Targeted silencing of BjMYB28 transcription factor gene directs development of low glucosinolate lines in oilseed Brassica juncea

Abstract

SummaryBrassica juncea (Indian mustard), a globally important oilseed crop, contains relatively high amount of seed glucosinolates ranging from 80 to 120 lmol/g dry weight (DW). One of the major breeding objectives in oilseed Brassicas is to improve the seed-meal quality through the development of low-seed-glucosinolate lines (<30 lmol/g DW), as high amounts of certain seed glucosinolates are known to be anti-nutritional and reduce the meal palatability. Here, we report the development of transgenic B. juncea lines having seed glucosinolates as low as 11.26 lmol/g DW, through RNAi-based targeted suppression of BjMYB28, a R2R3-MYB transcription factor family gene involved in aliphatic glucosinolate biosynthesis. Targeted silencing of BjMYB28 homologs provided significant reduction in the anti-nutritional aliphatic glucosinolates fractions, without altering the desirable nonaliphatic glucosinolate pool, both in leaves and seeds of transgenic plants. Molecular characterization of single-copy, low glucosinolate homozygous lines confirmed significant down-regulation of BjMYB28 homologs vis-a-vis enhanced accumulation of BjMYB28-specific siRNA pool. Consequently, these low glucosinolate lines also showed significant suppression of genes involved in aliphatic glucosinolate biosynthesis. The low glucosinolate trait was stable in subsequent generations of the transgenic lines with no visible off-target effects on plant growth and development. Various seed quality parameters including fatty acid composition, oil content, protein content and seed weight of the low glucosinolate lines also remained unaltered, when tested under containment conditions in the field. Our results indicate that targeted silencing of a key glucosinolate transcriptional regulator MYB28 has huge potential for reducing the glucosinolates content and improving the seed-meal quality of oilseed Brassica crops.

Transcriptome atlas of the Arabidopsis funiculus – a study of maternal seed subregions

“…Further investigation may reveal orthologs in relatives of Arabidopsis like members of the Brassicaceae, which are likely to have great economic and agricultural impact. For example in Brassica juncea, targeted suppression of regulators can lead to more desirable seed traits like a lower seed aliphatic GS content (Augustine et al, 2013). We have found a glucosinolate transporter, GTR1, that accumulates late in seed development in the funiculus and CZSC, indicating that there is a direct role for maternal compartments in glucosinolate loading into developing seeds.…”

Biotic elicitors and mechanical damage modulate glucosinolate accumulation by co-ordinated interplay of glucosinolate biosynthesis regulators in polyploid Brassica juncea

“…Glucosinolate extraction was done as per the protocols described earlier (Augustine et al, 2013a;Kraling et al, 1990). Briefly, 50 mg of the lyophilized tissue powder was used for extraction of glucosinolates in 70% boiling methanol.…”

“…Another area of glucosinolates research is the development of canola quality lines having less than 30 lmoles of glucosinolates per gram dry weight (lmoles g À1 dry wt.) of defatted seed meal in oilseed Brassica species, which are otherwise naturally high in anti-nutritional glucosinolates (Augustine et al, 2013a;Cartea and Velasco, 2008).…”

Characterization of RsMYB28 and RsMYB29 transcription factor genes in radish (Raphanus sativus L.)

“…In our study, the RsMYB28 and RsMYB29 proteins were grouped with the Bju_MYB28s and Bju_MYB29s proteins, respectively, indicating that RsMYB28 and RsMYB29 have a high level of sequence conservation with their homologues from the B. juncea genomes. Silencing of the BjMYB28 transcription factor gene caused a reduction in the GSL content in B. juncea (Augustine et al, 2013b). BoMYB29 has been shown to be a regulator for increasing methylsulfinyl GSL content in B. oleracea (Araki et al, 2013).…”