Identification of Single Nucleotide Polymorphism (SNP) Markers Associated with Botrytis Resistance in Lily (Lilium spp.)

The genus Lilium belongs to the Liliaceae family, which is economically important ornamental monocot flowering plant and highly susceptible to Botrytis infection. A bulk segregant analysis was conducted in susceptible or resistant pools constructed from a parental cross between Asiatic, Oriental and Trumpet lilies to identify genes associated with resistance to Botrytis. Based on previously obtained transcriptome data, 16 cDNA libraries produced from 8 hybridizations for the bulk segregant analysis were then analyzed using differential gene expression (DEG) profiling. Potential up-regulated candidate transcripts which suspected to be linked with disease resistance were identified during a DGE comparison between resistant and susceptible lines. A randomly chosen three candidate genes was evaluated by quantitative real-time PCR (qRT-PCR) to validate the DGE data. As the most common variation, allele specific single nucleotide polymorphisms (SNPs) in the candidate transcripts from the 16 progenies were analyzed and subsequently used to develop EST–SNP markers. A total of 14 different EST–SNP markers potential linked with Botrytis resistance were identified. Further, two EST-SNP markers (mRNA_310345 and mRNA_296428) were tested on DNA sample panels of 16 lily lines to characterize their functional PCR amplification and polymorphisms using high resolution melting (HRM) analysis. It was found that both EST-SNP markers have generated 10 unique HRM profiles and showed a clear classification of 6–7 different lily genotypes in almost accordance to their EST sequence characteristics indicating that HRM is an efficient tool for the rapid screening of sequence variations and allele discrimination. These result findings highlights the significance of SNP genotyping by HMR to identify the disease resistance genes in lily. With further validation, the allele specific EST–SNP markers which were identified in this study could be used for improving resistance to Botrytis in lily breeding programs.