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A GIFT SEED

Objective 1

Genetic Improvement & Production

Leadership

Dr. Jill Wegrzyn (Objective Leader), Dr. Ross Whetten, Dr. Justin Whitehill

Objective Goal & Deliverables

GOAL: Accelerate genetic improvement of fir (Abies spp.) Christmas tree species through the development of genetic and genomic tools.

DELIVERABLES

  1. Four functionally contiguous fir genome assemblies.
  2. Commercially available genotyping assays for Fraser complex, noble fir, and CoFirGE complex.
  3. Pedigree reconstructions of OP families using molecular markers to evaluate Plantation-Scale Studies with stakeholders.
  4. A commercial somatic embryo production system.

Objective Activities

Each of the following activities will contribute to the generation of the Objective 1 deliverables listed above.

The main focus of Activity 1.1. is to obtain functionally contiguous genome assemblies for Fraser, noble, Turkish, and momi fir.

The Objective 1 team will be doing this by selecting fir genotypes for genome sequencing (Sub-Activity 1.1.1.) and extracting and sequencing the genomic DNA (gDNA) of the firs (Sub-Activity 1.1.2.). From this data, the team will assemble genome assembly and annotate the fir genome (Sub-Activity 1.1.3.). Following genome assembly, the annotated genome and transcriptome sequences will be made available through TreeGenesDb.org, a public repository of genomic information for forest trees curated by Co-Project Director Dr. Jill Wegrzyn’s lab, an effort to further develop and expand current international conifer genomic resources (Sub-Activity 1.1.4.).

The main focus of Activity 1.2. is developing genotyping panels to facilitate pedigree reconstruction using molecular markers in fir.

In order to do this, the Objective 1 team first plans to develop the genotyping panels (Sub-Activity 1.2.1.) and then apply them, to facilitate recurrent selections from plantation-scale studies using retrospective analyses of Fraser, noble and CoFirGE complex trees (Sub-Activity 1.2.2.). The next step will then be planning and testing the genotyping panels for molecular breeding and genomic selection (Sub-Activity 1.2.3.).

The main focus of Activity 1.3. is rapid propagation of elite Christmas tree lines using somatic embryogenesis.

Seeds from elite parental crosses identified through Activity 1.2. will be used for rapid propagation of elite lines through a somatic embryogenesis pipeline using established methods in Project Director Dr. Justin Whitehill’s laboratory. Optimization experiments will be carried out in collaboration with the NC State Golden LEAF Biomanufacturing Training and Education Center to scale up somatic embryogenesis production. Once optimized, the Objective 1 team plans to work with commercial tissue culture laboratories to implement mass production of elite planting material from somatic embryogenesis cultures.

Year 1 Progress

Activity 1.1: Genomic Resource Development

Year 1 efforts have focused on developing the primary genomic resources for several fir species, with an initial emphasis on Fraser fir. To expand genomic resources, multiple DNA extraction protocols were tested using Fraser fir needle samples from a single accession. In addition to Fraser fir, extractions and sequencing runs were completed for momi fir, Trojan fir, and noble fir. Trojan fir produced the highest read quality and longest reads (N50 >25 Kb) using both extraction methods.

The next phase of the project will focus on completing the momi fir genome assembly and scaffolding, followed by structural annotation using in-house developed pipelines, including EASEL (Efficient, Accurate, Scalable Eukaryotic modeLs) and EnTAP (Eukaryotic Non-Model Transcriptome Annotation Pipeline).

Activity 1.2: Genotyping

For genotyping, samples have not yet been submitted for sequencing as we are still developing SNP markers and high throughput gDNA extraction methods. A protocol was developed for purification of gDNA from diploid needles that can be used on all focal fir species (Fraser, momi, noble, Trojan). Single-copy regions and SNPs in the Fraser fir genome will be used to design the first generation of fingerprinting assays for genotyping progeny tests and seed orchard trees of Fraser, Canaan, and balsam fir. For noble fir, Objective 1 leads met with Oregon State University and others about what noble fir genotypes for use in project including the collection of seeds.

Trojan/Turkish fir genotyping panels will be focused on in the next report.

Activity 1.3: Rapid Propagation Using Somatic Embyrogenesis

For rapid propagation using somatic embryogenesis, the Whitehill lab has drastically advanced our ability to generate clonal trees in tissue culture. Two lines of momi fir are being developed to facilitate grafting of elite Fraser fir germplasm to combat Phytophthora root rot and provide a practical solution to US Christmas tree farmers.