Fast neutron mutagenesis in soybean enriches for small indels and creates frameshift mutations

AbstractThe mutagenic effects of ionizing radiation have been used for decades to create novel variants in experimental populations. Fast neutron (FN) bombardment as a mutagen has been especially widespread in plants, with extensive reports describing the induction of large structural variants, i.e., deletions, insertions, inversions, and translocations. However, the full spectrum of FN-induced mutations is poorly understood. We contrast small insertions and deletions (indels) observed in 27 soybean lines subject to FN irradiation with the standing indels identified in 107 diverse soybean lines. We use the same populations to contrast the nature and context (bases flanking a nucleotide change) of single nucleotide variants. The accumulation of new single nucleotide changes in FN lines is marginally higher than expected based on spontaneous mutation. In FN treated lines and in standing variation, C→T transitions and the corresponding reverse complement G→A transitions are the most abundant and occur most frequently in a CpG local context. These data indicate that most SNPs identified in FN lines are likely derived from spontaneous de novo processes in generations following mutagenesis, rather than from the FN irradiation mutagen. However, small indels in FN lines differ from standing variants. Short insertions, from 1 – 6 base pairs, are less abundant than in standing variation. Short deletions are more abundant and prone to induce frameshift mutations that should disrupt the structure and function of encoded proteins. These findings indicate that FN irradiation generates numerous small indels, increasing the abundance of loss of function mutations that will impact single genes.Significance StatementIrradiation mutagenesis is commonly viewed as a method to induce large structural variants in genomes. We also find enrichment in small insertion and deletion (indel) variants. The radiation-mutagenized lines averaged 32 indels per line, far exceeding the number estimated to occur by spontaneous processes, indicating that these arose from the irradiation treatment. Nevertheless, naturally-occurring standing variation among soybean accessions is still four orders of magnitude higher than the level of diversity introduced by mutagenesis. Induced mutations from any source are likely to constitute a relatively small portion of the genetic variation present in crop species. However, irradiation mutagenesis is useful for altering genomes by introducing small indels into single genes or disrupting gene clusters by creating structural variants.

Download Full-text

An integrative approach to investigate the respective roles of single-nucleotide variants and copy-number variants in Attention-Deficit/Hyperactivity Disorder

Scientific Reports ◽

10.1038/srep22851 ◽

2016 ◽

Vol 6 (1) ◽

Cited By ~ 9

Author(s):

Leandro de Araújo Lima ◽

Ana Cecília Feio-dos-Santos ◽

Sintia Iole Belangero ◽

Ary Gadelha ◽

Rodrigo Affonseca Bressan ◽

...

Keyword(s):

Attention Deficit Hyperactivity Disorder ◽

Attention Deficit ◽

Copy Number ◽

De Novo ◽

Copy Number Variants ◽

Integrative Approach ◽

Single Nucleotide Variants ◽

Single Nucleotide ◽

Hyperactivity Disorder ◽

New Genes

Abstract Many studies have attempted to investigate the genetic susceptibility of Attention-Deficit/Hyperactivity Disorder (ADHD), but without much success. The present study aimed to analyze both single-nucleotide and copy-number variants contributing to the genetic architecture of ADHD. We generated exome data from 30 Brazilian trios with sporadic ADHD. We also analyzed a Brazilian sample of 503 children/adolescent controls from a High Risk Cohort Study for the Development of Childhood Psychiatric Disorders, and also previously published results of five CNV studies and one GWAS meta-analysis of ADHD involving children/adolescents. The results from the Brazilian trios showed that cases with de novo SNVs tend not to have de novo CNVs and vice-versa. Although the sample size is small, we could also see that various comorbidities are more frequent in cases with only inherited variants. Moreover, using only genes expressed in brain, we constructed two “in silico” protein-protein interaction networks, one with genes from any analysis, and other with genes with hits in two analyses. Topological and functional analyses of genes in this network uncovered genes related to synapse, cell adhesion, glutamatergic and serotoninergic pathways, both confirming findings of previous studies and capturing new genes and genetic variants in these pathways.

