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Gene expression analysis in pre- and early post-anthesis stage ovules of Citrus sinensis

ABSTRACT: Nucellar embryony is a form of apomixis found in citrus where somatic nucellar cells differentiate into embryos and are included in the seed resulting from the normal sexual process. The nucellar cells giving rise to adventive embryo start proliferating prior to anthesis and fully differentiate obtaining nourishment from sexually derived endosperm. To identify transcripts differentially expressed during nucellar embryo initiation we have taken RNA samplesfrom different developing stages of ovules from polyembryonic (cv. Vaniglia Sanguigno) and monoembryonic (cv. Temple) cultivars. We used microarray for a detailed analysis of global gene expression during nucellar embryony initiation and development. We have further validated the differentially expressed genes using qRT-PCR.

ORGANISM(S): Citrus sinensis

PROVIDER: GSE36188 | GEO | 2012/03/01

SECONDARY ACCESSION(S): PRJNA151663

REPOSITORIES: GEO

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Project description:Somatic embryogenesis in nucellar tissues is widely recognized to induce polyembryony in major citrus varieties such as sweet oranges, satsuma mandarins and lemons. This capability for apomixis is attractive in agricultural production systems using hybrid seeds, and many studies have been performed to elucidate the molecular mechanisms of various types of apomixis. To identify the gene responsible for somatic embryogenesis in citrus, a custom oligo-DNA microarray including predicted genes in the citrus polyembryonic locus was used to compare the expression profiles in reproductive tissues between monoembryonic and polyembryonic varieties. The full length of CitRKD1, which was identified as a candidate gene responsible for citrus somatic embryogenesis, was isolated from satsuma mandarin and its molecular function was investigated using transgenic ‘Hamlin’ sweet orange by antisense-overexpression. The candidate gene CitRKD1, predominantly transcribed in reproductive tissues of polyembryonic varieties, is a member of the plant RWP-RK domain proteins. CitRKD1 of satsuma mandarin comprised two alleles (CitRKD1-mg1 and CitRKD1-mg2) at the polyembryonic locus controlling embryony type (mono/polyembryony) that were structurally divided into two types with or without a miniature inverted-repeat transposable element (MITE)-like insertion in the upstream region. CitRKD1-mg2 with the MITE insertion was the predominant transcript in flowers and young fruits where somatic embryogenesis of nucellar cells occurred. Loss of CitRKD1 function by antisense-overexpression abolished somatic embryogenesis in transgenic sweet orange and the transgenic T1 plants were confirmed to derive from zygotic embryos produced by self-pollination by DNA diagnosis. Genotyping PCR analysis of 95 citrus traditional and breeding varieties revealed that the CitRKD1 allele with the MITE insertion (polyembryonic allele) was dominant and major citrus varieties with the polyembryonic allele produced polyembryonic seeds.

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Gene expression analysis in pre- and early post-anthesis stage ovules of Citrus sinensis

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