The commitment of barley microspores into embryogenesis correlates with miRNA-directed regulation of members of the SPL, GRF and HD-ZIPIII transcription factor families.

Microspore embryogenesis is a model for developmental plasticity and cell fate decisions. To investigate the role of miRNAs in this development, we sequenced sRNAs and the degradome of barley microspores collected prior to (day 0) and after (days 2 and 5) the application of a stress treatment known to induce embryogenesis. Microspores isolated at these timepoints were uniform in both appearance and in their complements of sRNAs. We detected 68 miRNAs in microspores. The abundance of 51 of these miRNAs differed significantly during microspore development. One group of miRNAs was induced when the stress treatment was applied, prior to being repressed when microspores transitioned to embryogenesis. Another group of miRNAs were up-regulated in day-2 microspores and their abundance remained stable or increased in day-5 microspores, a timepoint at which the first clear indications of the transition toward embryogenesis were visible. Collectively, these miRNAs might play a role in the modulation of the stress response, the repression of gametic development, and/or the gain of embryogenic potential. A degradome analysis allowed us to validate the role of miRNAs in regulating 41 specific transcripts. We showed that the transition of microspores toward the embryogenesis pathway involves miRNA-directed regulation of members of the ARF, SPL, GRF, and HD-ZIPIII transcription factor families. We noted that 41.5% of these targets were shared between day-2 and day-5 microspores while 26.8% were unique to day-5 microspores. The former set may act to disrupt transcripts involved in pollen development while the latter set may drive the commitment to embryogenesis.

How Much Knowledge is Worth Knowing? An American Intellectual Historian's Thoughts on the Geschichte des Wissens.

This essay investigates the origins and assesses the advantages and disadvantages of the new field known as Wissensgeschichte from the perspective of an American intellectual historian. It argues that while some historians of science may be ready to embrace a new identity as historians of knowledge, this terminology remains baggy and invites facile applications of Foucauldian theory. The essay concludes with the hope that the history of knowledge approach may instead open up new avenues for conversation and collaboration between historians of science and garden variety historians.

Differential Expression Profiling of Microspores During the Early Stages of Isolated Microspore Culture Using the Responsive Barley Cultivar Gobernadora.

In barley, it is possible to induce embryogenesis in the haploid and uninucleate microspore to obtain a diploid plant that is perfectly homozygous. To change developmental fates in this fashion, microspores need to engage in cellular de-differentiation, interrupting the pollen formation, and restore totipotency prior to engaging in embryogenesis. In this work, we used the barley cultivar Gobernadora to characterize the transcriptome of microspores prior to (day 0) and immediately after (days 2 and 5) the application of a stress pretreatment. A deep RNA-seq analysis revealed that microspores at these three time points exhibit a transcriptome of ∼14k genes, ∼90% of which were shared. An expression analysis identified a total of 3,382 differentially expressed genes (DEGs); of these, 2,155 and 2,281 DEGs were respectively identified when contrasting expression at days 0 and 2 and at days 2 and 5. These define 8 expression profiles in which DEGs share a common up- or down-regulation at these time points. Up-regulation of numerous glutathione S-transferase and heat shock protein genes as well as down-regulation of ribosomal subunit protein genes was observed between days 0 and 2. The transition from microspores to developing embryos (days 2 vs. 5) was marked by the induction of transcription factor genes known to play important roles in early embryogenesis, numerous genes involved in hormone biosynthesis and plant hormonal signal transduction in addition to genes involved in secondary metabolism. This work sheds light on transcriptional changes accompanying an important developmental shift and provides candidate biomarkers for embryogenesis in barley.

Improving the efficiency of isolated microspore culture in six-row spring barley: II-exploring novel growth regulators to maximize embryogenesis and reduce albinism.

Two alternative cytokinins, thidiazuron and meta-topoline, were tested in isolated microspore culture on recalcitrant barley genotypes (six-row, spring), and green plant regeneration was improved substantially. Doubled-haploid (DH) plants are coveted in plant breeding and in genetic studies, since they are rapidly obtained and perfectly homozygous. In barley, DHs are produced mainly via androgenesis, and isolated microspore culture (IMC) constitutes the method offering the greatest potential efficiency. However, IMC can often be challenging in some genotypes because of low yield of microspores, low regeneration and high incidence of albinism. Six-row spring-type barleys, the predominant type grown in Eastern Canada, are considered recalcitrant in this regard. Our general objective was to optimize an IMC protocol for DH production in six-row spring barley. In particular, we explored the use of alternative hormones in the induction medium (thidiazuron and dicamba), and in the regeneration medium (meta-topoline). This optimization was performed on two typical six-row spring (ACCA and Léger), a two-row spring (Gobernadora) and a two-row winter (Igri) barley cultivar. When 6-benzyl-aminopurine (BAP) was replaced by a combination of thidiazuron and dicamba in the induction medium, a 5.1-fold increase (P < 0.01) in the production of green plants resulted. This increase was mainly achieved by a reduction of albinism. Moreover, a 2.9-fold increase (P < 0.01) in embryo differentiation into green plants was obtained using meta-topoline instead of BAP in the regeneration medium. Together, these innovations allowed us to achieve a substantial improvement in the efficiency of IMC in this recalcitrant type of barley. These results were later successfully validated using sets of F1s from a six-row spring barley breeding program.

