Plant & Animal Genomes XIV Conference

January 14-18, 2006
Town & Country Convention Center
San Diego, CA

Impaired Splicing Of fad3 Pre-mRNA In A Low Linolenic Mutant Of Canola (Brassica napus L.)

Xueyi Hu , Mandy L. Sullivan-Gilbert , Manju Gupta , Raghav Ram , Steven A. Thompson

  Dow AgroSciences LLC, 9330 Zionsville Road, Indianapolis, IN 46268, USA

The canola (Brassica napus) fad3 gene encodes for endoplasmic delta-15 linoleate desaturase, which is responsible for the desaturation of linoleic acid (C18:2) into linolenic acid (C18:3). The canola mutant line DMS100, developed through ethyl methanesulphonate (EMS) mutagenesis carries a G-to-A base substitution at the 5’ splice site +1 G of fad3 intron 6 and contains reduced C18:3 content in oil seed. Reverse transcription (RT) PCR using the primers specific to the exon 5 and 7 amplified a fragment of larger size in the fad3 mutant than in wild type. DNA sequence analysis showed that the entire intron 6 is retained in the mature fad3 transcript of the mutant. Thus, the fad3 G-to-A mutation prevents intron splicing. The retained Intron 6 contains stop codons in all three possible reading frames, therefore could lead to early termination of translation and synthesis of a shorter polypeptide for fad3. The incomplete translation of fad3 can inactivate the enzyme and block the desaturation of linoleic acid (C18:2) to linolenic acid (C18:3), resulting in the decrease of C18:3 accumulation in canola seeds. This is consistent with the observation that the fad3 mutant has significantly lower C18:3 content (<3%) than the wild type (~7%).