Thinopyrum elongatum serves as an excellent gene pool for wheat improvement. Genes for resistance to many biotic and abiotic stresses have been transferred from Th. elongatum to wheat through chromosome manipulation. For breeding programs, molecular markers enable screening of a large number of genotypes for alien chromosome introgressions. The main objective of the present study was to develop and characterize EST (expressed sequence tags) and PLUG (PCR-based Landmark Unique Gene) markers that can distinguish Th. elongatum chromatin from the wheat genomes. A total of 258 mapped EST primer pairs and 46 PLUG primer pairs were tested on DNA from wheat Chinese Spring (CS) and CS-Th. elongatum addition lines. The results showed that 43 primer pairs could be effectively mapped to specific Th. elongatum chromosomes. Twenty-two of the 43 markers displayed similar homoeologous chromosome locations to hexaploid wheat. Nine markers mapped to different linkage groups between wheat and Th. elongatum, while 12 makers mapped on two or three different Th. elongatum chromosomes. A comparison of molecular marker locations indicated that Th. elongatum genome was closely related to the D genome of wheat, and chromosome rearrangements and duplication had occurred in Th. elongatum and the wheat genomes. The markers will be useful in comparative gene mapping, chromosome evolutionary analysis, and gene introgression for wheat improvement using Th. elongatum accessions as gene donors.

Stripe rust (caused by Puccinia striiformis) occurs annually in most wheat-growing areas of the world. Thinopyrum ponticum has provided novel rust resistance genes to protect wheat from this fungal disease. Wheat - Th. ponticum partial amphiploid line 7430 and a substitution line X005 developed from crosses between wheat and 7430 were resistant to stripe rust isolates from China. Genomic in situ hybridization (GISH) analysis using Pseudoroegneria spicata genomic DNA as a probe demonstrated that the partial amphiploid line 7430 contained ten J(s) and six J genome chromosomes, and line X005 had a pair of J(s)-chromosomes. Giemsa-C banding further revealed that both lines 7430 and X005 were absent of wheat chromosomes 6B. The EST based PCR confirmed that the introduced J(s) chromosomes belonging to linkage group 6, indicating that line X005 was a 6J(s)/6B substitution line. Both resistance observation and sequence characterized amplified region (SCAR) markers displayed that the introduced chromosomes 6J(s) were responsible for the stripe rust resistances. Therefore, lines 7430 and X005 can be used as a donor in wheat breeding for stripe rust resistance.

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Two cytologically stable wheat-Dasypyrum breviarisatatum addition lines, Y93-1-6-6 and Y93-1-A6-4, were identified by integrated molecular and cytogenetic techniques. C-banding and genomic in situ hybridization (GISH) showed that Y93-1-6-6 and Y93-1-A6-4 were different wheat-D. breviaristatum additions. A total of 51 markers (primer/enzyme combinations), including 6 PCR-based Landmark Unique Gene (PLUG) markers and 45 Sequence-Tagged-Site (STS) markers, were selected from 3,774 primer/enzyme combinations to further characterize these two additions. Marker haploytpes suggested that both D. breviaristatum chromosomes in Y93-1-6-6 and Y93-1-A6-4 were rearranged. Stem rust resistance screening indicated that both additions were highly resistant to race RKQQC, whereas only Y93-1-6-6 was resistant to race TTKSK (Ug99). Powdery mildew resistance screening showed that only Y93-1-6-6 was resistant. Pedigree analysis suggested that the stem rust and powdery mildew resistance of Y93-1-6-6 was derived from D. breviaristatum, indicating that the D. breviaristatum chromosomes in Y93-1-6-6 possess a new powdery mildew resistance gene(s), and new stem rust resistance gene(s). These two additions could be used as stem rust or powdery mildew resistance sources in wheat breeding programs.

A species-specific DNA sequence (marker) that can detect the presence of Secale cereale chromatin in common wheat background was developed by using wheat microsatellite primer Xgwm614. One rye-specific DNA amplified fragment of 416bp (pSa614416) was obtained from Secale africanum and a wheat -S. africanum amphiploid. The primer Xgwm614 also gave rise to specific bands in all Chinese Spring-Imperial rye addition lines 1R to 7R. Sequence analysis revealed that pSa614416 was strongly homologous to a miniature inverted transposable element (MITE) stowaway-like element. Results of fluorescence in situ hybridization showed that the signal of pSa614416 was distributed along all S. cereale. cv Jingzhou chromosomes, but the signal strengths were unbalanced on the seven rye genome chromosomes. This repetitive element may be useful as a molecular marker for the introgression of rye germplasm into the wheat genome.

Thinopyrum intermedium, which has many useful traits, is valuable for wheat breeding. A new wheat-Thinopyrum addition line, SN100109, was developed from the progeny of common wheat cultivar Yannong 15 and Th. intermedium. It was resistant to most races of Blumeria graminis f. sp tritici (Bgt), which caused powdery mildew in wheat, and its reactions were different from the reactions of gene Pm40 and Pm43. Genomic in situ hybridization (GISH) and molecular marker analysis were used to identify the genomic composition of SN100109. GISH results showed that SN100109 was a wheat-Th. intermedium disomic addition line containing one pair of J chromosomes, and the resistance gene was located on the alien additional chromosomes of SN100109. And four molecular markers BE425942, BF482714, Xgdm93 and BV679214 which were assigned to homologous group 2, were specific molecular markers of the additional chromosomes. All the results indicated that SN100109 contained one pair of 2J chromosomes. SN100109 can be used as a novel germplasm source for introducing powdery mildew resistance genes to wheat in breeding programs.