Comparative transcriptome analysis of ovules reveals stress related genes associated with nucellar polyembryony in citrus
2014
Kumar, Vajinder | Malik, S. K. (Surendra Kumar) | Pal, Digvender | Srinivasan, Ramamurthy | Bhat, Shripad Ramachandra
Adventive polyembryony is a form of apomixis where sporophytic cells of the ovule directly give rise to embryos nourished by the endosperm of autonomous or sexual origin. Molecular basis of adventive embryony is poorly understood. This is the first report of transcriptome analysis of developing ovules during citrus nucellar embryo initiation. Suppression subtractive hybridization (SSH) and microarray were employed to find differentially expressed transcripts during early stages of seed development. Results of the two approaches were congruent for enriched GO terms (response to stress and signalling) and differential gene expression levels. SSH added 85 novel unigenes to the citrus EST database and revealed 443 new unigenes not represented in the Citrus Affymetrix array. Examination of the most-specific-GO-terms showed upregulation (123) and over-representation of transcripts related to abiotic stress (HSPs, HSF, pyruvate decarboxylase) and downregulation (92) of signalling-related (kinases and WRKYs) transcripts at the pre-anthesis stage of ovule development, when nucellar cells destined for embryogenesis begin to differentiate. Microarray and quantitative real-time polymerase chain reaction (qRT-PCR) analyses of genes present in the polyembryony locus identified significant differential expression of heat-shock-related genes and three novel ORFs (orf21, 22, and 25) between ovules of poly- and mono-embryonic varieties. qRT-PCR and microarray data confirmed the up- and downregulation of HSP and WRKY transcripts, respectively, in the pre-anthesis stage ovules of the polyembryonic variety. Based on differential gene expression pattern, a possible mechanistic sequence of polyembryony is presented.
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