Genoscope is participating in the study of expressed sequences in the chaetognath Spadella cephaloptera. It has produced a collection of ESTs from a S. cephaloptera cDNA library constructed by the Oceanography Center of Marseille (titer: 5.5 x 10⁶ pfu/ml). The mRNAs were purified from various embryonic stages of this arrow worm (from 0 to 48 hours after hatching). The cDNAs have been cloned in the Lambda-triplex 2 vector; they are between 500 pb and 5 kb in length, with a mean size of around 1 kb. For sequencing, the primary cDNA library has been cloned into bacterial strain BM25.8, which expresses the CRE recombinase and allows the circularization of the phagemid to form a plasmid. Inserts with a size greater than 800 pb have been selected.
Genoscope has sequenced the 5' end of 11 000 clones from this library. The sequences were first submitted to quality control, trimming of vector sequences and elimination of contaminants and artefacts. Annotation was carried out in the Oceanography Center of Marseille. After elimination of the sequences corresponding to 12S and 16S rRNAs, the remaining sequences have been blasted against the protein databases Swissprot (2136 clones identified) and NR (2384 clones identified). The annotation was first performed with Swissprot, then with NR in cases of lack of homology with any Swissprot sequences. In this way, homology relations have been assigned to 2396 clones corresponding to the transcripts of 792 different genes. This annotation procedure did not involve a clustering step, because the high level of polymorphism in the library makes it difficult to distinguish between polymorphic orthologues and paralogues within the clusters. However, the results of clustering tests have validated the annotation.