Majority of the highly variable NLRs in maize share genomic location and contain additional target-binding domains.

Nucleotide-binding, Leucine Rich Repeat proteins (NLRs) are a major class of immune receptors in plants. NLRs include both conserved and rapidly evolving members, however their evolutionary trajectory in crops remains understudied. Availability of crop pan-genomes enables analysis of the recent events in the evolution of this highly complex gene family within domesticated species. Here, we investigated the NLR complement of 26 nested association mapping (NAM) founder lines of maize. We found that maize has just four main subfamilies containing rapidly evolving highly variable NLR (hvNLR) receptors. Curiously, three of these phylogenetically distinct hvNLR lineages are located in adjacent clusters on chromosome 10. Members of the same hvNLR clade show variable expression and methylation across lines and tissues, consistent with their rapid evolution. By combining sequence diversity analysis and AlphaFold2 computational structure prediction we predicted ligand binding sites in the hvNLRs. We also observed novel insertion domains in the LRR regions of two hvNLR subfamilies that likely contribute to target recogniton. To make this analysis accessible, we created NLRCladeFinder, a Google Colaboratory notebook, that accepts any newly identified NLR sequence, places it in the evolutionary context of the maize pan-NLRome, and provides an updated clade alignment, phylogenetic tree, and sequence diversity information for the gene of interest.

Quality control of large genome datasets.

The 1000 Genomes Project (TGP) is a foundational resource that serves the biomedical community as a standard reference cohort for human genetic variation. There are now seven public versions of these genomes. The TGP Consortium produced the first by mapping its final data release against human reference sequence GRCh37, then "lifted over" these genomes to the improved reference sequence (GRCh38) when it was released, and remapped the original data to GRCh38 with two similar pipelines. As best-practice quality validation, the pipelines that generated these versions were benchmarked against the Genome In A Bottle Consortium's "platinum quality" genome (NA12878). The New York Genome Center recently released the results of independently resequencing the cohort at greater depth (30×), a phased version informed by the inclusion of related individuals, and independently remapped the original variant calls to GRCh38. We performed a cross-comparison evaluation of all seven versions using genome fingerprinting, which supports ultrafast genome comparison even across reference versions. We noted multiple issues, including discrepancies in cohort membership, disagreement on the overall level of variation, evidence of substandard pipeline performance on specific genomes and in specific regions of the genome, cryptic relationships between individuals, inconsistent phasing, and annotation distortions caused by the history of the reference genome itself. We therefore recommend global quality assessment by rapid genome comparisons, alongside benchmarking as part of best-practice quality assessment of large genome datasets. Our observations also help inform the decision of which version to use, to support analyses by individual researchers.