Abstract
Unravelling the biosynthetic pathway of quinolizidine alkaloids (QAs), regarded as antinutritional compounds of narrow-leafed lupin (NLL) seeds, is fundamental to best exploit NLL as food or feed. We investigated 12 candidate genes connected to QA biosynthesis, selecting them by transcriptomic and genomic approaches, from the landscape of genes differentially expressed in leaves of the high- and low-alkaloid NLL accessions. Linkage analysis enabled the assessment of the location of the candidate genes in relation to iucundus, a major locus of unknown identity, that confers reduced QA content in seeds. The key finding was the identification of APETALA2/ethylene response transcription factor, RAP2-7, cosegregating with the iucundus locus and located within a region with highly significant QTLs that affect QA composition. We additionally identified a 4-hydroxy-tetrahydrodipicolinate synthase (DHDPS) gene involved in L-lysine biosynthesis as being closely linked to iucundus. The distributed location of other remaining candidates (including previously known QA genes) across different linkage groups, also indirectly supports the transcription factor as a possible regulator of lupin alkaloid biosynthesis. Our findings provide crucial insight into QA biosynthesis in NLL. Additionally, we evaluated and selected appropriate reference genes for qRT-PCRs to analyse the expression levels of QA genes in NLL.
Highlights
Quinolizidine alkaloids (QAs) are lysine-derived secondary metabolites that are distributed mainly in family Leguminosae and predominantly in genus Lupinus[1,2,3]
The high-alkaloid accession P27255 was chosen to assemble the reference transcriptome, as we expected that genes involved in quinolizidine alkaloids (QAs) biosynthesis are upregulated in bitter accessions
Based on the assumption that genes involved in QA biosynthesis are upregulated in leaves of bitter accessions, we assembled a reference transcriptome for high-alkaloid P27255, as the reference genotype very likely to express alkaloid-associated transcripts at consistent levels that would enable the assembly of good-quality, full-length transcripts
Summary
Quinolizidine alkaloids (QAs) are lysine-derived secondary metabolites that are distributed mainly in family Leguminosae and predominantly in genus Lupinus[1,2,3]. In the Polish Lupinus Gene Bank (Poznan Plant Breeders Ltd., Poland), we previously found accessions of narrow-leafed lupin (Lupinus angustifolius L.) and white lupin (Lupinus albus L.) with much lower alkaloid content in their seeds (0.0005% and 0.016% of the seed dry weight, respectively)[24,25]. This demonstrates the improvement of breeding, and shows the high potential of collection as the source of low alkaloid material. Genetic and genomic analyses of NLL were integrated using molecular cytogenetic tools, which provided insights into the cytomolecular organization and evolution of the NLL genome[36,37,38,39,40]
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