Journal Article

Isolation of an embryogenic line from non-embryogenic <i>Brassica napus</i> cv. Westar through microspore embryogenesis

Meghna R. Malik, Feng Wang, Joan M. Dirpaul, Ning Zhou, Joe Hammerlindl, Wilf Keller, Suzanne R. Abrams, Alison M. R. Ferrie and Joan E. Krochko

in Journal of Experimental Botany

Published on behalf of Society for Experimental Biology

Volume 59, issue 10, pages 2857-2873
Published in print July 2008 | ISSN: 0022-0957
Published online June 2008 | e-ISSN: 1460-2431 | DOI: http://dx.doi.org/10.1093/jxb/ern149

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Brassica napus cultivar Westar is non-embryogenic under all standard protocols for induction of microspore embryogenesis; however, the rare embryos produced in Westar microspore cultures, induced with added brassinosteroids, were found to develop into heritably stable embryogenic lines after chromosome doubling. One of the Westar-derived doubled haploid (DH) lines, DH-2, produced up to 30% the number of embryos as the highly embryogenic B. napus line, Topas DH4079. Expression analysis of marker genes for embryogenesis in Westar and the derived DH-2 line, using real-time reverse transcription-PCR, revealed that the timely expression of embryogenesis-related genes such as LEAFY COTYLEDON1 (LEC1), LEC2, ABSCISIC ACID INSENSITIVE3, and BABY BOOM1, and an accompanying down-regulation of pollen-related transcripts, were associated with commitment to embryo development in Brassica microspores. Microarray comparisons of 7 d cultures of Westar and Westar DH-2, using a B. napus seed-focused cDNA array (10 642 unigenes), identified highly expressed genes related to protein synthesis, translation, and response to stimulus (Gene Ontology) in the embryogenic DH-2 microspore-derived cell cultures. In contrast, transcripts for pollen-expressed genes were predominant in the recalcitrant Westar microspores. Besides being embryogenic, DH-2 plants showed alterations in morphology and architecture as compared with Westar, for example epinastic leaves, non-abscised petals, pale flower colour, and longer lateral branches. Auxin, cytokinin, and abscisic acid (ABA) profiles in young leaves, mature leaves, and inflorescences of Westar and DH-2 revealed no significant differences that could account for the alterations in embryogenic potential or phenotype. Various mechanisms accounting for the increased capacity for embryogenesis in Westar-derived DH lines are considered.

Keywords: Brassica napus; embryogenesis; microarray; microspore; transcript profiling

Journal Article.  8324 words.  Illustrated.

Subjects: Plant Sciences and Forestry

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