Transcriptome Analysis of Cucumber Fruit Development Using Next-Generation Sequencing Technologies

Cucumber (Cucumis sativus L.) fruit undergo a typical sigmoidal pattern of growth during early stages of development.  The first few days (0-4dpp) are dominated by cell division, followed by a period of rapid expansion (4-12 dpp) that slows as near full size is reached at 12-16 dpp.  Massively parallel 454 pyrosequencing analysis was used to study changes in gene expression during these early stages of cucumber fruit growth.  Expression libraries were prepared from fruits at 0, 4, 8, 12 and 16 dpp, yielding approximately 1.1 million expressed sequence tag (EST) reads.  Contig assembly resulted in ca. 28,000 contigs representing 82% of the total reads with an average contig length of 834 base pairs (bp) and mean number of 67 reads/contig.  Putative Arabidopsis homologs could be assigned for 85.6% of the contigs with greater than 10 ESTs/contig; another 4-5% had homologs in other species.  Approximately 7% of the contigs did not have homologs, including many highly expressed genes (>100 ESTs/contig), suggesting genes possibly unique to cucumber or cucurbits.  For contigs with Arabidopsis homologs, the overall distribution of putative gene function resembled the Arabidopsis genome.  However, at individual time points there was differential representation of the most highly expressed functional groups.  Gene expression levels were compared to the base timepoint of 0dpp to identify changes unique to the different timepoints.  The 4dpp timepoint showed high expression of cell division and protein synthesis associated genes including cyclins, histones, and ribosome-related genes, all of which decreased at later time points.  The 8dpp timepoint showed high level expression of many growth related transcripts such as aquaporins, phloem associated protiens, and cell structure related genes.  The 12 and 16 dpp fruit showed elevated expression of many abiotic and biotic stress related genes.  Principle component analysis showed that the timepoints fell into three groups based on patterns of gene expression, 0 and 4 dpp; 8dpp; and 12 and 16 dpp.  These groupings also reflect the different stages of growth: cell division, exponential expansion, and post-expansion, respectively.