Transcriptome sequencing and comparative analysis of cucumber flowers with different sex types

Abstract

BackgroundCucumber, Cucumis sativus L., is an economically and nutritionally important crop of the Cucurbitaceae family and has long served as a primary model system for sex determination studies. Recently, the sequencing of its whole genome has been completed. However, transcriptome information of this species is still scarce, with a total of around 8,000 Expressed Sequence Tag (EST) and mRNA sequences currently available in GenBank. In order to gain more insights into molecular mechanisms of plant sex determination and provide the community a functional genomics resource that will facilitate cucurbit research and breeding, we performed transcriptome sequencing of cucumber flower buds of two near-isogenic lines, WI1983G, a gynoecious plant which bears only pistillate flowers, and WI1983H, a hermaphroditic plant which bears only bisexual flowers.ResultUsing Roche-454 massive parallel pyrosequencing technology, we generated a total of 353,941 high quality EST sequences with an average length of 175bp, among which 188,255 were from gynoecious flowers and 165,686 from hermaphroditic flowers. These EST sequences, together with ~5,600 high quality cucumber EST and mRNA sequences available in GenBank, were clustered and assembled into 81,401 unigenes, of which 28,452 were contigs and 52,949 were singletons. The unigenes and ESTs were further mapped to the cucumber genome and more than 500 alternative splicing events were identified in 443 cucumber genes. The unigenes were further functionally annotated by comparing their sequences to different protein and functional domain databases and assigned with Gene Ontology (GO) terms. A biochemical pathway database containing 343 predicted pathways was also created based on the annotations of the unigenes. Digital expression analysis identified ~200 differentially expressed genes between flowers of WI1983G and WI1983H and provided novel insights into molecular mechanisms of plant sex determination process. Furthermore, a set of SSR motifs and high confidence SNPs between WI1983G and WI1983H were identified from the ESTs, which provided the material basis for future genetic linkage and QTL analysis.ConclusionA large set of EST sequences were generated from cucumber flower buds of two different sex types. Differentially expressed genes between these two different sex-type flowers, as well as putative SSR and SNP markers, were identified. These EST sequences provide valuable information to further understand molecular mechanisms of plant sex determination process and forms a rich resource for future functional genomics analysis, marker development and cucumber breeding.

Genome-Wide Mapping of Structural Variations Reveals a Copy Number Variant That Determines Reproductive Morphology in Cucumber

“…The duplicated region contains the ACS1 (Csa6G496450) gene as well as Csa6G496950 (a gene of unknown function), Csa6G496960 (encoding a truncated MYB transcription factor), and part of Csa6G496970 (encoding a branched-chain amino acid aminotransferase) ( Figure 3D). The previously reported differentially expressed genes between female and male flowers, such as auxin-related and short-chain dehydrogenase or reductase genes related to the development of unisexual flowers (Huang et al, 2009;Guo et al, 2010), were not found in this region; thus, they are likely downstream floweringrelated genes. To further confirm the association between the CNV and gynoecy, we resequenced six additional gynoecious lines (GCA07, GCA09, HAU106, IVF204, HAU107, and SJ08).…”

Complexity of the Alternative Splicing Landscape in Plants

“…Innovations in sequencing RNA (RNA-seq) led to the generation of millions of short sequence reads of cDNAs derived from poly(A)-enriched mRNA from different tissues or conditions in a cost effective manner, which were then mapped onto unique locations on the genome to predict AS events (Wang et al, 2009a;Filichkin et al, 2010;Hallegger et al, 2010;Marquez et al, 2012). This has been used to analyze the extent of AS and to identify novel introns and splice variants in Arabidopsis (Filichkin et al, 2010;Marquez et al, 2012;Syed et al, 2012), rice (Oryza sativa; Zhang et al, 2010b), cucumber (Cucumis sativus; Guo et al, 2010), grape (Vitis vinifera; Zenoni et al, 2010), and Brachypodium distachyon (Walters et al, 2013). These studies have revealed that up to 60% of introncontaining genes are alternatively spliced.…”

Profiling of the transcriptome of Porphyra yezoensis with Solexa sequencing technology

“…Expressed sequence tags (ESTs) are important for studying the structure and function of functional genome. ESTs have played significant roles in accelerating gene discovery, improving genome annotation, elucidating phylogenetic relationships, facilitating breeding programs by providing SSR and SNP markers, analyzing large-scale gene expression and rapidly identifying transcripts involved in specific biological processes [6]. During the last few years, EST analysis of P. yezoensis has been conducted in many laboratories [7][8][9], and approximately 22000 ESTs have been deposited into the dbEST database at NCBI.…”