Theoretical and Applied Genetics

Microsatellite markers are increasingly being used in crop plants to discriminate among genotypes and as tools in marker-assisted selection. Here we evaluated the use of microsatellite markers to quantify the genetic diversity within as well as among accessions sampled from the world germplasm collection of sorghum. Considerable variation was found at the five microsatellite loci analysed, with an average number of alleles per locus equal to 2.4 within accessions and 19.2 in the overall sample of 25 accessions. The collection of sorghum appeared highly structured genetically with about 70% of the total genetic diversity occurring among accessions. However, differentiation among morphologically defined races of sorghum, or among geographic origins, accounted for less than 15% of the total genetic diversity. Our results are in global agreement with those obtained previously with allozyme markers. We were also able to show that microsatellite data are useful in identifying individual accessions with a high relative contribution to the overall allelic diversity of the collection.

The use of major resistance genes is the most cost-effective strategy for preventing stem rust epidemics in Australian wheat crops. The long-term success of this strategy is dependent on combining resistance genes that are effective against all predominant races of the pathogen, a task greatly assisted by the use of molecular markers linked to individual resistance genes. The wheat stem rust resistance genes Sr24 and Sr26 (derived from Agropyron elongatum) and SrR and Sr31 (derived from rye) are available in wheat as segments of alien chromosome translocated to wheat chromosomes. Each of these genes provides resistance to all races of wheat stem rust currently found in Australia .We have developed robust PCR markers for Sr24 and Sr26 (this study) and SrR and Sr31 (previously reported) that are applicable across a wide selection of Australian wheat germplasm. Wheat lines have recently become available in which the size of the alien segments containing Sr26, SrR and Sr31 has been reduced. Newly developed PCR-markers can be used to identify the presence of the shorter alien segment in all cases. Assuming that these genes have different gene-for-gene specificities and that the wheat industry will discourage the use of varieties carrying single genes only, the newly developed PCR markers will facilitate the incorporation of two or more of the genes Sr24, Sr26, SrR and Sr31 into wheat lines and have the potential to provide durable control to stem rust in Australia and elsewhere.

Development of improved Eucalyptus genotypes involves the routine identification of breeding stock and superior clones. Currently, microsatellites and random amplified polymorphic DNA markers are the most widely used DNA-based techniques for fingerprinting of these trees. While these techniques have provided rapid and powerful fingerprinting assays, they are constrained by their reliance on gel or capillary electrophoresis, and therefore, relatively low throughput of fragment analysis. In contrast, recently developed microarray technology holds the promise of parallel analysis of thousands of markers in plant genomes. The aim of this study was to develop a DNA fingerprinting chip for Eucalyptus grandis and to investigate its usefulness for fingerprinting of eucalypt trees. A prototype chip was prepared using a partial genomic library from total genomic DNA of 23 E. grandis trees, of which 22 were full siblings. A total of 384 cloned genomic fragments were individually amplified and arrayed onto glass slides. DNA fingerprints were obtained for 17 individuals by hybridizing labeled genome representations of the individual trees to the 384-element chip. Polymorphic DNA fragments were identified by evaluating the binary distribution of their background-corrected signal intensities across full-sib individuals. Among 384 DNA fragments on the chip, 104 (27%) were found to be polymorphic. Hybridization of these polymorphic fragments was highly repeatable (R2>0.91) within the E. grandis individuals, and they allowed us to identify all 17 full-sib individuals. Our results suggest that DNA microarrays can be used to effectively fingerprint large numbers of closely related Eucalyptus trees.

