Background The incorporation of root traits into elite germplasm is typically a slow process. Thus, innovative approaches are required to accelerate research and pre-breeding programs targeting root traits to improve yield stability in different environments and soil types. Marker-assisted selection (MAS) can help to speed up the process by selecting key genes or quantitative trait loci (QTL) associated with root traits. However, this approach is limited due to the complex genetic control of root traits and the limited number of well-characterised large effect QTL. Coupling MAS with phenotyping could increase the reliability of selection. Here we present a useful framework to rapidly modify root traits in elite germplasm. In this wheat exemplar, a single plant selection (SPS) approach combined three main elements: phenotypic selection (in this case for seminal root angle); MAS using KASP markers (targeting a root biomass QTL); and speed breeding to accelerate each cycle. Results To develop a SPS approach that integrates non-destructive screening for seminal root angle and root biomass, two initial experiments were conducted. Firstly, we demonstrated that transplanting wheat seedlings from clear pots (for seminal root angle assessment) into sand pots (for root biomass assessment) did not impact the ability to differentiate genotypes with high and low root biomass. Secondly, we demonstrated that visual scores for root biomass were correlated with root dry weight (r = 0.72), indicating that single plants could be evaluated for root biomass in a non-destructive manner. To highlight the potential of the approach, we applied SPS in a backcrossing program which integrated MAS and speed breeding for the purpose of rapidly modifying the root system of elite bread wheat line Borlaug100. Bi-directional selection for root angle in segregating generations successfully shifted the mean root angle by 30° in the subsequent generation (P ≤ 0.05). Within 18 months, BC2F4:F5 introgression lines were developed that displayed a full range of root configurations, while retaining similar above-ground traits to the recurrent parent. Notably, the seminal root angle displayed by introgression lines varied more than 30° compared to the recurrent parent, resulting in lines with both narrow and wide root angles, and high and low root biomass phenotypes. Conclusion The SPS approach enables researchers and plant breeders to rapidly manipulate root traits of future crop varieties, which could help improve productivity in the face of increasing environmental fluctuations. The newly developed elite wheat lines with modified root traits provide valuable materials to study the value of different root systems to support yield in different environments and soil types.

Background: The incorporation of root traits into elite germplasm is typically a slow process. Thus, innovative approaches are required to accelerate research and pre-breeding programs targeting root traits to improve yield stability in different environments and soil types. Marker-assisted selection (MAS) can help to speed up the process by selecting key genes or quantitative trait loci (QTL) associated with root traits. However, this approach is limited due to the complex genetic control of root traits and the limited number of well-characterised large effect QTL. Coupling MAS with phenotyping could increase the reliability of selection. Here we present a useful framework to rapidly modify root traits in elite germplasm. In this wheat exemplar, a single plant selection (SPS) approach combined three main elements: phenotypic selection (in this case for seminal root angle); MAS using KASP markers (targeting a root biomass QTL); and speed breeding to accelerate each cycle.Results: To develop a SPS approach that integrates non-destructive screening for seminal root angle and root biomass, two initial experiments were conducted. Firstly, we demonstrated that transplanting wheat seedlings from clear pots (for seminal root angle assessment) into sand pots (for root biomass assessment) did not impact the ability to differentiate genotypes with high and low root biomass. Secondly, we demonstrated that visual scores for root biomass were correlated with root dry weight (r = 0.73), indicating that single plants could be evaluated for root biomass in a non-destructive manner. To highlight the potential of the approach, we applied SPS in a backcrossing program which integrated MAS and speed breeding for the purpose of rapidly modifying the root system of elite bread wheat line Borlaug100. Bi-directional selection for root angle in segregating generations successfully shifted the mean root angle by 30o in the subsequent generation (P ≤ 0.05). Within 18 months, BC2F4:F5 introgression lines were developed that displayed a full range of root configurations, while retaining similar above-ground traits to the recurrent parent. Notably, the seminal root angle displayed by introgression lines varied more than 30° compared to the recurrent parent, resulting in lines with both narrow and wide root angles, and high and low root biomass phenotypes.Conclusion: The SPS approach enables researchers and plant breeders to rapidly manipulate root traits of future crop varieties, which could help improve productivity in the face of increasing environmental fluctuations. The newly developed elite wheat lines with modified root traits provide valuable materials to study the value of different root systems to support yield in different environments and soil types.

Aims Rhizoboxes allow non-invasive phenotyping of root systems and are often used as an alternative to evaluation in the field which typically requires excavation, a laborious endeavour. Semi-automated rhizobox methods can be used to screen large numbers of plants, but these platforms can be expensive due to the cost of customised components, assembly, and maintenance, which limits the accessibility for many root researchers. To widen access to the rhizobox method—for example for preliminary screening of germplasm for root system architecture traits—we present a method to build a simple, low-cost rhizobox method using widely available materials, which should allow any research group to conduct root experiments and phenotype root system architecture in their own laboratories and greenhouses. Methods The detailed construction of 80 wooden rhizoboxes is described (each 40 cm width x 90 cm height x 6 cm depth; total cost 1,786 AUD, or 22 AUD or [$15 USD] per rhizobox). Using a panel of 20 spring wheat lines, including parental lines and derived intro-selection lines selected for divergent seedling root traits (seminal root angle and root biomass), genotypic variation in root biomass distribution were examined in the upper (0–30 cm), middle (30–60 cm) and lower sections (60–90 cm) of the rhizobox. At the conclusion of the experiment, rhizobox covers were removed and the exposed roots were imaged prior to destructive root washing. Root morphological traits were extracted from the images using RhizoVision Explorer (Seethepalli and York 2020). Results There were significant genotypic differences in total root biomass in the upper and middle sections of the rhizobox, but differences were not detected in the deepest section. Compared with the recurrent elite parent Borlaug100, some of the intro-selection lines showed greater biomass (or less), depending on the status of the root biomass QTL on chromosome 5B. Genotypes also differed in shoot biomass and tiller number. The donor lines for high and low root biomass showed corresponding differences in shoot biomass. Additional root parameters such as total root length and branching frequency were obtained through image analysis and genotypic effects were detected at different depths. Conclusions The rhizobox set up is easy-to-build-and-implement for phenotyping the root distribution of wheat. This will support root research and breeding efforts to identify and utilise sources of genetic variation for target root traits that are needed to develop future wheat cultivars with improved resource use efficiency and yield stability.

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