Patricio Esteves

Les plantes haploides doublees (HD) sont des individus completement homozygotes qui peuvent etre obtenus via l’androgenese in vitro. Les HD sont tres apprecies comme outils de recherche en genetique et en amelioration vegetale. La culture de microspores isolees (CMI) est le moyen le plus efficace de produire des HD. Malheureusement, les orges de printemps a six rangs, le type predominant cultive dans l’Est du Canada, sont considerees comme recalcitrantes a cause d’une faible embryogenese et d’une propension a l’albinisme. Notre objectif etait de developper un protocole CMI adapte a ce type d’orge. Nous avons entame des travaux exploratoires sur quatre cultivars : ACCA et Leger (six rangs, printemps), Gobernadora (deux rangs, printemps) et Igri (deux rangs, hiver). Dans une premiere phase, nous avons evalue l’impact de quatre facteurs physiques. Nous avons trouve que l’optimisation du stade de recolte des tiges pouvait augmenter de 2 a 4 fois la recolte de microspores embryogeniques....

Isolated microspore culture (IMC) is the most efficient way to produce large numbers of doubled-haploid (DH) barley plants in a short time. Yet, while IMC is more cost-efficient and less labor-intensive than anther culture, it is technically more complex and requires more experienced personnel if it is to yield its full potential. In part, this is because of multiple and important interactions that exist between factors at its many different phases, including genotype effects as well. When every phase is fine-tuned, the protocol that is presented below yields a useful number of DHs with almost all genotypes and can allow the production of up to 300 DH plants from a single F1 plant in just a few months.

Gametic embryogenesis is defined as the process that allows immature pollen grains – microspores – to parthenogenetically become embryos. The process can successfully be induced at a high frequency under in vitro culture conditions in a wide number of crop species. Microspores are haploid cells carrying half the somatic number of chromosomes, but if either spontaneously or artificially their chromosomal complement is doubled during the embryogenic pathway, the resulting embryos will become diploid and perfectly homozygous. The products of gametic embryogenesis are therefore called doubled-haploid plants, which are coveted materials for research and for plant breeding. Yet, to be efficiently used in a plant breeding program, doubled haploids need to be produced at a high frequency and in a reproducible manner. The efficiency and reproducibility of DH production are tied to the control of key factors intervening in the process. As is the case in many in vitro procedures, growth regula...

Extent and overlap of segregation distortion regions in 12 barley crosses determined via a Pool-GBS approach. Segregation distortion is undesirable as it alters the frequency of alleles and can reduce the chances of obtaining a particular combination of alleles. In this work, we have used a pooled genotyping-by-sequencing (Pool-GBS) approach to estimate allelic frequencies and used it to examine segregation distortion in 12 segregating populations of barley derived from androgenesis. Thanks to the extensive genome-wide SNP coverage achieved (between 674 and 1744 markers), we determined that the proportion of distorted markers averaged 28.9 % while 25.3 % of the genetic map fell within segregation distortion regions (SDRs). These SDRs were characterized and identified based on the position of the marker showing the largest distortion and the span of each SDR. Summed across all 12 crosses, 36 different SDR peaks could be distinguished from a total of 50 SDRs and a majority of these SDRs (27 of 36) were observed in only one population. While most shared SDRs were common to only two crosses, two SDRs (SDR3.1 and SDR4.2) were exceptionally recurrent (seen in five and four crosses, respectively). Because of the broad span of most SDRs, an average of 30 % of crosses showed segregation distortion in any given chromosomal segment. In reciprocal crosses, although some SDRs were clearly shared, others were unique to a single direction. In summary, segregation distortion is highly variable in its extent and the number of loci underpinning these distortions seems to be quite large even in a narrow germplasm such as six-row spring barley.

SUMMARYMicrospore 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 towards embryogenesis were visible. Collectively, these miRNAs might play a role in ...