)
                     /gene="S. cerevisiae TRP1"
                     /label=TRP1 promoter
     primer_bind     complement(4006..4025)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4125..4147
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4185..4203)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     misc_feature    4244..4747
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     promoter        4784..4888
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4889..4957,4958..5749)
                     /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(5107..5126)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5920..6508
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6409..6428
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6662..6679
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 ggcagtgagc gcaacgcaat taatgtgagt tagctcactc attaggcacc ccaggcttta
       61 cactttatgc ttccggctcg tatgttgtgt ggaattgtga gcggataaca atttcacaca
      121 ggaaacagct atgaccatga ttacgccaag ctcgaaatta accctcacta aagggaacaa
      181 aagctggtac caattccttg aattttcaaa aattcttact ttttttttgg atggacgcaa
      241 agaagtttaa taatcatatt acatggcatt accaccatat acatatccat atctaatctt
      301 acttatatgt tgtggaaatg taaagagccc cattatctta gcctaaaaaa accttctctt
      361 tggaactttc agtaatacgc ttaactgctc attgctatat tgaagtacgg attagaagcc
      421 gccgagcggg tgacagccct ccgaaggaag actctcctcc gtgcgtcctc gtcttcaccg
      481 gtcgcgttcc tgaaacgcag atgtgcctcg cgccgcactg ctccgaacaa taaagattct
      541 acaatactag cttttatggt tatgaagagg aaaaattggc agtaacctgg ccccacaaac
      601 cttcaaatga acgaatcaaa ttaacaacca taggatgata atgcgattag ttttttagcc
      661 ttatttctgg ggtaattaat cagcgaagcg atgatttttg atctattaac agatatataa
      721 atgcaaaaac tgcataacca ctttaactaa tactttcaac attttcggtt tgtattactt
      781 cttattcaaa tgtaataaaa gtatcaacaa aaaattgtta atatacctct atactttaac
      841 gtcaaggaga aaaaaccccg gatcgaattc cctacttcat acattttcaa ttaagatgca
      901 gttacttcgc tgtttttcaa tattttctgt tattgcttca gttttagcac aggaactgac
      961 aactatatgc gagcaaatcc cctcaccaac tttagaatcg acgccgtact ctttgtcaac
     1021 gactactatt ttggccaacg ggaaggcaat gcaaggagtt tttgaatatt acaaatcagt
     1081 aacgtttgtc agtaattgcg gttctcaccc ctcaacaact agcaaaggca gccccataaa
     1141 cacacagtat gtttttaagg acaatagctc gacgattgaa ggtagatacc catacgacgt
     1201 tccagactac gctctgcagg ctagtggtgg aggaggctct ggtggaggcg gccgcggagg
     1261 cggagggtcg gctagccata tgaatatcac gaacctttgt cctttcggtg aggtcttcaa
     1321 tgctactaga ttcgcatccg tgtatgcatg gaatagaaag agaattagta attgtgtagc
     1381 ggactactct gtactttata actccgcctc cttctccaca ttcaagtgtt acggtgtatc
     1441 tcccaccaag ttgaatgatc tatgctttac aaacgtttac gccgatagtt tcgtaattag
     1501 aggcgatgaa gtgcgtcaga tcgcaccagg ccagacgggc aagatagcag actataatta
     1561 taagctgcct gatgacttca ccggctgtgt gatagcttgg aactcaaata atctagattc
     1621 caaggtggga ggcaattaca attatttgta ccgtctgttc cgtaaaagca atttgaaacc
     1681 atttgaaaga gacattagca ctgaaattta tcaagcaggg tccaccccgt gcaacggcgt
     1741 aaagggcttt aactgttatt tcccattaca gtcttatggt ttccaaccta cgaacggagt
     1801 cgggtatcag ccgtacaggg ttgtggttct ttcatttgaa ctgctgcacg cgcccgcaac
     1861 cgtatgcggg ccgaagaaat caacgctcga ggggggcggt tccgaacaaa agcttatttc
     1921 tgaagaggac ttgtaataga gatctgataa caacagtgta gatgtaacaa aatcgacttt
     1981 gttcccactg tacttttagc tcgtacaaaa tacaatatac ttttcatttc tccgtaaaca
     2041 acatgttttc ccatgtaata tccttttcta tttttcgttc cgttaccaac tttacacata
     2101 ctttatatag ctattcactt ctatacacta aaaaactaag acaattttaa ttttgctgcc
     2161 tgccatattt caatttgtta taaattccta taatttatcc tattagtagc taaaaaaaga
     2221 tgaatgtgaa tcgaatccta agagaattga gctccaattc gccctatagt gagtcgtatt
     2281 acaattcact ggccgtcgtt agatcggaag agcacaactc cagtcactta caacgtcgtg
     2341 actgggaaaa ccctggcgtt acccaactta atcgccttgc agcacatccc cctttcgcca
     2401 gctggcgtaa tagcgaagag gcccgcaccg atcgcctttc ccaacagttg cgcagcctga
     2461 atggcgaatg gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg
     2521 cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc
     2581 ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg
     2641 gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc
     2701 acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt
     2761 ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc
     2821 ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta
     2881 acaaaaattt aacgcgaatt ttaacaaaat attaacgctt acaatttcct gatgcggtat
     2941 tttctcctta cgcatctgtg cggtatttca caccgcatag atcggcaagt gcacaaacaa
