Simultaneous over-expressing of an acyl-ACP thioesterase (FatB) and silencing of acyl-acyl carrier protein desaturase (SAD) by artifical microRNAs increases saturated fatty acid levels in Brassica napus seeds

摘要

No temperate oilseeds crops are available that produce oil with a high saturated fatty acid content. To achieve such a profile, Brassica napus cv. DH12075 was engineered by simultaneous seed-specific over-expression of a native fatty acyl-ACP thioesterase B [BnFatB(2)] and artificial microRNA-mediated down-regulation of eight endogenous genes encoding putative stearoyl-ACP desaturases (BnSADs). Semi-quantitative RT-PCR analysis of transformed lines showed that the BnFatB(2) gene was highly over-expressed and the eight putative SAD genes were strongly down-regulated in developing seed demonstrating the successful application of microRNA as a tool for down-regulation of genes in the allotetraploid plant B. napus. Analysis of seed triacylglycerol (TAG) composition revealed that all lines contained high levels of palmitic acid (16:0) and moderately increased levels of stearic acid (18:0). Total saturated fatty acid content was increased from 7.4% in the control to 37.3-45.6% in the transformed lines, with FatB over-expression as the dominant trait. A twofold increase in 16:0 was observed in seed polar lipids. The melting point of oil from mature seeds was increased from -10 \ub0C in DH12075 to 15 \ub0C in the line with the highest saturated fatty acid content. TAG composition showed a shift from predominantly C54 TAG to C50 and C52 TAGs enriched in palmitic acid. Seedling establishment at low temperature was compromised in lines with high saturated fatty acid content. Results suggested that transcript encoding stearoyl-ACP desaturase in developing B. napus seeds is present in considerable excess of the level required for efficient desaturation of 18:0.