Download Full-text

Implications of Genetic Distance to Reference and De Novo Genome Assembly for Clinical Genomics in Africans

10.1101/2020.09.25.20201780 ◽

2020 ◽

Author(s):

Daniel Shriner ◽

Adebowale Adeyemo ◽

Charles Rotimi

Keyword(s):

Genetic Distance ◽

De Novo ◽

Reference Sequence ◽

Sequencing Data ◽

Single Nucleotide Variants ◽

De Novo Genome Assembly ◽

Single Nucleotide ◽

Clinical Genomics ◽

Advantages And Disadvantages ◽

False Discovery

In clinical genomics, variant calling from short-read sequencing data typically relies on a pan-genomic, universal human reference sequence. A major limitation of this approach is that the number of reads that incorrectly map or fail to map increase as the reads diverge from the reference sequence. In the context of genome sequencing of genetically diverse Africans, we investigate the advantages and disadvantages of using a de novo assembly of the read data as the reference sequence in single sample calling. Conditional on sufficient read depth, the alignment-based and assembly-based approaches yielded comparable sensitivity and false discovery rates for single nucleotide variants when benchmarked against a gold standard call set. The alignment-based approach yielded coverage of an additional 270.8 Mb over which sensitivity was lower and the false discovery rate was higher. Although both approaches detected and missed clinically relevant variants, the assembly-based approach identified more such variants than the alignment-based approach. Of particular relevance to individuals of African descent, the assembly-based approach identified four heterozygous genotypes containing the sickle allele whereas the alignment-based approach identified no occurrences of the sickle allele. Variant annotation using dbSNP and gnomAD identified systematic biases in these databases due to underrepresentation of Africans. Using the counts of homozygous alternate genotypes from the alignment-based approach as a measure of genetic distance to the reference sequence GRCh38.p12, we found that the numbers of misassemblies, total variant sites, potentially novel single nucleotide variants (SNVs), and certain variant classes (e.g., splice acceptor variants, stop loss variants, missense variants, synonymous variants, and variants absent from gnomAD) were significantly correlated with genetic distance. In contrast, genomic coverage and other variant classes (e.g., ClinVar pathogenic or likely pathogenic variants, start loss variants, stop gain variants, splice donor variants, incomplete terminal codons, variants with CADD score ≥20) were not correlated with genetic distance. With improvement in coverage, the assembly-based approach can offer a viable alternative to the alignment-based approach, with the advantage that it can obviate the need to generate diverse human reference sequences or collections of alternate scaffolds.

Download Full-text

Recent Advances in Understanding the Genetic Architecture of Autism

Annual Review of Genomics and Human Genetics ◽

10.1146/annurev-genom-121219-082309 ◽

2020 ◽

Vol 21 (1) ◽

pp. 289-304 ◽

Cited By ~ 1

Author(s):

Caroline M. Dias ◽

Christopher A. Walsh

Keyword(s):

Genetic Architecture ◽

De Novo ◽

Clinical Care ◽

Copy Number Variants ◽

Autism Spectrum ◽

Loss Of Function ◽

Single Nucleotide Variants ◽

Single Nucleotide ◽

Recent Advances ◽

Insight Into

Recent advances in understanding the genetic architecture of autism spectrum disorder have allowed for unprecedented insight into its biological underpinnings. New studies have elucidated the contributions of a variety of forms of genetic variation to autism susceptibility. While the roles of de novo copy number variants and single-nucleotide variants—causing loss-of-function or missense changes—have been increasingly recognized and refined, mosaic single-nucleotide variants have been implicated more recently in some cases. Moreover, inherited variants (including common variants) and, more recently, rare recessive inherited variants have come into greater focus. Finally, noncoding variants—both inherited and de novo—have been implicated in the last few years. This work has revealed a convergence of diverse genetic drivers on common biological pathways and has highlighted the ongoing importance of increasing sample size and experimental innovation. Continuing to synthesize these genetic findings with functional and phenotypic evidence and translating these discoveries to clinical care remain considerable challenges for the field.

