Towards Stunt-Free Lonren Reniform Nematode Resistance by Dissection of an Alien R-Gene Chromosome Segment and Detection of Significant Fungal Interactions

Reniform (Rotylenchulus reniformis) nematodes cost US cotton growers over 130 million dollars per year. Genetic resistance of the wild African species Gossypium longicalyx to reniform nematodes was previously transferred into upland cotton (Gossypium hirsutum L.). LONREN and other cotton lines containing the responsible gene, Ren^(lon), are highly nematode-resistant, but when grown in nematode-infested fields, the seedlings and plants became "stunted". Hypothesized causes include hypersensitivity reaction, per se, and/or increased susceptibility to soil pathogens. My research objectives have been: [1] to develop Ren^(lon)-linked markers and map the Ren^(lon) region at high resolution, [2] to recombine Ren^(lon) and nearby loci and determine if ?stunting? is due to linkage drag, [3] to create genomic and germplasm resources for genetic manipulation of chromosome-11, especially Ren^(lon), and [4] to determine if "stunting" is due solely to plant-nematode interactions or involves additional pathogens.

Putative SNPs between G. hirsutum and G. longicalyx were collaboratively sequence-aligned to the D5 reference genome. By selecting SNPs aligned to the Ren^(lon)-homeologous region and screening them against a genetic panel, we identified 85 as Ren^(lon) -linked. To enable large-scale SNP applications, we developed inexpensive methods for high-throughput non-destructive seed DNA extraction for PCR-based genotyping. We then high-resolution mapped 10 SNPs in the proximal alien segment near Ren^(lon). We chose the two closest Ren^(lon)-flanking SNPs and used them for marker-assisted selection to identify 5 recombinants from 17,600 BC1F1 seed. Subsequent progeny tests indicated 2 of the 5 recombinants were free of "stunting" and that 1 seems to be segregating for nematode resistance. Contingent on verification of the resistance, the results indicate that stunting of LONREN seedlings is indeed due to linkage drag, not hypersensitivity, per se. To determine if "stunting" can be ascribed solely to hypersensitivity-induced root damage, we conducted three replicated growth chamber experiments with multiple genotypes, pasteurized and natural soils, and several pathogen treatments. The results showed that Thielaviopsis root rot alone can cause severe seedling "stunting", but not the nematodes, alone. Moreover, they showed that LONREN-like genotypes are differentially sensitive to the combined presence of reniform nematodes and Thielaviopsis, and become more severely stunted.