Lesley B. Turner

Synteny between a major heading-date QTL in perennial ryegrass (Lolium perenne L.) and the Hd3 heading-date locus in rice

The genetic control of induction to flowering has been studied extensively in both model and crop species because of its fundamental biological and economic significance. An ultimate aim of many of these studies has been the application of the understanding of control of flowering that can be gained from the study of model species, to the improvement of crop species. The present study identifies a region of genetic synteny between rice and Lolium perenne, which contains the Hd3 heading-date QTL in rice and a major QTL, accounting for up to 70% of the variance associated with heading date in L. perenne. The identification of synteny between rice and L. perenne in this region demonstrates the direct applicability of the rice genome to the understanding of biological processes in other species. Specifically, this syntenic relationship will greatly facilitate the genetic dissection of aspects of heading-date induction by enabling the magnitude of the genetic component of the heading-date QTL in L. perenne to be combined with the sequencing and annotation information from the rice genome.

Light-induced isomerization and dimerization of cinnamic acid derivatives in cell walls

Abstract The role of light in the isomerization and cyclodimerization of the cinnamic acid component of plant cell walls has been examined in a number of ways. Cinnamic acid derivatives were extracted from neutral detergent fibre cell walls by alkaline hydrolysis and quantified by GC. Amounts of these acids in leaf and stem tissues of barley straw which had naturally been exposed to very different levels of radiation were compared. Leaf cell walls contained less p-coumaric acid than stem cell walls, but a higher proportion of this was found as the (Z)-isomer and cyclodimer (4,4′-dihydroxy-α-truxillic acid). Samples of wheat straw from a sequence of 10 consecutive years, grown under conditions where meteorological conditions were the major variable, showed considerable year-to-year differences; the (Z)-/(E)-ratio and dimer/monomer ratio varied three-fold. Additional studies compared the direct effect of light on the cinnamic acid derivatives in growing barley seedlings (live), freeze-dried seedlings (dead) and freshly harvested straw samples (dead). There were no significant changes in the cinnamic acid derivatives extracted from the cell walls of dead tissue which had been stored in the dark. Storage in light resulted in increased (Z)-isomer and cyclodimer contents. These changes occurred whether the tissue was alive or dead. They can, therefore, be directly attributed to the effect of light, and do not appear to be mediated by any enzymic reactions.

Comparison of growth and carbohydrate accumulation in seedlings of two varieties of Lolium perenne

Turner, L. B., Humphreys, M. O., Cairns, A. J., Pollock, C. J. (2001). Comparison of growth and carbohydrate accumulation in seedlings of two varieties of Lolium perenne. Journal of Plant Physiology, 158, (7), 891-897. Sponsorship: BBSRC RASP Initiative

Identifying genetic components controlling fertility in the outcrossing grass species perennial ryegrass (Lolium perenne) by quantitative trait loci analysis and comparative genetics

Mutational load and resource allocation factors and their effects on limiting seed set were investigated in ryegrass by comparative mapping genomics and quantitative trait loci (QTL) analysis in two perennial ryegrass (Lolium perenne) mapping families sharing common genetic markers. Quantitative trait loci for seed-set were identified on chromosome (LG) 7 in both families and on LG4 of the F2/WSC family. On LG7, seed-set and heading date QTLs colocalized in both families and cannot be unequivocally resolved. Comparative genomics suggests that the LG7 region is syntenous to a region of rice LG6 which contains both fertility (S5(n)) and heading date (Hd1, Hd3a) candidate genes. The LG4 region is syntenous to a region of rice LG3 which contains a fertility (S33) candidate gene. QTL maxima for seed-set and heading date on LG4 in the F2/WSC family are separated by c. 8 cm, indicating distinct genetic control. Low seed set is under the control of recessive genes at both LG4 and LG7 locations. The identification of QTLs associated with seed set, a major component of seed yield in perennial ryegrass, indicates that mutational load associated with these genomic regions can be mitigated through marker-assisted selection.

