Bloom and Starr Labs SARS-CoV-2 RBD SSM Libraries
(Pooled Libraries #1000000182, #1000000183, #1000000184, #1000000185, #1000000186, #1000000187, #1000000188, #1000000231, #1000000232, #1000000246, #1000000247, #1000000248)

• Purpose:

  Site-saturation mutagenesis (SSM) libraries of yeast-displayed SARS-CoV-2 receptor-binding domain (RBD). Each SSM library is in the background of fixed mutations that define certain SARS-CoV-2 variants that have evolved, including the ancestral Wuhan-Hu-1 strain and variants Alpha (N501Y), Beta (K417N/E484K/N501Y), Delta (L452R/T478K), Eta (E484K), Omicron BA.1 (15 mutations), Omicron BA.2 (16 mutations), Omicron BQ.1.1 (20 mutations), Omicron XBB.1.5 (23 mutations), Omicron EG.5 (24 mutations), Omicron FLip (i.e. HK3, 25 mutations), and Omicron BA.2.86 (27 mutations) RBD. Each library has a downstream N16 barcode that was previously linked to mutant identity via PacBio sequencing.

  Libraries are useful for deep mutational scanning characterization of the impacts of SARS-CoV-2 mutations on receptor- and antibody-binding.

• Vector Backbone:

  pETcon(-)

• Depositing Labs: Jesse Bloom, Tyler Starr
• Publication:

  #1000000182, #1000000183, #1000000184, #1000000185, #1000000186:
  Starr et al Science. 2022 Jun 28:eabo7896. doi: 10.1126/science.abo7896.

  #1000000187, #1000000188:
  Starr et al PLoS Pathog. 2022 Nov 18;18(11):e1010951. doi: 10.1371/journal.ppat.1010951.

  #1000000231, #1000000232:
  Taylor et al. PLoS Pathog. 2023. doi: 10.1371/journal.ppat.1011901.

  #1000000246, #1000000247, #1000000248:
  Taylor et al. Virus Evol. 2024. doi: 10.1093/ve/veae067.

Library Details

• Mutagenized CDS: SARS-CoV-2 Receptor Binding Domain
• Mutagenized region size: 603 bp (The RBD insert expressed as a yeast-display fusion protein is 201 aa / 603 nt for EG.5 and FLip, 200 aa / 600 nt for BA.2.86.)

Each catalog item is a pair of duplicate libraries to be used separately as technical replicates, as described in (Starr and Greaney et al. 2020 Cell and Starr et al. 2022 bioRxiv).

Each catalog item also contains a separate bacterial stab of the parent control plasmid, as listed in the following table.

Pooled Library	Background	Parent Plasmid #	Plasmids/library
1000000182	Wuhan-Hu-1	166782	~50,000
1000000183	Alpha	184405	~50,000
1000000184	Beta	184406	~50,000
1000000185	Delta	184407	~100,000
1000000186	Eta	184404	~50,000
1000000187	Omicron BA.1	184408	~100,000
1000000188	Omicron BA.2	184409	~100,000
1000000231	Omicron BQ.1.1	208085	~100,000
1000000232	Omicron XBB.1.5	208086	~100,000
1000000246	Omicron EG.5	222232	~100,000
1000000247	Omicron FLip	222230	~100,000
1000000248	Omicron BA.2.86	222231	~100,000

The parent plasmid is included with each library purchased. Please do NOT order this plasmid in addition to the library.

Library Shipping

Each library will be delivered as two pooled DNA libraries in microcentrifuge tubes on blue ice (tubes labeled lib1 and lib2). The tube contents will not necessarily be frozen. For best results, minimize freeze-thaws.

• Volume: ∼10 µL
• Concentration: 25 ng/µL

You will also receive a bacterial stab of the parental-control plasmid.

