An RNA-Seq Based Reference Transcriptome for Citrus

RNA-seq studies in citrus have focused on physiological processes relevant for fruit quality and productivity of the major species, especially sweet orange. However, little or no attention has been paid to vegetative or reproductive tissues, and most of the Citrus species have never been analyzed. In this work we characterized the transcriptome of vegetative and reproductive tissues from a wide representation of Citrus species, with members from all the main phylogenetic groups. Our aims were to acquire a complete view of the citrus transcriptome landscape, to improve previous functional annotations and to obtain genetic markers (SNPs and Indels) associated to genes of agronomic interest.

28 samples were used for RNA-Seq analysis, obtained from 12 citrus species: C. medica, C. aurantifolia, C. limon, C. bergamia, C. clementina, C. deliciosa, C. reshni, C. maxima, C. paradisi, C. aurantium, C. sinensis, and Poncirus trifoliata. Four different organs were analyzed: root, bark, leaf, and flower.

A total of 3421 million Illumina reads were produced and mapped against the reference C. clementina genome sequence. Transcript discovery pipeline revealed 3326 new genes, the number of genes with alternative splicing was increased to 19739, and a total of 73797 transcripts were identified, which represents a 3 fold increase with respect the previous results from C. clementina.

Differential expression studies between the 4 tissues were performed with EDGE-R. Variants discovery analysis revealed the presence of indels and SNPs in genes relevant for fruit quality and productivity that could be used for breeding. Some relevant pathways in citrus were also analyzed: flavonoids and flavonols, ethylene and auxin.