Identification Of Disease-Resistant Wheat-Thinopyrum Alien Substitution Lines Using Molecular Cytogenetics

The ongoing improvement of wheat cultivars is dependent on a continuous supply of genetic variability. In wheat breeding programs, the production of (partial) amphiploids and substitution lines between wheat and relative species is an important intermediate step for such a gene transfer, because they allow the reliable analysis of the effects of alien genes in the genetic background of wheat and their fertility allows gene transfer even when an Fi hybrid is almost completely sterile. The wild relatives, Thinopyrum intermedium (2n=6x=42) with the genomic formulus StJJs, has a wealth of genetic variation for the improvement of resistance to fungal diseases including powdery mildew and rusts, the main diseases in wheat-producing regions of the world, but no substitution or translocation conferring the resistances to powdery mildew and stem rust from this species to wheat chromosomes was reported to date.CH08-141 is a homogeneous BC1F6-derived resistant wheat germplasm line developed in our laboratory by crossing common wheat cultivar Jintai 170 twice with partial amphiploid TAI7047 derived from Th. intermedium accession Z1141 with pedigree ’Taiyuan 768/Z1141//Jinchun 5’. In the present study, an attempt has been made to determine the chromosome composition and genomic origins of the alien chromosomes of CH08-141 by genomic in situ hybridization (GISH) and multi color genomic in situ hybridization (mcGISH) and to evaluate its potential as a novel source for resistance to powdery mildew and stem rust pathogens of wheat. The following results were obtained:1. CH08-141 and all the parents including wheat, partial amphiploid TAI7047 and wild accession Z1141 were screened with Ug99, the most widely virulent and prevalent race of stem rust in the world, and E09, the most prevalent race of powdery mildew in northern China. CH08-141, TAI7047 and Th. intermedium were completely resistant (immune or hypersensitive flecks), whereas the parental wheat cultivars Jintai 170, Jinchun 5 were highly susceptible to all the races tested. The disease evaluation showed the resistance of CH08-141 to these two fungal pathogens was similar to that of its putative donor parent TAI7047 and wild parent Th. intermedium. Since CH08-141 is a derivative of that species and all wheat parents involved in its pedigree were highly susceptible, these resistances were derived from Th. intermedium and expressed in this line.2. Chromosome counting showed the chromosome number of CH08-141 varied from In=41 to In=42, with 42 being the most frequent. After the chromosome counting, the GISH technique was used to determine the presence or absence, number and genomic composition of the alien chromosomes in CH08-141. The GISH pattern from probing with genomic DNA of Th. intermedium revealed that one pair of chromosomes was labeled over their entire length and belonged to the alien genome, and the remaining 20 chromosome pairs or 39 chromosomes to the genomes of wheat, indicating that this line is a wheat-alien substitution in which one chromosome pair was derived from Th. intermedium.3. The mcGISH patterns using DNA from A and B genomes as probes showed that one pair of chromosomes from Th. intermedium substituted for one pair of the B-genomic chromosomes of wheat (Triticum aestivum) With two-colour fluorescence in situ hybridization (mcFISH) using the pSc119.2 and pAsl repetitive DNA clones as probes demonstrated that one pair of 6B chromosomes in CH08-141 was replaced by one pair of J-genome chromosomes derived from Th. intermedium in partial amphiploid TAI7047. Meanwhile, the molecular marker analysis using 48 pairs SSR primers evenly distributed on all the 6-group chromosomes of wheat confirmed the absence of 6B chromosomes in CH08-141.Evidences from GISH and mcFISH, and disease screening demonstrated that CH08-141 is a new Th. intermedium-derived wheat alien substitution line in which the wheat chromosome 6B pair was replaced by one pair of Th. intermedium chromosome 6J, and that this line appears to serve as a novel source for the transfer of resistance genes for multiple fungal pathogens to wheat.