Download Full-text

Haplotype-aware variant calling enables high accuracy in nanopore long-reads using deep neural networks

10.1101/2021.03.04.433952 ◽

2021 ◽

Author(s):

Kishwar Shafin ◽

Trevor Pesout ◽

Pi-Chuan Chang ◽

Maria Nattestad ◽

Alexey Kolesnikov ◽

...

Keyword(s):

De Novo ◽

Sequence Data ◽

Variant Calling ◽

High Accuracy ◽

Superior Performance ◽

Read Length ◽

Single Nucleotide Variants ◽

Single Nucleotide ◽

Short Read ◽

Long Read

Long-read sequencing has the potential to transform variant detection by reaching currently difficult-to-map regions and routinely linking together adjacent variations to enable read based phasing. Third-generation nanopore sequence data has demonstrated a long read length, but current interpretation methods for its novel pore-based signal have unique error profiles, making accurate analysis challenging. Here, we introduce a haplotype-aware variant calling pipeline PEPPER-Margin-DeepVariant that produces state-of-the-art variant calling results with nanopore data. We show that our nanopore-based method outperforms the short-read-based single nucleotide variant identification method at the whole genome-scale and produces high-quality single nucleotide variants in segmental duplications and low-mappability regions where short-read based genotyping fails. We show that our pipeline can provide highly-contiguous phase blocks across the genome with nanopore reads, contiguously spanning between 85% to 92% of annotated genes across six samples. We also extend PEPPER-Margin-DeepVariant to PacBio HiFi data, providing an efficient solution with superior performance than the current WhatsHap-DeepVariant standard. Finally, we demonstrate de novo assembly polishing methods that use nanopore and PacBio HiFi reads to produce diploid assemblies with high accuracy (Q35+ nanopore-polished and Q40+ PacBio-HiFi-polished).

Download Full-text

Next-generation forward genetic screens: using simulated data to improve the design of mapping-by-sequencing experiments in Arabidopsis

Nucleic Acids Research ◽

10.1093/nar/gkz806 ◽

2019 ◽

Vol 47 (21) ◽

pp. e140-e140

Author(s):

David Wilson-Sánchez ◽

Samuel Daniel Lup ◽

Raquel Sarmiento-Mañús ◽

María Rosa Ponce ◽

José Luis Micol

Keyword(s):

Point Mutations ◽

Sequencing Depth ◽

Nucleotide Polymorphisms ◽

Next Generation ◽

Genetic Screens ◽

Induced Mutations ◽

Single Nucleotide Variants ◽

Single Nucleotide ◽

Forward Genetic Screens ◽

Mapping By Sequencing

Abstract Forward genetic screens have successfully identified many genes and continue to be powerful tools for dissecting biological processes in Arabidopsis and other model species. Next-generation sequencing technologies have revolutionized the time-consuming process of identifying the mutations that cause a phenotype of interest. However, due to the cost of such mapping-by-sequencing experiments, special attention should be paid to experimental design and technical decisions so that the read data allows to map the desired mutation. Here, we simulated different mapping-by-sequencing scenarios. We first evaluated which short-read technology was best suited for analyzing gene-rich genomic regions in Arabidopsis and determined the minimum sequencing depth required to confidently call single nucleotide variants. We also designed ways to discriminate mutagenesis-induced mutations from background Single Nucleotide Polymorphisms in mutants isolated in Arabidopsis non-reference lines. In addition, we simulated bulked segregant mapping populations for identifying point mutations and monitored how the size of the mapping population and the sequencing depth affect mapping precision. Finally, we provide the computational basis of a protocol that we already used to map T-DNA insertions with paired-end Illumina-like reads, using very low sequencing depths and pooling several mutants together; this approach can also be used with single-end reads as well as to map any other insertional mutagen. All these simulations proved useful for designing experiments that allowed us to map several mutations in Arabidopsis.

