Selection of candidate genes from QTL mapping
In order to select and prioritize positional candidate genes from the QTL confidence intervals, we conducted a literature screen and anin silico analysis (explained in more detail in the Materials and Methods section) that were based on functional annotations, gene expression data and tissue distribution of the selected genes. We analyzed loci from the Dol-associated QTL (DOL1) and from the three Pren-associated QTLs (PRE1, PRE2, PRE3). We selected the intervals that were characterized by the highest percentage of phenotypic variance related to each QTL and the highest LOD score values linked with the lowest number of loci (Table S1). As a result of the above-described procedure of selection and prioritization, we generated four sets of genes ‒ three for Prens (Table S2) and one for Dol (Table S3).
Within a set of potential candidate genes for Pren (Table S2), there was the AT5G45940 gene encoding the Nudix hydrolase 11 (Kupke, Caparrós-Martín, Malquichagua Salazar, & Culiáñez-Macià, 2009) with putative IPP isomerase activity. For Dol biosynthesis, we identified three loci that might be directly implicated in the process: AT2G17570, encoding a cis -prenyltransferase 3 (CPT3), AT2G17370, encoding HMGR2 (hydroxymethylglutaryl Coenzyme-A reductase 2, also called HMG2, a highly regulated enzyme that constitutes a rate-limiting step in the MVA pathway), and AT2G18620, encoding a putative GGPPS2 (geranylgeranyl diphosphate synthase 2). A brief comment on the putative role of the two latter genes in the Dol pathway is presented in Table S3, while an in-depth characteristic of AT2G17570 (CPT3) is presented below.