Barrio, A. M. | Search bioRxiv

bioRxiv2023

Molecular Pixelation: Single cell spatial proteomics by sequencing

The spatial distribution of cell surface proteins govern vital processes of the immune system such as inter-cell communication and mobility. However, tools for studying these at high multiplexing scale, resolution, and throughput needed to drive novel discoveries are lacking. We present Molecular Pixelation, a DNA-sequencing based method for single cell analysis to quantify protein abundance, spatial distribution, and colocalization of targeted proteins using Antibody Oligonucleotide Conjugates (AOCs). Relative locations of AOCs are inferred by sequentially associating these into local neighborhoods using DNA-pixels containing unique pixel identifier (UPI) sequences, forming >1,000 connected spatial zones per single cell in three dimensions. DNA-sequencing reads are computationally arranged into spatial single cell maps for 76 proteins without cell compartmentalization. By studying immune cell dynamics and using spatial statistics on graph representations of the data, previously known and novel patterns of protein spatial polarization and co-localization were found in chemokine-stimulated T-cells.

immunology↗

bioRxiv2019

A robust benchmark for germline structural variant detection

New technologies and analysis methods are enabling genomic structural variants (SVs) to be detected with ever-increasing accuracy, resolution, and comprehensiveness. Translating these methods to routine research and clinical practice requires robust benchmark sets. We developed the first benchmark set for identification of both false negative and false positive germline SVs, which complements recent efforts emphasizing increasingly comprehensive characterization of SVs. To create this benchmark for a broadly consented son in a Personal Genome Project trio with broadly available cells and DNA, the Genome in a Bottle (GIAB) Consortium integrated 19 sequence-resolved variant calling methods, both alignment- and de novo assembly-based, from short-, linked-, and long-read sequencing, as well as optical and electronic mapping. The final benchmark set contains 12745 isolated, sequence-resolved insertion and deletion calls [≥]50 base pairs (bp) discovered by at least 2 technologies or 5 callsets, genotyped as heterozygous or homozygous variants by long reads. The Tier 1 benchmark regions, for which any extra calls are putative false positives, cover 2.66 Gbp and 9641 SVs supported by at least one diploid assembly. Support for SVs was assessed using svviz with short-, linked-, and long-read sequence data. In general, there was strong support from multiple technologies for the benchmark SVs, with 90 % of the Tier 1 SVs having support in reads from more than one technology. The Mendelian genotype error rate was 0.3 %, and genotype concordance with manual curation was >98.7 %. We demonstrate the utility of the benchmark set by showing it reliably identifies both false negatives and false positives in high-quality SV callsets from short-, linked-, and long-read sequencing and optical mapping.

genomics↗