Festuca pratensis (meadow fescue) as the most frost-tolerant species within the Lolium-Festuca complex was used as a model for research aimed at identifying the cellular components involved in the cold acclimation (CA) of forage grasses. The work presented here also comprises the first comprehensive proteomic research on CA in a group of monocotyledonous species which are able to withstand winter conditions. Individual F. pratensis plants with contrasting levels of frost tolerance, high frost tolerant (HFT) and low frost tolerant (LFT) plants, were selected for comparative proteomic research. The work focused on the analysis of leaf protein accumulation before and after 2, 8, and 26 h, and 3, 5, 7, 14, and 21 d of CA, using high-throughput two-dimensional electrophoresis, and on the identification of proteins which were accumulated differentially between the selected plants by the application of mass spectrometry. The analyses of approximately 800 protein profiles revealed a total of 41 (5.1%) proteins that showed a minimum of a 1.5-fold difference in abundance, at a minimum of one time point of CA for HFT and LFT genotypes. It was shown that significant differences in profiles of protein accumulation between the analysed plants appeared relatively early during cold acclimation, most often after 26 h (on the 2nd day) of CA and one-half of the differentially accumulated proteins were all parts of the photosynthetic apparatus. Several proteins identified here have been reported to be differentially accumulated during cold conditions for the first time in this paper. The functions of the selected proteins in plant cells and their probable influence on the level of frost tolerance in F. pratensis, are discussed.

Festulolium hybrids are being increasingly used worldwide as forage grasses. This is due to their superior agronomic characteristics, which combine yield performance of ryegrasses (Lolium multiflorum and L. perenne) and tolerance against abiotic stress of fescues (Festuca pratensis, F. arundinacea and F. arundinacea var. glaucescens). Despite the widespread use, only fragmentary information exists on their genomic constitution. We used genomic in situ hybridization (GISH) to analyze genomic constitution of over 600 plants from almost all commercially available cultivars of Festulolium. Our results revealed a surprisingly large range of variation in the proportions of parental genomes and in the extent of intergenomic recombination. Using fluorescence in situ hybridization (FISH) with probes for ribosomal DNA, we assessed the frequency of recombination and elimination of particular chromosomes and chromosome groups in three contrasting Festulolium cultivars. This study provides novel information that will aid in understanding the relationship between a genetic make-up and the phenotype of Festulolium hybrids. Our results indicate that GISH might be a useful tool to aid in Festulolium breeding and provide data for a more detailed description of registered cultivars.

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Ghesquiere, M., Humphreys, M.W., Zwierzykowski, Z. (2010). Festulolium. In: ' Eucarpia Fodder Crops and Amenity Grasses (Handbook on Plant Breeding )', Veronesi, F., Posselt, U., Beart, B. (Eds). , pp. 293-316 RONO: 1310 3001Festulolium refers to natural or synthetic intergeneric hybrids between obligate outbreeding species of the Festuca (fescue) and Lolium (ryegrass) genera, species considered frequently as ideal components of agricultural or turf-grass systems. Intermediate forms between the two genera have long been recognized in nature and considered as hybrids (? Festulolium spp.) by taxonomists (e.g., Hubbard 1992) mostly on the basis of the inflorescence shape and their suspected progenitor species? combinations

The phylogeny of Festuca arundinacea Schreb. (2n = 6x = 42) was determined using GISH. Total genomic DNA of putative ancestral species was labelled with rhodamine and hybridized to chromosome preparations of hybrids involving these species and F. arundinacea. The degree of hybridization to chromosomes known to be homologous to the probe DNA was compared with that found simultaneously on chromosomes of the genome of F. arundinacea. It was concluded that the tetraploid species Festuca arundinacea var. glaucescens contributed two genomes and the diploid species Festuca pratensis one, to create the allohexaploid species F. arundinacea.Peer reviewe

