Barley Seeds miRNome Stability during Long-Term Storage and Aging.

Seed aging is a complex biological process that has been attracting scientists' attention for many years. High-throughput small RNA sequencing was applied to examine microRNAs contribution in barley seeds senescence. Unique samples of seeds that, despite having the same genetic makeup, differed in viability after over 45 years of storage in a dry state were investigated. In total, 61 known and 81 novel miRNA were identified in dry seeds. The highest level of expression was found in four conserved miRNA families, i.e., miR159, miR156, miR166, and miR168. However, the most astonishing result was the lack of significant differences in the level of almost all miRNAs in seed samples with significantly different viability. This result reveals that miRNAs in dry seeds are extremely stable. This is also the first identified RNA fraction that is not deteriorating along with the loss of seed viability. Moreover, the novel miRNA hvu-new41, with higher expression in seeds with the lowest viability as detected by RT-qPCR, has the potential to become an indicator of the decreasing viability of seeds during storage in a dry state.

Low RIN Value for RNA-Seq Library Construction from Long-Term Stored Seeds: A Case Study of Barley Seeds.

Seed aging is a complex biological process and its fundamentals and mechanisms have not yet been fully recognized. This is a key issue faced by research teams involved in the collection and storage of plant genetic resources in gene banks every day. Transcriptomic changes associated with seed aging in the dry state have barely been studied. The aim of the study was to develop an efficient protocol for construction of RNA-Seq libraries from long-term stored seeds with very low viability and low RNA integrity number (RIN). Here, barley seeds that have almost completely lost their viability as a result of long-term storage were used. As a control, fully viable seeds obtained in the course of field regeneration were used. The effectiveness of protocols dedicated to RNA samples with high and low RIN values was compared. The experiment concluded that library construction from low viable or long-term stored seeds with degraded RNA (RIN < 3) should be carried out with extraordinary attention due to the possibility of uneven degradation of different RNA fractions.

A multilevel exploration of Avena strigosa diversity as a prelude to promote alternative crop.

BACKGROUND: Sand oat (Avena strigosa Schreb.), one of the four cultivated species of the genus Avena, could be considered as another alternative crop. In gene banks 865 germplasm samples of this species have been preserved that have not been thoroughly investigated so far. The results of phenotyping (36 traits), isoenzymatic (12 systems) and genetic (8 pairs of Sequence-related amplified polymorphism markers) variation were used to obtain the complete description of 56 accessions diversity originated from different parts of world. RESULTS: Breeded and weedy forms represented similar pool of morphological traits that indicated a short-term and extensive breeding process, albeit all accessions which we classified as cultivated were characterized by better grain and green mass parameters compared to the weedy ones. Isoenzymes showed relationships with geographical origin, which was not possible to detect by SRAP markers. There was no similarity between morphological and biochemical results. The polymorphism level of SRAP markers was lower than indicated by the available literature data for other species, however it may result from the analysis of pooled samples of accessions with a high internal variability. The extensive type of breeding and its relatively short duration was also reflected in the population structure results. Joint analysis revealed that a secondary centre of diversity is being created in South America and that it has its genealogy from the Iberian Peninsula. CONCLUSIONS: Despite the relatively large representation of this species is in various gene banks, it is highly probable that the vast majority of stored worldwide accessions are duplicates, and the protected gene pool is relatively narrow. Sand oat meets all the requirements for an alternative crop species, but further studies are needed to identify the genotypes/populations with the most favourable distribution of utility and quality parameters.

Differences in Ear Rot Resistance and Fusarium verticillioides-Produced Fumonisin Contamination Between Polish Currently and Historically Used Maize Inbred Lines.

Poland is the fifth largest European country, in terms of maize production. Ear rots caused by Fusarium spp. are significant diseases affecting yield and causing grain mycotoxin contamination. Inbred lines, which are commonly used in Polish breeding programs, belong, mostly, to two distinct genetic categories: flint and dent. However, historically used lines belonging to the heterotic Lancaster, IDT and SSS groups were also present in previous Polish breeding programs. In the current study, 98 inbred lines were evaluated across a 2-year-long experiment, after inoculation with F. verticillioides and under natural infection conditions. Lancaster, IDT, SSS and SSS/IDT groups were characterized as the most susceptible ones and flint as the more resistant. Based on the results obtained, the moderately resistant and most susceptible genotypes were defined to determine the content of fumonisins (FBs) in kernel and cob fractions using the HPLC method. Fumonisin's content was higher in the grain samples collected from inoculated plants than in cobs. The association of visible Fusarium symptoms with fumonisin concentration in grain samples was significant. Conversely, the cobs contained more FB1 under natural infection, which may be related to a pathogen's type of growth, infection time or presence of competitive species. Using ddRADseq genome sampling method it was possible to distinguish a basal relationship between moderately resistant and susceptible genotypes. Genetic distance between maize genotypes was high. Moderately resistant inbreed lines, which belong to IDT and IDT/SSS belong to one haplotype. Genotypes which belong to the flint, dent or Lancaster group, and were characterized as moderately resistant were classified separately as the same susceptible one. This research has demonstrated that currently grown Polish inbred lines, as well the ones used in the past are a valid source of resistance to Fusarium ear rot. A strong association was observed between visible Fusarium symptoms with fumonisin concentration in grain samples, suggesting that selection in maize for reduced visible molds should reduce the risk of mycotoxin contamination. NGS techniques provide new tools for overcoming the long selection process and increase the breeding efficiency.