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The Nec3 gene is a putative negative regulator of pathogen induced programmed cell death in barley

Gazala Ameen: Department of Plant Pathology, North Dakota State University

<div>To defend against biotrophic pathogens, plants rely on programmed cell death (PCD) responses that are initiated to isolate and impede colonization. However, necrotrophs often hijack these immunity responses to complete their lifecycle and proliferate. Thus, elucidating mechanisms of PCD suppression or enhancement is important. However, mutagenesis and forward genetics is one of the only options to study these conserved pathways. The <em>nec3</em> mutants of barley, predominantly show distinctive large cream to orange necrotic lesions specifically during infection with several necrotrophic pathogens. A g-irradiation induced <em>nec3</em> mutant in cultivar Bowman background, designated nec3-γ1, segregated as a single recessive gene in a nec3-γ1 x cv Quest F<sub>2</sub> population when inoculated with <em>B. sorokiniana</em> isolate ND85F. The homozygous F<em><sub>2</sub></em> susceptible progeny, were genotyped with 4 SSR and 25 SNP markers spanning the barley ch. 6H region where <em>nec3</em> was previously mapped at low resolution. The 29 markers were perfectly ordered on the newly released barley genome sequence and <em>nec3</em> was delimited to an ~0.14 cM region spanning ~16.96 Mb of genomic sequence containing 80 annotated high confidence genes. Exome capture sequencing of three independent <em>nec3</em> mutants failed to identify a candidate gene. However, RNAseq analysis of pathogen challenged and control nec3-γ1 and wildtype Bowman identified two genes in the delimited region with >3 fold down regulation in the nec3-γ1 mutant, representing candidate <em>nec3</em> genes.</div>