Festuca arundinacea is a drought tolerant species. Lolium multiflorum has better forage quality but lower tolerance to abiotic stresses. Their hybrids offer an opportunity to perform research on the molecular basis of tolerance to drought. The aim of this work was to recognise the mechanisms of response to short-term drought (11 days) in a glasshouse in two L. multiflorum/F. arundinacea introgression forms with distinct levels of tolerance to long-term drought (14 weeks) in the field. Measurements of physiological parameters, analyses of protein accumulation profiles using two-dimensional gel electrophoresis, and mass spectrometry identification of proteins, which were accumulated differentially between the selected genotypes during short-term drought, were performed. Genotype 7/6, with lower yield potential during 14 weeks of drought, and lower ability to re-grow after watering, had a higher capacity for photosynthesis during 11 days of drought. Genotype 4/10, more tolerant to long-term drought, was able to repair damaged cell membranes after watering and was also characterised by lower transpiration during short-term drought. A total of 455 proteins were analysed, and the 17 that were differentially accumulated between the two genotypes were identified. The results of physiological and proteomic research led to a hypothesis that the higher photosynthetic capacity of genotype 7/6 could be due to a more efficient Calvin cycle, supported by higher accumulation of crucial proteins involving chloroplast aldolase.

Summary• Mechanisms of photosynthetic acclimation to cold were investigated on androgenic plants generated from Festuca pratensis × Lolium multiflorum (4x) cultivars Felopa and Sulino and on parental material.• Photosynthetic acclimation and resistance to high-light induced inactivation of PSII at low temperature were studied using chlorophyll fluorescence techniques in relation to winter hardiness, frost resistance and cold acclimation in field and controlled conditions.• In the field increased energy dissipation before winter through a lower maximum quantum yield of PSII was correlated with improved winter survival of these genotypes. In controlled conditions winter hardy plants were more resistant to cold-induced inactivation of PSII. During cold acclimation of winter hardy plants nonphotochemical quenching (NPQ) increased, except in one genotype where photochemical quenching increased.• The use of androgenic lines revealed gene combinations that determined alternative photoinhibition avoidance mechanisms in the parental genome. Increased dissipation of light energy is an alternative process to the increased photosynthetic capacity reported previously to be the main mechanism of photosynthetic acclimation to cold in herbaceous Poaceae . AbbreviationsF 0 , fluorescence of leaves in the dark when all PSII reaction centres are open; , fluorescence in leaves previously exposed to light darkened just before measurement; F v , variable fluorescence ( F v = F m − F 0 ); F m and , fluorescence when all PSII reaction centres are closed in dark-and light-exposed leaves, respectively; F s , steady state fluorescence in light exposed leaves; F v : F m , maximum quantum yield of PSII; PPFD, photosynthetic photon flux density; HL, high-light treatment, RT 50 , temperature causing a 50% reduction in re-growth rate after freezing; PSII, photosystem II; φ PSII current quantum yield of PSII; NPQ, nonphotochemical quenching of chlorophyll a fluorescence; q P , photochemical quenching of chlorophyll a fluorescence; HF and HS followed by a number are the androgenic genotypes derived from F. pratensis ×

In the allotetraploid, Festuca pratensis Huds. (2n = 4x = 28) x Lolium perenne L. (2n = 4x = 28) the balance of chromatin, as determined by GISH, changes over successive generations of open pollination in favour of L. perenne. There is extensive recombination between chromosomes of the two parental genomes, as well as substitution of whole Festuca chromosomes by whole Lolium chromosomes. The total number of Lolium chromosomes increased from a mean 14.36 in the F(2) to 16.26 in the F(6), and the total number of Festuca chromosomes decreased correspondingly from a mean of 13.57 to a value of 11.56. The number of recombinant chromosomes and recombination breakpoints per genotype also increased from generation to generation, although the respective values of both characters were higher for Festuca (0.86-8.41 and 1.14-15.22) than for Lolium (0.68-4.59 and 0.68-6.0). The proportion of total genome length contributed by the L. perenne chromatin increased from about 50% in F(2) to 59.5% in F(6). The results are based on the sample of 134 plants studied (26-28 plants per generation), and are discussed in terms of the dominance of Lolium chromosomes over those of Festuca, and possible mechanisms underlying this phenomenon of chromatin substitution.