Comparative analysis of genetic diversity in Canadian barley assessed by SSR, DarT, and pedigree data.

The aim of this study was to measure genetic diversity and population structure among 92 Canadian barley cultivars using two types of molecular markers (SSRs and DArTs) and pedigree data. A total of 368 alleles were identified at 50 SSR loci. The number of alleles per locus ranged between 2 and 13 ([Formula: see text] = 7.36) and PIC values ranged from 0.34 to 0.86 ([Formula: see text] = 0.69). For the biallelic DArT markers, the genetic distance matrix was based on 971 markers whose PIC values ranged between 0.06 and 0.50 ([Formula: see text] = 0.39). A third distance matrix was computed based on the kinship coefficient. Clustering of genotypes was performed based on the genetic distance matrix and the three dendrograms obtained showed the genetic relationships among barley cultivars. The topological similarity of the three dendrograms was estimated using a congruence index and showed the three dendrograms to be in very good agreement. Statistical analysis also showed a highly significant correlation between the SSR and DArT matrices (r = 0.80, p < 0.002) compared with lower yet significant correlations of the pedigree data with both marker types (r = 0.46, p < 0.002; r = 0.52, p < 0.002). Finally, we assessed linkage disequilibrium in this germplasm and found it to be quite extensive, as the mean distance between marker pairs with significant (P < 0.001) r(2) values >0.5 was 3.8 cM. Information obtained from comparing results of different genetic diversity estimation methods should be useful for the improvement and conservation of barley genetic resources.

Population structure and linkage disequilibrium in barley assessed by DArT markers.

Diversity Array Technology (DArT) markers were used to investigate the genetic diversity, population structure, and extent of linkage disequilibrium (LD) on a genome-wide level in Canadian barley (Hordeum vulgare L.). Approximately 1,000 DArT markers were polymorphic and scored with high confidence among a collection of 170 barley lines composed mostly of Canadian cultivars and breeding lines. The reproducibility of DArT markers proved very high, as 99.9% of allele calls were identical among seven replicated samples. The polymorphism information content (PIC) of DArT markers ranged between 0.04 and 0.50 with an average of 0.38. Using principal coordinate analysis (PCoA), most lines fell into one of two major groups reflecting inflorescence type (two-row versus six-row). Within these two large groups, evidence of geographic clustering of genotypes was also observed. A cluster analysis Unweighted Pair Group Method with Algorithmic Mean suggested the existence of three subgroups within the two-row group and four subgroups within the six-row group. An analysis of molecular variance (AMOVA) revealed highly significant (P < 0.001) genetic variance within subgroups, among subgroups, and among groups. Values of LD, expressed as r (2), declined with increasing genetic distance, and mean values of r (2) fell below 0.2 for markers located 2.6 cM apart. Approximately 8% of marker pairs located on the same chromosome and 3.4% of pairs located on different chromosomes were in LD (r ( 2 ) > 0.2). Within both the subsets of two-row and six-row lines, LD extended slightly further (3.5 cM) than for the entire set, while 7.5% of intra-chromosomal locus pairs and <2% of inter-chromosomal pairs were in LD. We discuss the implications of these findings with regard to the prospects of association mapping of complex traits in barley.

Androgenic response of barley accessions and F1s with Fusarium head blight resistance.

Most Fusarium head blight (FHB) resistant barley (Hordeum vulgare L.) accessions perform relatively poorly from an agronomic point of view. Due to the polygenic inheritance of FHB resistance, introgression of this complex trait into well-adapted elite germplasm will likely require multiple cycles of hybridization and selection to combine resistance and agronomic performance. The use of anther culture to produce doubled haploids would seem well justified to reduce the time required to achieve this goal. Unfortunately, little is known concerning the androgenic response of the small number of genotypes with known partial FHB resistance. To make the best use of such FHB resistance donors in a barley improvement program, we first characterized the FHB resistance of eight reported FHB resistance sources (Chevron, Gobernadora, Seijo II, Shyri, Svanhals, Zhedar I, F104-250-9 and C97-21-38-3) in our own FHB nursery in Quebec City (QC, Canada). In parallel, we assessed the androgenic response of these same eight lines with that of three cultivars (ACCA, Léger and Cadette) of known androgenic response. Finally, the androgenic response of F(1) hybrids involving some of these genotypes used as parents was measured and compared to that of the parental genotypes. Very large and significant differences were observed in the number of green plants produced by the different accessions and F(1)s. Although anther culture seemed very promising for some accessions, for others, the androgenic response was so low that a conventional approach would seem more appropriate.