Chúng tôi mô tả cách công nghệ đa dạng gen (DArT) có thể kết hợp với việc phân loại nhiễm sắc thể để tăng mật độ của các bản đồ di truyền trong các vùng ở gen cụ thể. Nhiễm sắc thể 3B và cánh ngắn của nhiễm sắc thể 1B (1BS) của lúa mì đã được phân lập bằng phương pháp lọc dòng tế bào và được sử dụng để phát triển các mảng genotip được làm giàu cho nhiễm sắc thể và cánh nhiễm sắc thể chứa 2.688 dòng 3B và 384 dòng 1BS. Phân tích liên kết cho thấy 553 trong số 711 marker dị hợp sắc thể 3B (78%) được bản đồ hóa vào nhiễm sắc thể 3B, và 59 trong số 68 marker dị hợp sắc thể 1BS (87%) được bản đồ hóa vào nhiễm sắc thể 1BS, xác nhận tính hiệu quả của phương pháp phân loại nhiễm sắc thể. Để chứng minh tiềm năng làm dày các bản đồ di truyền, chúng tôi đã xây dựng một bản đồ đồng thuận của nhiễm sắc thể 3B bằng cách sử dụng 19 quần thể bản đồ, bao gồm một số quần thể đã được genotip hóa bằng mảng làm giàu 3B. Các marker DArT có nguồn gốc từ 3B đã gấp đôi số lượng vị trí di truyền được bao phủ. Bản đồ đồng thuận thu được, có thể là bản đồ di truyền dày đặc nhất của 3B tính đến thời điểm này, chứa 939 marker (779 DArT và 160 marker khác) phân ly trên 304 vị trí di truyền khác nhau. Quan trọng là chỉ 2.688 dòng 3B (probe) cần được sàng lọc để có được gần gấp đôi số marker 3B dị hợp (510) so với việc sàng lọc khoảng 70.000 dòng gen từ toàn bộ bộ gen (269). Do một mảng DArT làm giàu có thể được phát triển từ dưới 5 ng DNA nhiễm sắc thể, một lượng có thể thu được trong vòng 1 giờ sau khi phân loại, phương pháp này có thể dễ dàng được áp dụng cho bất kỳ loại cây trồng nào mà việc phân loại nhiễm sắc thể có sẵn.

Aluminum (Al) toxicity is a major constraint for wheat production in acidic soils. Wheat producers now routinely use Al-resistant cultivars as one cost-effective means to reduce risks associated with acidic soils. To date, diverse Al-resistant materials have been identified, but their genetic relationship has not been well characterized. A total of 57 wheat accessions, including the majority of the parents of Al-resistant accessions we identified in a previous study, were evaluated for Al resistance and analyzed with 49 simple sequence repeat (SSR) markers and 4 markers for Al-activated malate transporter (ALMT1). Pedigree and principle coordinate analysis (PCA) both separated Al-resistant accessions into four groups labeled according to common ancestry or geographical origin: US-Fultz, Polyssu, Mexican and Chinese. Al resistance in the four groups may have three independent origins given their distinct geographic origins and gene pools. Fultz originated in the USA as a major ancestor to soft red winter wheat, Polyssu originated in Brazil as a major source of Al resistance used in most genetic studies worldwide, and the Chinese group originated in China. Based on ALMT1 marker haplotypes, the Al resistance in the Polyssu and Mexico groups was likely derived from Polyssu, while most Al-resistant cultivars developed in the USA most likely inherited most of Al resistance from Fultz. Fultz was released about 50 years earlier than Polyssu. Norin 10 likely played a pivotal role in passing Al-resistant gene(s) from Fultz to better adapted, semi-dwarf wheat cultivars developed in the USA. Further characterization of Al resistance in the three different sources could reveal multiple Al-resistant mechanisms in wheat.

Fusarium head blight (FHB) resistance was identified in the alien species Leymus racemosus, and wheat-Leymus introgression lines with FHB resistance were reported previously. Detailed molecular cytogenetic analysis of alien introgressions T01, T09, and T14 and the mapping of Fhb3, a new gene for FHB resistance, are reported here. The introgression line T09 had an unknown wheat-Leymus translocation chromosome. A total of 36 RFLP markers selected from the seven homoeologous groups of wheat were used to characterize T09 and determine the homoeologous relationship of the introgressed Leymus chromosome with wheat. Only short arm markers for group 7 detected Leymus-specific fragments in T09, whereas 7AS-specific RFLP fragments were missing. C-banding and genomic in situ hybridization results indicated that T09 has a compensating Robertsonian translocation T7AL·7Lr#1S involving the long arm of wheat chromosome 7A and the short arm of Leymus chromosome 7Lr#1 substituting for chromosome arm 7AS of wheat. Introgression lines T01 (2n = 44) and T14 (2n = 44) each had two pairs of independent translocation chromosomes. T01 had T4BS·4BL-7Lr#1S + T4BL-7Lr#1S·5Lr#1S. T14 had T6BS·6BL-7Lr#1S + T6BL·5Lr#1S. These translocations were recovered in the progeny of the irradiated line Lr#1 (T5Lr#1S·7Lr#1S). The three translocation lines, T01, T09, and T14, and the disomic addition 7Lr#1 were consistently resistant to FHB in greenhouse point-inoculation experiments, whereas the disomic addition 5Lr#1 was susceptible. The data indicated that at least one novel FHB resistance gene from Leymus, designated Fhb3, resides in the distal region of the short arm of chromosome 7Lr#1, because the resistant translocation lines share a common distal segment of 7Lr#1S. Three PCR-based markers, BE586744-STS, BE404728-STS, and BE586111-STS, specific for 7Lr#1S were developed to expedite marker-assisted selection in breeding programs.