Improving the time and space complexity of the WFA algorithm and generalizing its scoring

Modern genomic sequencing data is trending toward longer sequences with higher accuracy. Many analyses using these data will center on alignments, but classical exact alignment algorithms are infeasible for long sequences. The recently proposed WFA algorithm demonstrated how to perform exact alignment for long, similar sequences in O(sN) time and O(s2) memory, where s is a score that is low for similar sequences (Marco-Sola et al., 2021). However, this algorithm still has infeasible memory requirements for longer sequences. Also, it uses an alternate scoring system that is unfamiliar to many bioinformaticians. We describe variants of WFA that improve its asymptotic memory use from O(s2) to O(s3/2) and its asymptotic run time from O(sN) to O(s2 + N). We expect the reduction in memory use to be particularly impactful, as it makes it practical to perform highly multithreaded megabase-scale exact alignments in common compute environments. In addition, we show how to fold WFA's alternate scoring into the broader literature on alignment scores.

Chromosome-scale assembly of the highly heterozygous genome of red clover (Trifolium pratense L.), an allogamous forage crop species

Red clover (Trifolium pratense L.) is used as a forage crop due to a variety of favorable traits relative to other crops. Improved varieties have been developed through conventional breeding approaches, but progress could be accelerated and gene discovery facilitated using modern genomic methods. Existing short-read based genome assemblies of the ~420 Megabase (Mb) genome are fragmented into >135,000 contigs with numerous errors in order and orientation within scaffolds, likely due to the biology of the plant which displays gametophytic self-incompatibility resulting in inherent high heterozygosity. A high-quality long-read based assembly of red clover is presented that reduces the number of contigs by more than 500-fold, improves the per-base quality, and increases the contig N50 statistic by three orders of magnitude. The 413.5 Mb assembly is nearly 20% longer than the 350 Mb short read assembly, closer to the predicted genome size. Quality measures are presented and full-length isoform sequence of RNA transcripts reported for use in assessing accuracy and for future annotation of the genome. The assembly accurately represents the seven main linkage groups present in the genome of an allogamous (outcrossing), highly heterozygous plant species.

Envisaging an Effective Global Long-Term Agrobiodiversity Conservation System That Promotes and Facilitates Use

Genebanks were established out of a recognised need not just to provide genetic variation to support breeding objectives but to prevent crop diversity from being lost entirely for future users. Such conservation objectives may have led, over the past few decades, to a gradually diminishing connection between genebanks and current users of diversity. While there continues to be large-scale distribution of germplasm from genebanks to recipients worldwide, relatively little is known or published about the detailed trends in the demand for genebank materials. Meanwhile, the rapid expansion of the applications and uses of modern genomic technologies and approaches is, undoubtedly, having a transformational impact on breeding, research and the demand for certain genetic resources and associated data. These trends will require genebanks to be responsive and to adapt. They also provide important opportunities for genebanks to reorganize and become more efficient individually and as a community. Ultimately, future challenges and opportunities are likely to drive more demand for genetic diversity and provide an important basis for genebanks to gear up.

OTC intron 4 variations mediate pathogenic splicing patterns caused by the c.386G>A mutation in humans and spfash mice, and govern susceptibility to RNA-based therapies

Background Aberrant splicing is a common outcome in the presence of exonic or intronic variants that might hamper the intricate network of interactions defining an exon in a specific gene context. Therefore, the evaluation of the functional, and potentially pathological, role of nucleotide changes remains one of the major challenges in the modern genomic era. This aspect has also to be taken into account during the pre-clinical evaluation of innovative therapeutic approaches in animal models of human diseases. This is of particular relevance when developing therapeutics acting on splicing, an intriguing and expanding research area for several disorders. Here, we addressed species-specific splicing mechanisms triggered by the OTC c.386G>A mutation, relatively frequent in humans, leading to Ornithine TransCarbamylase Deficiency (OTCD) in patients and spfash mice, and its differential susceptibility to RNA therapeutics based on engineered U1snRNA. Methods Creation and co-expression of engineered U1snRNAs with human and mouse minigenes, either wild-type or harbouring different nucleotide changes, in human (HepG2) and mouse (Hepa1-6) hepatoma cells followed by analysis of splicing pattern. RNA pulldown studies to evaluate binding of specific splicing factors. Results Comparative nucleotide analysis suggested a role for the intronic +10-11 nucleotides, and pull-down assays showed that they confer preferential binding to the TIA1 splicing factor in the mouse context, where TIA1 overexpression further increases correct splicing. Consistently, the splicing profile of the human minigene with mouse +10-11 nucleotides overlapped that of mouse minigene, and restored responsiveness to TIA1 overexpression and to compensatory U1snRNA. Swapping the human +10-11 nucleotides into the mouse context had opposite effects. Moreover, the interplay between the authentic and the adjacent cryptic 5′ss in the human OTC dictates pathogenic mechanisms of several OTCD-causing 5′ss mutations, and only the c.386+5G>A change, abrogating the cryptic 5′ss, was rescuable by engineered U1snRNA. Conclusions Subtle intronic variations explain species-specific OTC splicing patterns driven by the c.386G>A mutation, and the responsiveness to engineered U1snRNAs, which suggests careful elucidation of molecular mechanisms before proposing translation of tailored therapeutics from animal models to humans.

