Molecular cytogenetic characterization of pre-breeding material produced

with perennial Thinopyrum species in Martonvásár

Gabriella Linc – Gyula Vida – Ottó Veisz – Márta Molnár-Láng

Agricultural Institute, Centre for Agricultural Research, Hungarian Academy of Sciences

Martonvásár 2462, Brunszvik Str.2. Hungary

linc.gabriella@agrar.mta.hu

This research was supported by the Hungarian National Grant (OTKA) nr. 104382 and a Janos Bolyai research fellowship to G.L.

Introduction

Wild relatives of cultivated wheat in the tribe Triticeae represent a rich potential source of

genetic variation for many agriculturally significant characteristics. Perennial species such as wheat

grasses includes diploid and polyploid species, containing genomes that are non-homologous to those

of wheat, are important as tertiary gene pools for wheat improvement. Species belonging to the

Thinopyrum genus are known to possess genes conferring resistance to various diseases and are

capable to produce hybrids with bread wheat.

Materials and methods

• probe labelling by Nick translation using DIG- or Biotin NT-mix and detected with streptavidin- FITC and anti-

digoxigenin-Rhodamine

• FISH and GISH according to Linc et al. 2012.

• rows of varieties were inoculated in development stage 37-39 on the Zadoks scale (Zadoks et al. 1974) using the

uredospore mixture

• the extent of infection at development stage 77-83 was evaluated in terms of severity (according to the modified Cobb

scale) and host response (resistant, moderately resistant, intermediate, moderately susceptible and susceptible; Stubbs et

al. 1986).

Aim

Our main goal is the characterization of pre-breeding material produced previously in Martonvásár

using BE-1 (wheat–Thinopyrum ponticum partial amphiploid) and different wheat genotypes (leaf

rust resistant and susceptible genotypes) using molecular cytogenetic methods.

Results and future prospects

Pre-breeding material developed earlier in Martonvásár were

maintained in nursery and greenhouse (Fig 1). After the 12

generations, new partial amphiploid lines with higher chromosome

number (50-56) were isolated and Thinopyrum chromosomes were

detected by GISH (Fig 2). Several stable pre-breeding line with 42

chromosomes were selected and analyzed by FISH using repetitive

DNA sequences (Fig 3). Chromosome 3B of the LRMAS 10 line showed

polymorphism compared to the 3B FISH chromosome pattern of the

GK Öthalom genotype (Fig 4a, b). Their leaf rust resistance was

evaluated in an artificially inoculated nursery. LRMAS 10 line

produced by BE-1 and GK Öthalom (leaf rust susceptible wheat

genotype) crosses was selected as a leaf rust resistant genotype

after the artificial infection (Fig 5). Further experiments in order to

detect alien DNA in the wheat background by GISH and molecular

marker analysis is in progress.

References

Zadoks JC, Chang TT, Konzak CF (1974) A decimal code for the growth stages of cereals. Weed Res 14, 415-421.

Stubbs RW, Prescott EE, Saari EE, Dubin HJ (1986) Cereal disease methodology manual. CIMMYT, Mexico

Linc G, Sepsi A, Molnár-Láng M (2012) A FISH karyotype to study chromosome polymorphisms for the Elytrigia elongata E genome. Cytogenet. Genome Res.

136:138-144 DOI: 10.1159/000334835

Fig 1. LRMAS 10 line maintained in nursery,Martonvásár.

Pseudoroegneria spicata Triticum aestivum

Fig 2. LRMAS 9 line chromosomes after

GISH. Pseudoroegneria spicata DNA

detected anti-DIG-Rhodamine (red colour),

wheat chromosomes counterstained with

DAPI (blue colour).

Fig 5. The extent of infection 1 week after

the inoculation. a; BE1, b; GK Öthalom,

c; LRMAS 10.

a b c

Fig 3. FISH chromosome pattern of the LRMAS 10 line.

pSc119.2 Afa family

a b

Fig 4. FISH pattern of 3B chromosome of the

genotypes a; GK Öthalom and b; LRMAS 10 using

pSc119.2 (green) and Afa family (red) repetitiv DNA

sequences.