     3001 tacttaaata aatactactc agtaataacc tatttcttag catttttgac gaaatttgct
     3061 attttgttag agtcttttac accatttgtc tccacacctc cgcttacatc aacaccaata
     3121 acgccattta atctaagcgc atcaccaaca ttttctggcg tcagtccacc agctaacata
     3181 aaatgtaagc tttcggggct ctcttgcctt ccaacccagt cagaaatcga gttccaatcc
     3241 aaaagttcac ctgtcccacc tgcttctgaa tcaaacaagg gaataaacga atgaggtttc
     3301 tgtgaagctg cactgagtag tatgttgcag tcttttggaa atacgagtct tttaataact
     3361 ggcaaaccga ggaactcttg gtattcttgc cacgactcat ctccatgcag ttggacgata
     3421 tcaatgccgt aatcattgac cagagccaaa acatcctcct taggttgatt acgaaacacg
     3481 ccaaccaagt atttcggagt gcctgaacta tttttatatg cttttacaag acttgaaatt
     3541 ttccttgcaa taaccgggtc aattgttctc tttctattgg gcacacatat aatacccagc
     3601 aagtcagcat cggaatctag agcacattct gcggcctctg tgctctgcaa gccgcaaact
     3661 ttcaccaatg gaccagaact acctgtgaaa ttaataacag acatactcca agctgccttt
     3721 gtgtgcttaa tcacgtatac tcacgtgctc aatagtcacc aatgccctcc ctcttggccc
     3781 tctccttttc ttttttcgac cgaattaatt cttaatcggc aaaaaaagaa aagctccgga
     3841 tcaagattgt acgtaaggtg acaagctatt tttcaataaa gaatatcttc cactactgcc
     3901 atctggcgtc ataactgcaa agtacacata tattacgatg ctgttctatt aaatgcttcc
     3961 tatattatat atatagtaat gtcgtgatct atggtgcact ctcagtacaa tctgctctga
     4021 tgccgcatag ttaagccagc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc
     4081 ttgtctgctc ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg
     4141 tcagaggttt tcaccgtcat caccgaaacg cgcgagacga aagggcctcg tgatacgcct
     4201 atttttatag gttaatgtca tgataataat ggtttcttag acggatcgct tgcctgtaac
     4261 ttacacgcgc ctcgtatctt ttaatgatgg aataatttgg gaatttactc tgtgtttatt
     4321 tatttttatg ttttgtattt ggattttaga aagtaaataa agaaggtaga agagttacgg
     4381 aatgaagaaa aaaaaataaa caaaggttta aaaaatttca acaaaaagcg tactttacat
     4441 atatatttat tagacaagaa aagcagatta aatagatata cattcgatta acgataagta
     4501 aaatgtaaaa tcacaggatt ttcgtgtgtg gtcttctaca cagacaagat gaaacaattc
     4561 ggcattaata cctgagagca ggaagagcaa gataaaaggt agtatttgtt ggcgatcccc
     4621 ctagagtctt ttacatcttc ggaaaacaaa aactattttt tctttaattt ctttttttac
     4681 tttctatttt taatttatat atttatatta aaaaatttaa attataatta tttttatagc
     4741 acgtgatgaa aaggacccag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg
     4801 tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat
     4861 gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat
     4921 tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt
     4981 aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag
     5041 cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa
     5101 agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg
     5161 ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct
     5221 tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac
     5281 tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca
     5341 caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat
     5401 accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact
     5461 attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc
     5521 ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga
     5581 taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg
     5641 taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg
     5701 aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca
     5761 agtttactca tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta
     5821 ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca
     5881 ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg
     5941 cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga
     6001 tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa
     6061 tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc
     6121 tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg
     6181 tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac
     6241 ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct
     6301 acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc
     6361 ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg
     6421 gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg
     6481 ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct
     6541 ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga
     6601 taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg
     6661 cagcgagtca gtgagcgagg aagcggaaga gcgcccaata cgcaaaccgc ctctccccgc
     6721 gcgttggccg attcattaat gcagctggca cgacaggttt cccgactgga aagcg
//