Identification and mode of action of self-compatibility loci in Lolium perenne L

The two-locus gametophytic incompatibility system in perennial ryegrass (Lolium perenne L.) is not always fully effective: obligate selfing of plants sieves self-compatible pollen mutants, and self-fertility becomes fixed in subsequent generations. Self-compatibility (SC) was investigated in an F2 family. In vitro self-pollinations were analysed and recorded and plants were classified as being either partially or fully compatible. Distorted segregation ratios of markers on linkage group (LG) 5 were found, which indicate the possible presence of a gametophytic SC locus. Interval linkage analysis of pollen compatibility after selfing confirmed that this distortion was due to a locus (T) analogous to the S5 locus of rye. However, even though markers in this region were, on average, less than 1 cM apart, the minimum number of plants possessing the unfavoured allele was never less than 6% for any marker locus. We proved that this was because of the presence of another SC locus, exhibiting gametophytic selection, segregating in this population and identified by interval mapping analysis of compatibility classes of in vitro self-pollinations. This locus was located on LG1, and probably corresponds to the S locus. We show that the T locus, a relic of a multilocus system, functions through interaction with the S locus: F2 segregation of incompatibility phenotypes and linked markers demonstrated that the S/t pollen genotype combination, expected to be compatible on selfing, was sometimes incompatible. Further evidence is presented to show that this interaction must be dependent on yet another locus located on LG2. A prime candidate would be the Z incompatibility locus.

Expressed sequence tag-derived microsatellite markers of perennial ryegrass (Lolium perenne L.)

An expressed sequence tag (EST) library of the key grassland species perennial ryegrass (Lolium perenne L.) has been exploited as a resource for microsatellite marker development. Out of 955 simple sequence repeat (SSR) containing ESTs, 744 were used for primer design. Primer amplification was tested in eight genotypes of L. perenne and L. multiflorum representing (grand-) parents of four mapping populations and resulted in 464 successfully amplified EST-SSRs. Three hundred and six primer pairs successfully amplified products in the mapping population VrnA derived from two of the eight genotypes included in the original screening and revealed SSR polymorphisms for 143 ESTs. Here, we report on 464 EST-derived SSR primer sequences of perennial ryegrass established in laboratory assays, providing a dedicated tool for marker assisted breeding and comparative mapping within and among forage and turf grasses.

Starch metabolism in the fructan-grasses: Patterns of starch accumulation in excised leaves of Lolium temulentum L.

Summary Excised and illuminated leaves of temperate gramineae have previously provided a powerful system for the analysis of the regulation of partitioning of photosynthate between sucrose and fructan. This paper complements previous data by describing the concomitant accumulation of transitory starch, an assimilate which has been ignored in this system. Leaves of Lolium temulentum were depleted to near-zero reserve carbohydrate content. In this condition, the tissue contained no detectable fructan or starch and only a trace of sucrose. When subsequently excised and illuminated continuously, the accumulation of sucrose and starch commenced immediately and occurred simultaneously. Starch anabolism was constitutive and the polymer was apparently synthesised de novo . The maximal rate of starch accumulation, at 0.6 mg g −1 fresh mass, was 5-10-fold lower than that of sucrose. Fructan was also synthesised de novo and was induced after a lag of 8 h, after which rates of net sucrose and starch accumulation slowed, ceasing completely by 14-16 h. Whilst total non-structural carbohydrate concentration increased continuously, reaching 56 mg g −1 at 30 h, starch concentration was asymptotic and was limited to a maximum of only 7 mg g −1 reached at 14-16 h. Hence, net starch accumulation was not restricted by the continued production of photosynthate. The illumination period spanned 1.25 diurnal cycles. Despite continued photoassimilation, there was no resumption of net starch synthesis at the beginning of the second cycle and hence the limitation of net starch accumulation was not imposed by an endogenous circadian rhythmicity. When assimilate partitioning into fructan was abolished by transpirational feeding of cycloheximide, sucrose concentration increased 2.7-fold to 49 mg g −1 . This abnormally high concentration did not feedback to enhance starch accumulation. Transpirational feeding of 10 mmol/L mannose inhibited sucrose accumulation by c .90 %, but this inhibition was not accompanied by an increase net starch accumulation, rather, starch accumulation was inhibited by 60 %. Mannose caused the formation of maltose in the tissue. Starch synthesis, in common with sucrose and fructan syntheses, was shown to be localised predominantly in mesophyll cells where presumably the control of partitioning between these three assimilates is exercised. The low rates, restricted tissue concentration, sucrose-insensitivity and mannose-inhibition of starch synthesis differ from many species where starch is the primary reserve. Although the available information is limited, some of these features are in common with other fructan grasses. The low capacity for starch accumulation provides a possible explanation for the evolution of fructan synthesis in these species.

