Transcriptional profiling of BnDGAT1 genes

Transcriptional profiling of the four BnDGAT1 genes from four developmental stages of the embryo was conducted as described (Troncoso-Ponce et al., 2011 (link)). Briefly, the seeds of greenhouse-grown B. napus were collected at intervals of 18–20, 23–25, 28–30, and 33–35 days after flowering (DAF) and frozen. After collection, total RNA was extracted using TRIZOL (Invitrogen, http://www.invitrogen.com/), followed first by mRNA purification with a commercial kit (Illustra, GE Healthcare, http://www.gehealthcare.com/) then by conversion to cDNA with Superscript III (Life Technologies, http://www.lifetechnologies.com/us/en/home.html) according to the manufacturer’s instructions. For sequence analysis, cDNA was processed with a Roche Library Preparation Kit (http://www.roche.com/) and sequenced with a 454 sequencer (GS FLX, Roche). Expression of the gene family was quantified taking into account the similarity between the gene sequences. For a given gene, the repartitioning of reads is based on the ratio of the number of reads mapped uniquely on this gene to the total number of reads mapped uniquely on the genes sharing the common reads. FPKM = number of reads × (1000/gene length) × (1 million/total number of reads).