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Gaspero, A.

Publications and source records attributed to Gaspero, A..

2 recordsLinked to original sources

Soft Windowing Application to Improve Analysis of High-throughput Phenotyping Data

MotivationHigh-throughput phenomic projects generate complex data from small treatment and large control groups that increase the power of the analyses but introduce variation over time. A method is needed to utlize a set of temporally local controls that maximises analytic power while minimising noise from unspecified environmental factors.\n\nResultsHere we introduce \"soft windowing\", a methodological approach that selects a window of time that includes the most appropriate controls for analysis. Using phenotype data from the International Mouse Phenotyping Consortium (IMPC), adaptive windows were applied such that control data collected proximally to mutants were assigned the maximal weight, while data collected earlier or later had less weight. We applied this method to IMPC data and compared the results with those obtained from a standard non-windowed approach. Validation was performed using a resampling approach in which we demonstrate a 10% reduction of false positives from 2.5 million analyses. We applied the method to our production analysis pipeline that establishes genotype-phenotype associations by comparing mutant versus control data. We report an increase of 30% in significant p-values, as well as linkage to 106 versus 99 disease models via phenotype overlap with the soft windowed and non-windowed approaches, respectively, from a set of 2,082 mutant mouse lines. Our method is generalisable and can benefit large-scale human phenomic projects such as the UK Biobank and the All of Us resources.\n\nAvailability and ImplementationThe method is freely available in the R package SmoothWin, available on CRAN http://CRAN.R-project.org/package=SmoothWin.

bioinformatics↗

Employing single-stranded DNA donors for the high-throughput production of conditional knockout alleles in mice

The International Mouse Phenotyping Consortium is generating null allele mice for every protein-coding gene in the genome and characterizing these mice to identify gene-phenotype associations. To test the feasibility of using CRISPR/Cas9 gene editing to generate conditional knockout mice for this large-scale resource, we employed Cas9-mediated homology driven repair (HDR) with short and long single-stranded oligodeoxynucleotides (ssODNs and lssODNs). Using pairs of guides and ssODNs donating loxP sites, we obtained putative conditional allele founder mice, harboring both loxP sites, for 23 of 30 genes targeted. LoxP sites integrated in cis in at least one F0 for 18 of 23 targeted genes. However, loxP sites were mutagenized in 4 of 18 in cis lines. HDR efficiency correlated with Cas9 cutting efficiency but was not influenced by ssODN homology arm symmetry. By contrast, using pairs of guides and a single lssODN to introduce a loxP-flanked exon, conditional allele founders were generated for all 4 genes targeted. Our studies demonstrate that Cas9-mediated HDR with pairs of ssODNs can generate conditional null alleles at many loci, but reveal inefficiencies when applied at scale. In contrast, lssODNs are amenable to high-throughput production of conditional alleles when they can be employed.

genetics↗