LncRNAs LY86-AS1 and VIM-AS1 Distinguish Plasma Cell Leukemia Patients from Multiple Myeloma Patients

Long non-coding RNAs (lncRNAs) are functional RNAs longer than 200 nucleotides. Due to modern genomic techniques, the involvement of lncRNAs in tumorigenesis has been revealed; however, information concerning lncRNA interplay in multiple myeloma (MM) and plasma cell leukemia (PCL) is virtually absent. Herein, we aimed to identify the lncRNAs involved in MM to PCL progression. We investigated representative datasets of MM and PCL patients using next-generation sequencing. In total, 13 deregulated lncRNAs (p < 0.00025) were identified; four of them were chosen for further validation in an independent set of MM and PCL patients by RT-qPCR. The obtained results proved the significant downregulation of lymphocyte antigen antisense RNA 1 (LY86-AS1) and VIM antisense RNA 1 (VIM-AS1) in PCL compared to MM. Importantly, these two lncRNAs could be involved in the progression of MM into PCL; thus, they could serve as promising novel biomarkers of MM progression.

Genic microsatellite marker characterization and development in little millet (Panicum sumatrense) using transcriptome sequencing

Little millet is a climate-resilient and high-nutrient value plant. The lack of molecular markers severely limits the adoption of modern genomic approaches in millet breeding studies. Here the transcriptome of three samples were sequenced. A total of 4443 genic-SSR motifs were identified in 30,220 unigene sequences. SSRs were found at a rate of 12.25 percent, with an average of one SSR locus per 10 kb. Among different repeat motifs, tri-nucleotide repeat (66.67) was the most abundant one, followed by di- (27.39P), and tetra- (3.83P) repeats. CDS contained fewer motifs with the majority of tri-nucleotides, while 3′ and 5′ UTR carry more motifs but have shorter repeats. Functional annotation of unigenes containing microsatellites, revealed that most of them were linked to metabolism, gene expression regulation, and response to environmental stresses. Fifty primers were randomly chosen and validated in five little millet and 20 minor millet genotypes; 48% showed polymorphism, with a high transferability (70%) rate. Identified microsatellites can be a noteworthy resource for future research into QTL-based breeding, genetic resource conservation, MAS selection, and evolutionary genetics.

Experience of using modern genomic technologies to study microorganisms and their communities

Objective. The aim of this work was to review the main results of genomic studies of microorganisms and their communities performed on the base of the Research Laboratory of Gomel State Medical University.Materials and methods. Genomic, transcriptomic and metagenomic analysis of the microorganisms of the stomach and respiratory tract.Results. The capabilities of modern next-generation sequencing platforms have been analyzed, and the authors` own results of the use of genomic, transcriptomic and metagenomic analysis of the microbiota in patients with various gastric and respiratory pathologies have been described.Conclusion. The analysis of the obtained data has revealed peculiarities of the structure of the microbial communities of the stomach of the patients infected with H. pylori with different gastric pathology: the proportion participation of H. pylori in the metagenome of the samples with different forms of gastric cancer did not exceed 25 %, of gastritis — 6 %, of peptic ulcer — 1 %. At the same time, the minimal amount of H. pylori in all the cases could reach 0.1 %. A signifcant degree of CagA and CagY loci variability of H. pylori was detected. Streptoccocus genus bacteria dominated (36 %) in the bacterial microbiome of a patient diagnosed with the coronavirus disease, and in the viral microbiome, SARS-CoV-2 constituted 59 % of the total number of viruses in the material. The analysis of 13 strains of Klebsiella pneumoniae with multiple and extreme resistance to antibiotics has found that the studied strains belong to fve MLST-types, three of which are classifed as high epidemic risk groups.

Seq Your Destiny: Neural Crest Fate Determination in the Genomic Era

Neural crest stem/progenitor cells arise early during vertebrate embryogenesis at the border of the forming central nervous system. They subsequently migrate throughout the body, eventually differentiating into diverse cell types ranging from neurons and glia of the peripheral nervous system to bones of the face, portions of the heart, and pigmentation of the skin. Along the body axis, the neural crest is heterogeneous, with different subpopulations arising in the head, neck, trunk, and tail regions, each characterized by distinct migratory patterns and developmental potential. Modern genomic approaches like single-cell RNA- and ATAC-sequencing (seq) have greatly enhanced our understanding of cell lineage trajectories and gene regulatory circuitry underlying the developmental progression of neural crest cells. Here, we discuss how genomic approaches have provided new insights into old questions in neural crest biology by elucidating transcriptional and posttranscriptional mechanisms that govern neural crest formation and the establishment of axial level identity. Expected final online publication date for the Annual Review of Genetics, Volume 55 is November 2021. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.

Anti-evolution Drugs: A New Paradigm to Combat Drug Resistance

: The drug resistance confronts chemotherapy of neoplasm and microbial infections. A vast array of molecular mechanisms was implicated in drug resistance, including, generation of drug efflux transporters, mutation of drug targets, and alteration of drug metabolism. With the alarming rate of increase in drug resistance, pathogens are bolstering in such a way that many new drugs face efficacy problems within a short span of entry into the market. Evolution is the driving force towards the development of drug resistance. By adopting the modern genomic and functionomic analytical techniques, scientists have now identified novel genes and signalling proteins involved in the evolution of drug resistance in microorganisms. Given the current knowledge of bacterial evolution, the antibiotic drug discovery is ready for a paradigm shift to explore the newer ways to tackle drug resistance. The article discusses such recent developments and reviews their merits and demerits in an attempt to envisage the findings in this new domain of medicine.