1. Please state project objectives and what work was done this quarter to address them:
Objective 1: Screen breeding populations for new individuals with exceptional HLB tolerance and fruit quality as populations mature and begin to fruit.
Data has been collected on new material, including fruit and juice blends derived from these materials. Tolerant material has been sampled from material early in the harvest season. Drones have been flown over sites with new populations to select the healthiest trees for sampling. The late season oranges and orange-like accessions will be sampled during the late season. Accessions that were treated with OTC were compared versus the controls (non-injected).
Objective 2: Plant new and manage ongoing Stage 2 semi-commercial trials of the selections most likely to succeed in the HLB environment versus commercial standard controls.
One Stage II with sweet orange is ongoing trial is 1.5 years old and is not fruiting yet, but the trees are growing off well and we anticipate the first fruit of that trial coming in next season. Another two trials have trees that are ready to plant but we will wait until after the threat of freeze has passed before planting those. There are other trials that are ongoing and data is being or will be collected from them as the fruit become ready. Also, we have reserved several acres for Stage II trials.
Objective 3: Utilize bioinformatic tools to identify genes or other variants involved in HLB tolerance and possible resistance for plant transformation and conventional breeding.
The first Material has been sent to the ICBR at University of Florida for deep sequencing (1-4-1 Hamlin) to discover why, on a molecular level, the N-13-32 accession typically grows off better than Hamlin 1-4-1. We also recently discovered a Star Ruby grapefruit individual that may be a mutant with enhanced tolerance. We ran the SSR markers to determine that the budline was Star Ruby and the rootstock was US-941 and they were. The next step will be to see if there is mutagenesis that would confer HLB tolerance. We will also test the tree for HLB to see if it is resistant. The SSR work was done without significant expenditure because that work was done in-house with existing inputs.
2. Please state what work is anticipated for next quarter:
Next quarter, we will collect tree and fruit and juice quality data from the late season accessions in trial and in other plantings. We will propagate trees for more Stage II trials. We will reserve more space for Stage II trials. We will collect leaf material during late winter/early spring for the sequencing of the 100 accessions. We will also be in a position to receive and process the genomic data from 1-4-1 Hamlin and will collect transcriptomic data from this accession during flowering to annotate the 1-4-1 genome so that we can compare it to N13-32 Hamlin to look for differences in genome that could confer HLB tolerance in sweet orange and allow for identification of this tolerant budline. We are expecting to meet with the Laurence Berkeley National Laboratory group to discuss the coverage depth and platform of Illumina sequencing for the 100 accessions.
3. Please state budget status (underspend or overspend, and why):
We are 11% underspent (13% time spent vs 2% spent) due to the paperwork with the Laurence Berkeley National Laboratory and the careful planning of the sequencing effort with this group, who will be the ones focused on gene and variant discovery for CRISPR and transgenic work downstream at the Crop Transformation Center. We expect to be close to on track by the next reporting period. Additionally, we have had a change in federal administration which has come with some unanticipated outcomes that we are assessing.