LOCUS       Exported                3488 bp ds-DNA     circular SYN 04-AUG-2021
DEFINITION  Encodes the SARS-CoV-2 Spike Receptor Binding Domain (RBD) with the 
            E484K mutation and can be combined with pJS697 (Addgene plasmid # 
            168778) through homologous recombination.
ACCESSION   .
VERSION     .
KEYWORDS    pIFU001
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3488)
  AUTHORS   Urdaniz IF, Steiner PJ, Kirby MB, Zhao F, Haas CM, Barman S, Rhodes 
            ER, Peng L, Sprenger KG, Jardine JG, Whitehead TA
  TITLE     One-shot identification of SARS-CoV-2 S RBD escape mutants using 
            yeast screening
  JOURNAL   bioRxiv 2021.03.15.435309
REFERENCE   2  (bases 1 to 3488)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 4, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..3488
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     protein_bind    106..127
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        142..172
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    180..196
                     /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     185..207
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     204..220
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     204..220
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             271..282
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             283..309
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             994..1023
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     primer_bind     complement(1128..1145)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1128..1144)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1137..1159)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(1353..1372)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1472..1494
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1532..1550)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1618..1722
                     /gene="bla"
                     /label=AmpR promoter
     CDS             1723..2538
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(1796..1815)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      2718..3306
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3207..3226
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3460..3477
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
       61 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaatg tgagttagct
      121 cactcattag gcaccccagg ctttacactt tatgcttccg gctcgtatgt tgtgtggaat
      181 tgtgagcgga taacaatttc acacaggaaa cagctatgac catgattacg ccaagcttgc
      241 atgcctgcag gtcgacgcgg ccgctcgacg attgaaggta gatacccata cgacgttcca
      301 gactacgctc tgcaggctag tggtggagga ggctctggtg gaggcggtag cggaggcgga
      361 gggtcgacaa acttgtgccc ttttggtgaa gtttttcaag ccaccagatt tgcatctgtt
      421 tatgcttgga acaggaagag aatcagcaac tgtgttgctg attattctgt cctatataat
      481 tccgcatcat tttccacttt taagtgttat ggagtgtctc ctactaaatt aaatgatctc
      541 tgctttacta atgtctatgc agattcattt gtaattagag gtgatgaagt cagacaaatc
      601 gctccagggc aaactggaaa gattgctgat tataattata aattaccaga tgattttaca
      661 ggctgcgtta tagcttggaa ttctaacaat cttgattcta aggttggtgg taattataat
      721 tacctgtata gattgtttag gaagtctaat ctcaaacctt ttgagagaga tatttcaact
      781 gaaatctatc aggccggtag cacaccttgt aatggtgtta aaggttttaa ttgttacttt
      841 cctttacaat catatggttt ccaacccact aatggtgttg gttaccaacc atacagagta
      901 gtagtacttt cttttgaact tctacatgca ccagcaactg tttgtggacc taaaaagtct
      961 actaatttgg ttaaaaacaa agggggcgga tccgaacaaa agcttatttc tgaagaggac
     1021 ttgtaataga gatctgataa caacagtgta gatgtaacaa aatcgacttt gttcccactg
     1081 tacttttagc ggccgccctc agcgatatcg gtaccgagct cgaattcact ggccgtcgtt
     1141 ttacaacgtc gtgactggga aaaccctggc gttacccaac ttaatcgcct tgcagcacat
     1201 ccccctttcg ccagctggcg taatagcgaa gaggcccgca ccgatcgccc ttcccaacag
     1261 ttgcgcagcc tgaatggcga atggcgcctg atgcggtatt ttctccttac gcatctgtgc
     1321 ggtatttcac accgcatatg gtgcactctc agtacaatct gctctgatgc cgcatagtta
     1381 agccagcccc gacacccgcc aacacccgct gacgcgccct gacgggcttg tctgctcccg
     1441 gcatccgctt acagacaagc tgtgaccgtc tccgggagct gcatgtgtca gaggttttca
     1501 ccgtcatcac cgaaacgcgc gagacgaaag ggcctcgtga tacgcctatt tttataggtt
     1561 aatgtcatga taataatggt ttcttagacg tcaggtggca cttttcgggg aaatgtgcgc
     1621 ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa
     1681 taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagcca tattcaacgg
     1741 gaaacgtctt gctctaggcc gcgattaaat tccaacatgg atgctgattt atatgggtat
     1801 aaatgggctc gcgataatgt cgggcaatca ggtgcgacaa tctatcgatt gtatgggaag
     1861 cccgatgcgc cagagttgtt tctgaaacat ggcaaaggta gcgttgccaa tgatgttaca
     1921 gatgagatgg tcagactaaa ctggctgacg gaatttatgc ctcttccgac catcaagcat
     1981 tttatccgta ctcctgatga tgcatggtta ctcaccactg cgatccccgg gaaaacagca
     2041 ttccaggtat tagaagaata tcctgattca ggtgaaaata ttgttgatgc gctggcagtg
     2101 ttcctgcgcc ggttgcattc gattcctgtt tgtaattgtc cttttaacag cgatcgcgta
     2161 tttcgtctcg ctcaggcgca atcacgaatg aataacggtt tggttgatgc gagtgatttt
     2221 gatgacgagc gtaatggctg gcctgttgaa caagtctgga aagaaatgca taaacttttg
     2281 ccattctcac cggattcagt cgtcactcat ggtgatttct cacttgataa ccttattttt
     2341 gacgagggga aattaatagg ttgtattgat gttggacgag tcggaatcgc agaccgatac
     2401 caggatcttg ccatcctatg gaactgcctc ggtgagtttt ctccttcatt acagaaacgg
     2461 ctttttcaaa aatatggtat tgataatcct gatatgaata aattgcagtt tcatttgatg
     2521 ctcgatgagt ttttctaacc tcagcaactg tcagaccaag tttactcata tatactttag
     2581 attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat
     2641 ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa
     2701 aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca
     2761 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt
     2821 ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg
     2881 tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc
     2941 ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga
     3001 cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc
     3061 agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc
     3121 gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca
     3181 ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg
     3241 tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta
     3301 tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct
     3361 cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag
     3421 tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa
     3481 gcggaaga
//