Resource Information

• Protocols:
  • Library Transformation Protocol (PDF, 35 KB)
  • PCR protocol for Illumina sequencing preparation (PDF, 34 KB)
• Depositor Data:
  • Table of primers (Bloom Lab Github)
  • Table of codon variants [Wuhan-Hu-1, Alpha, Beta, Eta] (Bloom Lab Github)
  • Table of codon variants [Delta] (Bloom Lab Github)
  • Table of codon variants [Omicron BA.1, BA.2] (Bloom Lab Github)
  • Table of codon variants [Omicron BQ.1.1, XBB.1.5] (Starr Lab Github)
  • Table of codon variants [Omicron EG5, FLip, BA.2.86] (Starr Lab Github)
  • Representative Illumina sequencing results [Omicron BQ.1.1, XBB.1.5] (Starr Lab Github)
  • Representative Illumina sequencing results [Omicron EG5, FLip, BA.2.86] (Starr Lab Github)
  • Representative plasmid map [Omicron BQ.1.1] (GB, 31 KB)
  • Representative plasmid map [Omicron XBB.1.5] (GB, 33 KB)
  • Representative plasmid map [Omicron EG.5] (GB, 51 KB)
  • Representative plasmid map [Omicron BA.2.86] (GB, 35 KB)
  • Representative plasmid map [Omicron FLip] (GB, 45 KB)
• Articles Citing this Pooled Library:
  • Wuhan-Hu-1 SSM (#1000000182)
  • Omicron BA.1 SSM (#1000000187)
  • Omicron BA.2 SSM (#1000000188)

Terms and Licenses

Academic/Nonprofit Terms:

• UBMTA

Industry Terms:

• Not Available to Industry

Trademarks:

• Zeocin® is an InvivoGen trademark.

Depositor Comments

The following GitHub repositories are provided:

• SARS-CoV-2-RBD_DMS_variants Includes algorithms for analyzing the PacBio and Illumina sequencing data for the Wuhan-Hu-1, Alpha, Beta, and Eta libraries
• SARS-CoV-2-RBD_Delta Includes algorithms for analyzing the PacBio and Illumina sequencing data for the Delta library
• SARS-CoV-2-RBD_DMS_Omicron Includes algorithms for analyzing the PacBio and Illumina sequencing data for the Omicron BA.1 and BA.2 libraries
• SARS-CoV-2-RBD_DMS_Omicron-XBB-BQ Includes algorithms for analyzing the PacBio and Illumina sequencing data for the Omicron BQ.1.1 and XBB.1.5 libraries
• SARS-CoV-2-RBD_DMS_Omicron-EG5-FLip-BA286 Includes algorithms for analyzing the PacBio and Illumina sequencing data for the Omicron EG5, FLip, and BA.2.86 libraries

The SARS-CoV-2 RBD Beta SSM Library is also described in Greaney et al. 2022 "A SARS-CoV-2 variant elicits an antibody response with a shifted immunodominance hierarchy" PLoS Pathog. doi: 10.1371/journal.ppat.1010248

The SARS-CoV-2 RBD Delta SSM Library is also described in Greaney et al. 2022 "The SARS-CoV-2 Delta variant induces an antibody response largely focused on class 1 and 2 antibody epitopes" bioRxiv. doi: 10.1101/2022.03.12.484088

The SARS-CoV-2 RBD Omicron BQ.1.1 SSM and XBB.1.5 Libraries are also described in Taylor et al. 2023 "Deep mutational scans of XBB.1.5 and BQ.1.1 reveal ongoing epistatic drift during SARS-CoV-2 evolution" bioRxiv. doi: 10.1101/2023.09.11.557279

The SARS-CoV-2 RBD Omicron EG5, FLip, and BA286 Libraries are also described in Taylor et al. 2024 "Deep mutational scanning of SARS-CoV-2 Omicron BA.2.86 and epistatic emergence of the KP.3 variant" bioRxiv. doi: 10.1101/2024.07.23.604853

• Overview of the method for library assembly and barcode-variant association.