In SP3 of the NUE-CROPS project the challenge of addressing NitUE in oilseed rape was approached by phenotyping and genotyping two relatives of oilseed rape (Brassica napus); Arabidopsis thaliana and Brassica rapa (Fig. 1) as well as oilseed rape itself.
Fig. 1. Three closely related species used to study NitUE in oilseed rape
Arabidopsis thaliana is the best studied dicot plant model species, belonging to the Brassicaceae family. In the NUE-CROPS project this species was studied to identify genes involved in Nitrogen Use Efficiency (Nit-UE) which can be used as syntenic markers for breeding strategies of target Brassicaceae crops Brassica napus and Brassica rapa.
Genome-Wide Association (GWA) mapping was used as a tool to identify genes that are involved in Nit-UE, for which there is genetic variation in the population. The population used consisted of a set of 350 distinct natural accessions of Arabidopsis thaliana, collected world-wide: the HapMap population. This population is very densely genotyped using 250,000 single nucleotide polymorphism (SNP) markers (one marker every 400 bp), which makes the use of GWA analysis ideal for analysis of the population.
The HapMap population was screened in a climate chamber (Fig. 2) at two levels of N supply: 1 mM NH4+/NO3 - (N-deficiency) and 5 mM NH4+/NO3- (control).
Fig. 2 Arabidopsis thaliana grown under N-deficiency (left) and control conditions (right). The pictures were taken 5 weeks after germination.
This work resulted in 15 candidate QTL being identified.
Researchers at the Chinese Academy of Agricultural Sciences used Brassica rapa as a model species to study the genetics of NitUE. Their goal was to quantify genetic variation in NitUE in a wide collection of B. rapa germplasm and investigate the phenotypic response to contrasting N supplies. This led them to eventually identify genetic markers specific to genes/loci involved in NitUE. This work involved both field and pot experiments (Fig. 3).
Fig. 3. Top: field trials in China with Brassica rapa grown at two levels of N input (0 and 120 kg N/ha); Bottom: pot trials growing Brassica rapa at two N input levels (3 and 10 mM nutrient solution)
Work on oilseed rape (Brassica napus) focussed on:
A population of 150 double haploid (DH) lines derived from a cross between a dwarf and a normal type parent was developed by KWS. These DH lines were crossed with a tester to produce 75 semi-dwarf (Fig. 4) and 75 normal type testcross hybrids.
In the first growing season 2010/11, these 150 testcrosses were grown in replicated field trials at two locations without additional nitrogen fertilization (low N input).
In the second growing season 2011/12, a subset of these testcrosses was grown in replicated field trials at two locations (Fig. 5). Besides the low nitrogen fertilization treatment a second level of nitrogen fertilization with optimal supply was added.
In every season there were two different harvest dates. The first one at the end of flowering, to measure the biomass and the amount of nitrogen taken up, and the second harvest date at grain maturity, to measure the straw and seed yield and to analyse their N-content.
The phenotypic data from these two years of field trials was used to map QTL influencing NUE related traits.
Fig. 4 Semi-dwarf and normal type at Low N-input conditions
Fig. 5 Field Trials in Einbeck (KWS) in the growing season 2011/12
Semi-dwarf varieties yielded more than normal varieties at low levels of N input and had a higher harvest index. This suggests that they have some traits that make them able to take up and utilize N efficiently when supplies are low.
In conclusion, researchers in SP3 found that:
A number of publications were produced from this work, as listed below:
Wu J, Wei K, Cheng F, Li S, Wang Q, Zhao J, Bonnema G, Wang X (2012) A naturally occurring InDel variation in BraA.FLC.b (BrFLC2) associated with flowering time variation in Brassica rapa. BMC Plant Biol 12 (1):151
Wang Y, Sun S, Liu B, Wang H, Deng J, Liao Y, Wang Q, Cheng F, Wang X, Wu J (2011) A sequenced-based genetic linkage map as a reference for Brassica rapa pseudochromosome assembly. BMC Genomics 12 (1):239
Wang X, Wang H, Wang J, Sun R, Wu J, Liu S, Bai Y, Mun J-H, Bancroft I, Cheng F, Huang S, Li X et al. (2011) The genome of the mesopolyploid crop species Brassica rapa. Nat Genet 43 (10):1035-1040