Aneeta Pradhan

Trigenomic bridges for Brassica improvement

We introduce and review Brassica crop improvement via trigenomic bridges. Six economically important Brassica species share three major genomes (A, B, and C), which are arranged in diploid (AA, BB, and CC) and allotetraploid (AABB, AACC, and BBCC) species in the classical triangle of U. Trigenomic bridges are Brassica interspecific hybrid plants that contain the three genomes in various combinations, either triploid (ABC), unbalanced tetraploid (e.g., AABC), pentaploid (e.g., AABCC) or hexaploid (AABBCC). Through trigenomic bridges, Brassica breeders can access all the genetic resources in the triangle of U for genetic improvement of existing species and development of new agricultural species. Each of the three Brassica genomes occurs in several species, where they are distinguished as subgenomes with a tag to identify the species of origin. For example, the A subgenome in B. juncea (2n = AABB) is denoted as Aj and the A subgenome in B. napus (2n = AACC) as An. Trigenomic bridges have been used to increase genetic diversity in allopolyploid Brassica crop species, such as a new-type B. napus with subgenomes from B. rapa (Ar) and B. carinata (Cc). Recently, trigenomic bridges from several sources have been crossed together as the ‘founders’ of a potentially new allohexaploid Brassica species (AABBCC). During meiosis in a trigenomic bridge, crossovers are expected to form between homologous chromosomes of related subgenomes (for example Ar and An), but cross-overs may also occur between non-homologous chromosomes (for example between A and C genome chromosomes). Irregular meiosis is a common feature of new polyploids, and any new allotetraploid or allohexaploid Brassica genotypes derived from a trigenomic bridge must achieve meiotic stability through a process of diploidisation. New sequencing technologies, at the genomic and epigenomic level, may reveal the genetic and molecular basis of diploidization, and accelerate selection of stable allotetraploids or allohexaploids. Armed with new genetic resources from trigenomic bridges, Brassica breeders will be able to improve yield and broaden adaptation of Brassica crops to meet human demands for food and biofuel, particularly in the face of abiotic constraints caused by climate change.

Evaluation and breeding of tedera for Mediterranean climates in southern Australia

Abstract. Tedera (Bituminaria bituminosa C.H. Stirton var. albomarginata and var. crassiuscula) has been identified as one of the most productive and drought-tolerant species of herbaceous perennial legumes based on 6 years of field evaluation in Western Australia in areas with Mediterranean climate and annual rainfall ranging from 200 to 600 mm. Importantly, tedera demonstrated broad adaptation to diverse soils, and some accessions have shown moderate levels of tolerance to waterlogging and salinity. Tedera exhibits minimal leaf shedding during summer and autumn. Economic modelling strongly suggests that giving livestock access to green tedera in summer and autumn will dramatically increase farm profit by reducing supplementary feeding. The breeding program (2006–12) evaluated the available genetic diversity of tedera for its field performance in seven nurseries with 6498 spaced plants in total covering a wide variation in rainfall, soils and seasons. Best overall plants were selected using a multivariate selection index generated with best linear unbiased predictors (BLUPs) of dry matter cuts and leaf retention traits. The breeding program also evaluated tedera for grazing tolerance, grazing preference by livestock, waterlogging tolerance, seed production, cold tolerance, disease susceptibility and presence of secondary compounds. Tedera is a diploid, self-pollinated species. Therefore, 28 elite parents were hand-crossed in several combinations to combine outstanding attributes of parents; F1 hybrids were confirmed with the aid of highly polymorphic, simple sequence repeat markers. The F1s were progressed to F4s by single-seed descent breeding. Elite parent plants were selfed for two generations to be progressed in the breeding program without hybridisation. Over time, selections from the crossing and selfing program will deliver cultivars of three ideotypes: (i) drought-tolerant, (ii) cold- and drought-tolerant, (iii) waterlogging- and drought-tolerant.

Trigenomic hybrids from interspecific crosses between Brassica napus and B. nigra

Interspecific hybridisation was carried out between five cultivars of Brassica napus and five accessions of B. nigra in all possible cross combinations including reciprocals. Crossing success was higher when B. napus genotypes were used as female parents. Pollination of 799 B. napus flowers with B. nigra pollen resulted in 433 pods set and 2063 putative hybrid seeds. In the reciprocal direction, pollination of 877 B. nigra flowers with B. napus pollen resulted in 281 pods set and 113 putative hybrid seeds. Pod and seed set varied with genotype and only 19 out of 25 combinations of B. napus × B. nigra and 14 out of 25 combinations of B. nigra × B. napus yielded seeds. Hybridity of 2176 putative hybrid seeds (2063 from B. napus × B. nigra and 113 from B. nigra × B. napus) was tested. Microsatellite markers with known locations for the A, B and C genomes indicated that six plants were true hybrids and one more plant remained unconfirmed for hybrid status. All other plants from putative hybrid seeds had the same DNA banding patterns and similar morphological characters as the female parent. However, the true hybrids had DNA bands from both parents and an intermediate morphology for colour and hairiness of leaf, stem and petiole. Anthers were shrunken and thin with a very limited number of sterile pollen grains. Cytological examination confirmed the triploid status of the hybrid with 27 chromosomes. The unconfirmed hybrid had 9% pollen viability and chromosome count was 27 as with the true hybrid; however, there was no clear B-genome marker from B. nigra.

Correlation of morphological traits with molecular markers in radish (Raphanus sativus)

Radish (Raphanus sativus L.) is an important vegetable crop in Asia. Lack of uniformity in the crop at harvest is due to genetic variability and environmental conditions. Molecular markers associated with morphological traits of seed and seedlings were identified. Seed (100) of radish cvv. Fire Ball and Long White Chinese from Australia and Mino Early, Pyuthane Red, Tokinasi, White Neck and 40 Days from Nepal. The seed was germinated between paper (20°C, 6 days) and seedlings were grown in pots under glasshouse conditions (20°C, further 3 weeks). Morphological characters were measured in seed and seedlings at 6 days and 4 weeks after germination. Deoxyribonucleic acid was extracted from selected samples and Random Amplified Polymorphic DNA (RAPD) markers were identified using 10 primers. Multivariate analysis based on principle coordinates analysis was used to correlate morphological traits with molecular markers within and across cultivars. Several markers associated with high or low seed weight, germination proportion, seedling length and fresh weight were identified. This new method for identifying potential RAPD markers may be useful for marker assisted breeding and selection of improved radish varieties.