Location: Horticultural Crops Disease and Pest Management Research Unit
Project Number: 2072-22000-044-011-G
Project Type: Grant
Start Date: Aug 1, 2020
End Date: Oct 31, 2023
Objective:
1. Develop new diagnostic markers that distinguish among Maine, Ontario and Michigan sources of aphid resistance.
2. Verify developed markers in breeding selections.
3. Summarize and share results with stakeholders and the scientific community.
Approach:
Plant material: there are three study populations: 145 seedlings from the Ontario source: ORUS 4580-2 x ORUS 4305-102 (ON); 137 seedlings from the Maine source: ORUS 4304-128 (ME) x ORUS 4580-2, and 255 seedlings from the Michigan source: ORUS 4310-2 (MI) x ORUS 4580-2. These progeny were tested for aphid response as previously described by Dossett and Finn (2010). Selections used as parents in the breeding program will be tested to confirm resistance.
To study gene expression, RNA was collected and sequenced on using both long-read and short-read sequencing technologies. The aim of the long-read sequencing approach was to capture and identify full-length genes as they were being expressed. Short-read sequencing is currently underway to estimate the level of gene expression – that is, the number of messenger RNA copies that are transcribed from particular genes and are on their way to being translated into functional proteins. Candidate genes differentially expressed in resistant and susceptible individuals will then be compared to the black raspberry genetic linkage map to find candidates mapping to the previously-identified aphid resistance region on linkage group 6 (RLG 6). A quantitative trait locus (QTL) analysis will be conducted to identify DNA differences correlated with aphid responses. Molecular markers will be developed targeting these regions.
A subset of 32 simple sequence repeat (SSR) markers and high resolution melting (HRM) markers that previously mapped to RLG 6 were screened to look for differences between the parents of each population. Based on this initial screening, 16-20 markers will then be used to genotype (distinguish DNA targets) in the populations. Additional markers will be identified from the chromosome corresponding to RLG 6 in the new black raspberry genome assembly and used to saturate RLG6. Markers developed from candidate genes identified through the Iso-Seq approach will also be used to genotype the populations. We will then construct updated linkage maps with Joinmap 4.0 to visualize results. Finally, markers associated with resistance from each of the three sources will be developed and validated in breeding selections that were tested for aphid response.