A novel grass hybrid to reduce flood generation in temperate regions

We report on the evaluation of a novel grass hybrid that provides efficient forage production and could help mitigate flooding. Perennial ryegrass (Lolium perenne) is the grass species of choice for most farmers, but lacks resilience against extremes of climate. We hybridised L. perenne onto a closely related and more stress-resistant grass species, meadow fescue Festuca pratensis. We demonstrate that the L. perenne × F. pratensis cultivar can reduce runoff during the events by 51% compared to a leading UK nationally recommended L. perenne cultivar and by 43% compared to F. pratensis over a two year field experiment. We present evidence that the reduced runoff from this Festulolium cultivar was due to intense initial root growth followed by rapid senescence, especially at depth. Hybrid grasses of this type show potential for reducing the likelihood of flooding, whilst providing food production under conditions of changing climate.

Breeding for Bio-ethanol Production in Lolium perenne L.: Association of Allelic Variation with High Water-Soluble Carbohydrate Content

Increasing the extractable sugar yield from perennial crops is one strategy to generate renewable fuels such as bio-ethanol. Lolium perenne L. (perennial ryegrass) can contain significant (>30% dry matter) water-soluble sugars in the form of polymeric fructan which is readily extracted, broken down and fermented to bio-ethanol. A population of L. perenne generated from four parents which differed in water-soluble carbohydrate (WSC) content was subjected to multiple rounds of selection and recombination on the basis of early spring WSC content to produce a high WSC, and a low WSC population. A control population was generated by selecting the same number of plants at random. The alleles present at six candidate gene loci were analysed before and after selection and correlated to WSC content. Significant differences in the allele frequency of L. perenne soluble-acid invertase1:4 were observed between the three populations with one haplotype significantly associated with the high WSC C2S+ population (after three rounds of selection and two rounds of recombination). Moreover, WSC content was also associated with biomass accumulation. Thus, in addition to a 2.84-fold increase in WSC yield, the C2S+ population also had 1.48-fold more biomass per plant, resulting in 3.9-fold higher WSC yield per plant than the control population.

Carbon assimilation and partitioning into non-structural carbohydrate in contrasting varieties of Lolium perenne

Summary Rates of photosynthetic carbon assimilation and the partitioning of carbon into different sugar fractions have been compared in Aurora and Perma, two varieties of perennial ryegrass ( Lolium perenne L.). Perma had a higher photosynthetic rate than Aurora at all irradiances, and numerically a considerably higher apparent maximum quantum yield although this was not statistically significant. On a leaf area basis Perma had heavier leaves that contained more chlorophyll a , chlorophyll b and carotenoid. Carbohydrate was predominantly present in the leaves of both varieties as sucrose and fructan. Fructose, glucose and starch were undetectable or only present in trace amounts. Perma leaf blades and sheaths contained more sucrose and more small (DP3) fructan than Aurora, but less polymeric fructan. Following depletion of carbohydrate pools in low light, excised leaves were induced to accumulate fructan at high light. Perma accumulated 88 % more water-soluble carbohydrate and 90 % more starch than Aurora. However after 24 h the polymeric fructan content of Perma leaves was still lower than that of Aurora. The increase in carbohydrate content of Aurora leaves was nearly all explained by the increase in polymeric fructan, whereas Perma partitioned less into polymeric fructan and contained significantly more glucose, sucrose and DP3 fructan. Additionally, the average chain length of Perma polymeric fructan was shorter than that of Aurora.