Identification of SNPs associated with magnesium and sodium uptake and the effect of their accumulation on micro and macro nutrient levels in Vitis vinifera

Rachel P. Naegele, Jason P. Londo, Cheng Zou and Peter Cousins

Peer Jl, February 8, 2021. DOI 10.7717/peerj.10773

By Michelle Podolec

The Takeaway

• Micronutrients are needed in small quantities for healthy plant growth and development. Levels fluctuate in the plant throughout the life span, and may vary based on genotype and type of tissue sampled.
• The ability of grapes to access micronutrients varies. Different genotypes may have different abilities to access and metabolize nutrients.
• Researchers previously identified regions on chromosome 13 related to Fe, Na, and Mg micronutrient usage. Chromosome mapping will help researchers understand complex traits such as micronutrient uptake and accumulation.
• Na levels were consistently associated in deficiency symptoms in this study. Na accumulation was associated with single nucleotide polymorphisms (SNPs) on chromosome 11, 3 and 18.
• Effects of individual SNPs in this research population varied. Symptoms may be related to interactions between micronutrient ions like Al, Na, Fe, and Mg. Most SNPs identified were not associated with known genic regions.
• Chromosome 3 was the major contributor to phenotypic variation observed in the population and significant SNPs for the Na and Mg were found on chromosome 3.
• Overall, heritability was low to no (Cu) for most nutrient concentrations measured. Heritability for micro and macro nutrients is complex.

Background

Researchers examined the genetics of an F₁ breeding population derived from a cross between V. vinifera cultivars ‘Verdejo’ and ‘Gewürztraminer’ to identify relationships between leaf nutrients and nutrient deficiency symptoms, and heritability of these traits. The primary goal was to identify genomic regions and potential candidate genes responsible for micronutrient accumulation and utilization, and to compare results of the SNP detections across the two different genomic reference populations.

Methods

Researchers examined two hundred forty nine own rooted, non-grafted seedlings of the F₁ breeding population of a cross between V. vinifera cultivars ‘Verdejo’ and ‘Gewurztraminer’ planted in Ripperdan, CA. They examined the leaves for foliar micronutrient deficiency symptoms in August and September using a standardized visual assessment, rating vines for plant stunting and leaf chlorosis. Nutrient analysis was performed on petiole and blade samples collected from fruiting canes, encompassing nitrogen, sulfur, phosphorus, potassium, magnesium, calcium, sodium, iron, aluminum, manganese, boron, copper and zinc. A follow-up study examined a subset of vines for root stunting.

Results were analyzed with statistical software to identify relationships within and between the two years of the study, and calculated broad sense heritability with year, genotype and nutrient concentration modeled as random effects.

DNA was extracted and genomes were sequenced using the young leaves collected from each F₁ vine in July. Thompson Seedless and wine grapes PN40024 and PN40024xv2 reference genomes were analyzed for variability in SNPs present in each of the reference genomes. SNPs identified were further analyzed and filtered to determine frequency of minor and major alleles and narrow SNPs based on their physical position. Using kinship matrixes, the researchers again narrowed SNPs to nutrient traits and further analyzed using a variety of methods to determine SNP significance.

Results

Of the total 249 vines, roughly 25% of the 249 vines tested exhibited symptoms of micronutrient deficiencies in leaf tissue, but root tissues showed no visible differences. This ratio suggests the leaf phenotype is segregating as a recessive trait within the population. Symptoms of leaf stunting and marginal chlorosis were consistent among plants in both years. Comparisons between symptomatic and asymptomatic plants showed differences in Na, Mg, Fe and Al levels. Correlations with other nutrients were observed but were not consistent between years.

Using the genetic markers, the researchers were able to demonstrate that 29 SNPs on chromosome 3 were associated with the observations of leaf symptoms and nutrient differences of symptomatic vines. These 29 SNPs spanned much of chromosome 3, complicating the identification of specific genes related to these leaf symptoms. Most SNP locations did not align to gene regions in the two reference genomes, indicating that the phenotype may be due to currently unknown genes. Genes that were close to genetic markers included membrane proteins, transcription factors, and hormone related genes.

Conclusions and Practical Considerations

Using a genetic mapping population between cultivars ‘Verdejo’ and ‘Gewurztraminer’, this study sought to understand the relationship between leaf nutrients and genetic regions with the goal of identifying genes responsible for leaf chlorosis and stunting. Genetic mapping of results from two growing seasons allowed researchers to identify chromosome 3 as the primary genetic region contributing to this phenotype. Significant correlations were observed for nutrient concentrations of Na and Mg, but they did not map to any previously identified genes in grapevine.

This result demonstrates how much more research is needed in grapevine to help identify genes responsible for balanced leaf nutrient levels. Foundational research documenting healthy grapevine leaf nutrient levels across many environments is needed to better contextualize these results. Additionally, reference genomes for more of cultivated grape varieties would increase the resolution and ability of researchers to identify specific genes affecting leaf traits.

Heritability for micro and macro nutrients is complex for perennial crops like grapes. Additional research will be needed to more fully explain the connection between observed symptoms and plant genes.

Michelle Podolec is extension support specialist with the statewide viticulture extension program, based at Cornell AgriTech in Geneva, NY.