We have been analyzing the gene expression patterns of the nematode C.elegans systematically by EST sequencing and whole mount in situ hybridization. All the results are integrated with the genome map based at NEXTDB
http://nematode.lab.nig.ac.jp. As an extension of the project we performed whole transcriptome analysis using the SOLEXA sequencer. Poly-A RNA were prepared from 4 samples; embryo, larvae, and adult of wild type hermaphrodites, and mixed stage population of male rich worms and then converted to double-stranded cDNA using the random hexamers primers. The resulting cDNA were subjected to the SOLEXA sequencer, and then the short reads (effective length of 32 bases) were mapped onto the C.elegans genome by the ELAND and our newly developed mapping program that allowed a gap to detect the reads from exon-intron junctions. As to the 5'' end, splice-leader capped sequences were detected and mapped. These data are integrated in NEXTDB and can be viewed as to exon structure, expression level, developmental stages, sex and so on. In parallel, we tried the de novo assembling of the transcripts using the total 1Gb reads from all the samples by the VELVET assembler which was developed at EBI, and obtained total 36,000 contigs with maximum length of 5 kb and N50 of 450 bases. Although an alternatively spliced region should be assembled to a branch structure, this assembler generates the branches and the trunk as separate contigs, which may cause the relatively short contigs. However, even with such short contigs we have made many corrections on the gene models, e.g., exon structure and splicing patterns. We are also conducting the genome sequencing of the other nematode Diploscapter coronatus in collaboration with Dr. Einhard Schierenberg at Univ. of Koeln; the nematode shows an interesting cell cleavage and arrangement pattern to gastrulation stage. Genomic DNA of the nematode was analyzed by the SOLEXA sequencer and 6Gb paired-end reads (each length of 36 bases) were obtained. They are being assembled in a hybrid manner together with low coverage Sanger shotgun reads. We have already done EST analysis of the nematode and identified 10,000 unique genes in which 5,800 were found to have homologs or orthologs to C.elegans. We will combine these genome and EST data.