Alien introgression in the grasses Lolium perenne (perennial ryegrass) and Festuca pratensis (meadow fescue): the development of seven monosomic substitution lines and their molecular and cytological characterization

Abstract

Background and aims: To address the issues associated with food security, environmental change and bioenergy in the context of crop plants, the production, identification and evaluation of novel plant phenotypes is fundamental. One of the major routes to this end will be wide hybridization and introgression breeding. The transfer of chromosomes and chromosome segments between related species (chromosome engineering or alien introgression) also provides an important resource for determining the genetic control of target traits. However, the realization of the full potential of chromosome engineering has previously been hampered by the inability to identify and characterize interspecific introgressions accurately.

Methods: Seven monosomic substitution lines have been generated comprising Festuca pratensis as the donor species and Lolium perenne as the recipient. Each of the seven lines has a different L. perenne chromosome replaced by the homoeologous F. pratensis chromosome (13 L. perenne + 1 F. pratensis chromosome). Molecular markers and genomic in situ hybridization (GISH) were used to assign the F. pratensis chromosomes introgressed in each of the monosomic substitutions to a specific linkage group. Cytological observations were also carried out on metaphase I of meiosis in each of the substitution lines.

Results: A significant level of synteny was found at the macro-level between L. perenne and F. pratensis. The observations at metaphase I revealed the presence of a low level of interspecific chromosomal translocations between these species.

Discussion: The isolation of the seven monosomic substitution lines provides a resource for dissecting the genetic control of important traits and for gene isolation. Parallels between the L. perenne/F. pratensis system and the Pooideae cereals such as wheat, barley, rye, oats and the model grass Brachypodium distachyon present opportunities for a comparison across the species in terms of genotype and phenotype.

Fig. 1.

Mitosis in the seven monosomic substitution lines. (Fig i–vii) Monosomic lines 1–7; the substituted chromosome is shown by an arrow. Inset images: (a) 4′,6-diamidino-2-phenylindole (DAPI)-stained image of the relevant substitution line; (b) phase contrast image of the relevant substitution line. (Fig ii) Inset images: (c) and d) show the 5s site in monosomic line 2. (Fig iii) Inset images: (c) and (d) show monosomic line 3 carrying the pta71 ribosomal site. (Fig viii) A colchicine doubled line of monosomic line 3 (2n = 28) carrying a disomic substitution. Inset image: (a) shows the disomic chromosome substitutions for monosomic line 3 carrying the two pta71 sites. Scale bar = 7·5 µm.

Fig. 2.

The analysis of chromosome pairing during meiotic metaphase I in L. perenne × F. pratensis monosomic substitution lines. (A–H) GISH images of meiotic metaphase I with F. pratensis (Fp) chromosome shown by a red arrow. (I–K) Phase images of pairing during meiotic metaphase I and drawn representations of the meiotic configuration shown by a blue arrow. (A) Monosomic line 3 showing a ring bivalent involving the Fp chromosome. (B) Monosomic line 5 showing a rod bivalent involving the Fp chromosome. (C) Monosomic line 4 showing the Fp chromosome as part of a quadrivalent. (D) Monosomic line 2 with a rod bivalent, but a quadrivalent is also present involving L. perenne (Lp) chromosomes shown by the green arrow. (E) Monosomic line 2 showing the Fp chromosome as part of a quadrivalent. (F) Proximal chiasma in monosomic line 1. (G) Proximal chiasma in monosomic line 5. (H) Monosomic line 2 showing the Fp chromosome as a univalent and also an Lp univalent (green arrow). (I) Monosomic line 3 showing two univalents. (J) Monosomic line 4 with one trivalent and three univalents. (K) Monosomic line 5 with one trivalent and one univalent.

Fig. 3.

The individual chromosomes of F. pratensis assigned to their respective Triticeae linkage group. A combination of arm ratio, chromosome length, 5S rDNA and 45S rDNA allows identification of each of the F. pratensis chromosomes. The chromosomes are drawn to scale.