Collection Citation: Warren van Loggerenberg, Daniel Zimmerman, Anna Axakova, Adrine de Souza, Frederick Roth 2026. Variant Effect Mapping Protocol Collection. protocols.io https://dx.doi.org/10.17504/protocols.io.5qpvo1n37g4o/v1
License: This is an open access collection distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this collection and it's working
Created: December 23, 2025
Last Modified: January 27, 2026
Collection Integer ID: 235740
Keywords: DMS, variant effect mapping, VE mapping, deep mutational scanning, human cell-based assay, yeast-based assay, functional assay, MAVE, Multiplex Assay of Variant Effect, MAVE, POPcode, double-tag POPcode, Precision Oligo-Pool Based Code Alteration, TileSeq, multiplexed assays of variant effect, supporting clinical variant interpretation, measurement of variant effect, clinical variant interpretation, comprehensive atlas of variant effect, measured variant function, functional impact of all possible missense variant, disease phenotype, variant effect mapping protocol collection advance, variant effect, based functional assay, variant effect map, functional assay, associated gene, gene, enabled multiplexed assay, including variant, directed mutagenesi, possible missense variant, variant, generating mutagenic library, based assay, functional score, mutagenic library
Funders Acknowledgements:
(NIH/NHGRI) Center of Excellence in Genomic Science Initiative
Grant ID: HG010461
(NIH/NHGRI) Impact of Genomic Variation on Function Initiative
Grant ID: UM1HG011989
NIH/NIAID
Grant ID: AI173827
Abstract
Advances in cellular engineering and sequencing have enabled Multiplexed Assays of Variant Effect (MAVEs), a technological approach to proactively measure the functional impact of all possible missense variants in a disease-associated gene, including variants not yet observed in the clinic. The resulting “lookup tables” of measured variant function are commonly referred to as variant effect maps. There is a growing international effort to generate a comprehensive atlas of variant effects, with value not only for defining sequence–structure–function relationships and supporting clinical variant interpretation, but also for demonstrating quantitative correlations between functional scores and disease phenotypes.
The protocols in this collection are organized sequentially and guide the user through preparing the gene of interest with a stop codon, performing site-directed mutagenesis, validating a scalable human cell- and yeast-based functional assay, generating mutagenic libraries, and executing a MAVE in a human cell line or in yeast. These workflows enable the measurement of variant effects en masse using complementary readouts — including growth-based assays and fluorophore reporters — to assess outputs such as total enzymatic activity and protein stability.
S Sun, F Yang, G Tan, M Costanzo, R Oughtred, J Hirschman, C Theesfeld, P Bansal, N Sahni, S Yi, A Yu, T Tyagi, C Tie, DE Hill, M Vidal, BJ Andrews, C Boone, K Dolinski & FP Roth†. An extended set of yeast-based functional assays accurately identifies human disease mutations. Genome Research 26(5):670-80 (2016). Full Text
J Weile*, S Sun*, AG Cote, J Knapp, M Verby, JC Mellor, Y Wu, C Pons, C Wong, N van Lieshout, F Yang, M Tasan, G Tan, S Yang, DM Fowler, R Nussbaum, JD Bloom, M Vidal, DE Hill, P Aloy & FP Roth†. A framework for exhaustively mapping functional missense variants. Molecular Systems Biology 13:157 (2017) Full Text
S Sun†, J Weile†, M Verby, Y Wu, Y Wang, AG Cote, I Fotiadou, J Kitaygorodsky, M Vidal, J Rine, P Jesina, V Kozich†, FP Roth†. A proactive genotype-to-patient-phenotype map for cystathionine beta-synthase. Genome Medicine 12(1):13 (2020). Full Text
W van Loggerenberg*, S Sowlati-Hashjin*, J Weile, R Hamilton, A Chawla, D Sheykhkarimli, M Gebbia, N Kishore, L Fresard, S Mustajoki, E Pischik, E Di Pierro, M Barbaro, Y Floderus, C Schmitt, L Gouya, A Colavin, R Nussbaum, ECH Friesema, R Kauppinen, J To-Figueras, AK Aarsand, RJ Desnick†, M Garton†, FP Roth†. Systematically testing human HMBS missense variants to reveal mechanism and pathogenic variation. American Journal of Human Genetics 110(10):1769-1786 (2023). Full Text
M Gebbia*, D Zimmerman*, R Jiang, M Nguyen, J Weile, R Li, M Gavac, N Kishore, S Sun, RA Boonen, R Hamilton, JN Dines, A Wahl, J Reuter, B Johnson, DM Fowler, F Couch, H van Attikum, FP Roth†. A missense variant effect map for the human tumour suppressor protein CHK2. American Journal of Human Genetics 111(12): 2675-2692 (2024). Full Text
A Axakova, M Ding, AG Cote, R Subramaniam, V Senguttuvan, H Zhang, J Weile, SV Douville, M Gebbia, A Al-Chalabi, A Wahl, J Reuter, J Hurt, A Mitchell, S Fradette, PM Andersen, W van Loggerenberg†, FP Roth†. Landscapes of missense variant impact for human superoxide dismutase 1. American Journal of Human Genetics 112:1-21 (2025). Full Text
D Zimmerman, A Cote, W van Loggerenberg, M Gebbia, N Kishore, J Weile, R Li, C Reno, A Marsh, F Hernandez, P Shahagadkar, L Grove, S Meier, H-J Wu, S Fengolia, L Ahronian, T Teng, AJ Waters, D Seward, M Taipale, M Aronson, ME Richardson, D Adams, FP Roth†. Comprehensively testing the function of missense variation in the STK11 tumour suppressor. bioRxiv https://doi.org/10.1101/2025.07.14.664734 (2025). Full Text
DR Tabet*, AG Cote*, MC Lancaster, J Weile, A Rayhan, I Fotiadou, N Kishore, R Li, D Kuang, JJ Knapp, CS Carrero, O Taverniti, A Axakova, JMP Castelli, MM Islam, S Sowlati-Hashjin, A Gandhi, R Maaieh, M Garton, K Matreyek, DM Fowler, M Bourbon, SG Pfisterer, AM Glazer, BM Kroncke, VN Parikh, EA Ashley, JW Knowles, M Claussnitzer, ET Cirulli, RA Hegele, DM Roden, CA MacRae, FP Roth†. The functional landscape of coding variation in the familial hypercholesterolemia gene LDLR. Science [First Release Oct 30] (2025). Full Text
A Axakova, A Berger, W van Loggerenberg, N Kishore, M Gebbia, MX Ding, SV Douville, DR Tabet, AG Cote, J Weile, S Johansson, E Bratland, FP Roth†. Systematic and proactive evaluation of AIRE missense variant effects. bioRxiv https://doi.org/10.1101/2025.11.18.688830 (2025). Full Text
