Identification of parental and recombined chromosomes in hybrid derivatives of Lolium multiflorum × Festuca pratensis by genomic in situ hybridization

Abstract

Genomic in situ hybridization (GISH) was used to identify Festuca chromatin in mitotic chromosomes of Lolium multiflorum (Lm) × Festuca pratensis (Fp) hybrids and hybrid derivatives. In two inverse autoallotriploids LmLmFp and LmFpFp, in situ hybridization was able to discriminate between the Lolium and Festuca chromosomes. In a third triploid hybrid produced by crossing an amphiploid of L. multiflorum × F. pratensis (2n=4x=28) with L. multiflorum (2n=2x=14), the technique identified chromosomes with interspecific recombination. Also, in an introgressed line of L. multiflorum which was homozygous for the recessive sid (senescence induced degradation) allele from F. pratensis, a pair of chromosome segments carrying the sid gene could be identified, indicating the suitability of GISH in showing the presence and location of introgressed genes. By screening backcross progeny for the presence of critical alien segments and the absence of other segments the reconstitution of the genome of the recipient species can be accelerated.

Introgression mapping in the grasses. II. Meiotic analysis of the Lolium perenne/Festuca pratensis triploid hybrid

“…Blocking DNA was prepared by autoclaving the DNA of Lp for 2 min, which produced fragments of approximately 200 bp in length. The in situ hybridization protocol used followed that described by Thomas et al (1994).…”

Introgression of crown rust (Puccinia coronata) resistance from meadow fescue (Festuca pratensis) into Italian ryegrass (Lolium multiflorum) and physical mapping of the locus

“…Excised root tips were kept in icecold water either for 16 h (for mitotic cell divisions) or 18 h (for in situ hybridisation) and then fixed in ethanol:acetic acid (3:1) for a minimum of 2 h. For chromosome counts, root tips were stained by the Feulgen method and squashed on a microscope slide in 1% acetocarmine. For in situ hybridisation, metaphase spreads were treated by the procedure described by Thomas et al (1994) and King et al (1998). Total genomic Fp DNA was used as a probe to detect Fp introgressions; this was labelled either with Cy3-dCTP (Amersham Pharmacia Biotech, UK) (red) or dig-11-dUTP (Boehringer Mannheim, UK) and detected by the fluorescein detection procedure (green).…”

“…On the other hand, the relationship is sufficiently distant that L. multiflorum (Lm) and F. pratensis (Fp) chromosomes can be distinguished in hybrid plants using genomic in situ hybridisation (GISH) (Thomas et al, 1994). The potential use of Lolium/Festuca introgression mapping to develop physical and genetic maps, determine the genetic control of important characters and develop new germplasm for plant breeders has been discussed previously (King et al, 1998).…”

GISH/FISH mapping of genes for freezing tolerance transferred from Festuca pratensis to Lolium multiflorum

“…Furthermore, a shift towards the Lolium parental genotype in amphiploid L. perenne or L. multiflorum  F. pratensis over several generations (up to F 8 ) has been observed by different research groups (Zwierzykowski et al, 1998(Zwierzykowski et al, , 2003Canter et al, 1999;Pašakinskienė and Jones, 2003). As recombination between homoeologous Lolium and Festuca chromosomes occurs frequently in hybrids, opportunities for gene introgression arise, enabling the transfer to Lolium of Festuca genes for improved persistency, through a backcross breeding programme (Thomas et al, 1994Humphreys et al, 1997Humphreys et al, , 2003Humphreys et al, , 2005King et al, 1998;Zwierzykowski et al, 1998Zwierzykowski et al, , 1999. The targeted inclusion of desirable Festuca gene combinations can be accompanied by the targeted exclusion of other Festuca genes considered detrimental to the high forage quality traits found in Lolium.…”