Download Full-text

From the periphery to centre stage: de novo single nucleotide variants play a key role in human genetic disease

Journal of Medical Genetics ◽

10.1136/jmedgenet-2013-101519 ◽

2013 ◽

Vol 50 (4) ◽

pp. 203-211 ◽

Cited By ~ 27

Author(s):

Chee-Seng Ku ◽

Eng King Tan ◽

David N Cooper

Keyword(s):

Genetic Disease ◽

De Novo ◽

Single Nucleotide Variants ◽

Human Genetic Disease ◽

Single Nucleotide

Download Full-text

Burden of de novo mutations and inherited rare single nucleotide variants in children with sensory processing dysfunction

BMC Medical Genomics ◽

10.1186/s12920-018-0362-x ◽

2018 ◽

Vol 11 (1) ◽

Cited By ~ 6

Author(s):

Elysa Jill Marco ◽

Anne Brandes Aitken ◽

Vishnu Prakas Nair ◽

Gilberto da Gente ◽

Molly Rae Gerdes ◽

...

Keyword(s):

Sensory Processing ◽

De Novo ◽

De Novo Mutations ◽

Single Nucleotide Variants ◽

Single Nucleotide

Download Full-text

Weakly deleterious natural genetic variation amplifies probability of resistance in multiplexed gene drive systems

10.1101/2021.12.23.473701 ◽

2021 ◽

Author(s):

Bhavin S Khatri ◽

Austin Burt

Keyword(s):

De Novo ◽

Large Population ◽

Stochastic Simulations ◽

Gene Drive ◽

End Joining ◽

Single Nucleotide ◽

Major Barrier ◽

Standing Variation ◽

Large Populations ◽

Drive Systems

Evolution of resistance is a major barrier to successful deployment of gene drive systems to suppress natural populations. Multiplexed guide RNAs that require resistance mutations in all target cut sites is a promising strategy to overcome resistance. Using novel stochastic simulations that accurately model evolution at very large population sizes, we explore the probability of resistance due to three important mechanisms: 1) non-homologous end-joining mutations, 2) single nucleotide mutants arising de novo or, 3) single nucleotide polymorphisms pre-existing as standing variation. If the fraction of functional end-joining mutants is rare, we show that standing variation dominates, via a qualitatively new phenomenon where weakly deleterious variants significantly amplify the probability of multi-site resistance. This means resistance can be probable even with many target sites in not very large populations. This result has broad application to resistance arising in multi-site evolutionary scenarios including the evolution of vaccine escape mutations in large populations.

Download Full-text

Quiescence unveils a novel mutational force in fission yeast

eLife ◽

10.7554/elife.27469 ◽

2017 ◽

Vol 6 ◽

Cited By ~ 12

Author(s):

Serge Gangloff ◽

Guillaume Achaz ◽

Stefania Francesconi ◽

Adrien Villain ◽

Samia Miled ◽

...

Keyword(s):

Cell Division ◽

Fission Yeast ◽

Spontaneous Mutation ◽

Whole Genome ◽

The Novel ◽

Single Nucleotide Variants ◽

Spontaneous Mutation Rate ◽

Single Nucleotide ◽

Phenotypic Variations ◽

Genome Level

To maintain life across a fluctuating environment, cells alternate between phases of cell division and quiescence. During cell division, the spontaneous mutation rate is expressed as the probability of mutations per generation (Luria and Delbrück, 1943; Lea and Coulson, 1949), whereas during quiescence it will be expressed per unit of time. In this study, we report that during quiescence, the unicellular haploid fission yeast accumulates mutations as a linear function of time. The novel mutational landscape of quiescence is characterized by insertion/deletion (indels) accumulating as fast as single nucleotide variants (SNVs), and elevated amounts of deletions. When we extended the study to 3 months of quiescence, we confirmed the replication-independent mutational spectrum at the whole-genome level of a clonally aged population and uncovered phenotypic variations that subject the cells to natural selection. Thus, our results support the idea that genomes continuously evolve under two alternating phases that will impact on their size and composition.

Download Full-text