A multiple alignment workflow shows the effect of repeat masking and parameter tuning on alignment in plants

Abstract

Alignments of multiple genomes are a cornerstone of comparative genomics, but generating these alignments remains technically challenging and often impractical. We developed the msa_pipeline workflow (https://bitbucket.org/bucklerlab/msa_pipeline) to allow practical and sensitive multiple alignment of diverged plant genomes and calculation of conservation scores with minimal user inputs. As high repeat content and genomic divergence are substantial challenges in plant genome alignment, we also explored the effect of different masking approaches and parameters of the LAST aligner using genome assemblies of 33 grass species. Compared with conventional masking with RepeatMasker, a masking approach based on k‐mers (nucleotide sequences of k length) increased the alignment rate of coding sequence and noncoding functional regions by 25 and 14%, respectively. We further found that default alignment parameters generally perform well, but parameter tuning can increase the alignment rate for noncoding functional regions by over 52% compared with default LAST settings. Finally, by increasing alignment sensitivity from the default baseline, parameter tuning can increase the number of noncoding sites that can be scored for conservation by over 76%. Overall, tuning of masking and alignment parameters can generate optimized multiple alignments to drive biological discovery in plants.

Genetic Incompatibilities and Evolutionary Rescue by Wild Relatives Shaped Grain Amaranth Domestication

“…We aligned 15 repeat-masked genomes of angiosperm species spanning a large taxonomic range to the reference genome of A. hypochondriacus (v2.1) ( Lightfoot et al 2017 ). We followed the pipeline of Wu et al (2022) . Briefly, we aligned genomes of 16 divergent species: Beta vulgaris EL10 1.0, Brachypodium distachyon v2.1, Chenopodium quinoa v1.0, Glycine max Wm82.a4.v1, Helianthus annuus r1.2, Medicago truncatula Mt4.0v1, Mimulus guttatus v2.0, Oryza sativa v7.0, Phaseolus vulgaris v2.1, Populus trichocarpa v4.1, Setaria viridis v2.1, Solanum lycopersicum ITAG4.0, Sorghum bicolor v3.1.1, Spinacia oleracea (Monoe Viroflay), and Vitis vinifera v2.1 from phytozome ( Goodstein et al 2012 ) to the A. hypochondriacus reference genome using the LAST aligner ( Kiełbasa et al 2011 ).…”

Whole Genome Alignment: Methods, Challenges, and Future Directions

“…Traditionally, aligning divergent genomes has been challenging due to the high levels of sequence divergence and the presence of unique genomic elements. However, tools like LASTZ and MULTIZ have been designed to tackle these challenges, enabling the alignment of divergent genomes [56,57].…”

Whole-Genome Alignment: Methods, Challenges, and Future Directions

“…Traditionally, aligning divergent genomes has been challenging due to the high levels of sequence divergence and the presence of unique genomic elements. However, tools like LASTZ and MULTIZ have been designed to tackle these challenges, enabling the alignment of divergent genomes [25,69].…”