Piero Carninci — Page 5

PubMed2002

The Drosophila gene collection: identification of putative full-length cDNAs for 70% of D. melanogaster genes.

Collections of full-length nonredundant cDNA clones are critical reagents for functional genomics. The first step toward these resources is the generation and single-pass sequencing of cDNA libraries that contain a high proportion of full-length clones. The first release of the Drosophila Gene Collection Release 1 (DGCr1) was produced from six libraries representing various tissues, developmental stages, and the cultured S2 cell line. Nearly 80,000 random 5' expressed sequence tags (5' expressed sequence tags [ESTs]from these libraries were collapsed into a nonredundant set of 5849 cDNAs, corresponding to ~40% of the 13,474 predicted genes in Drosophila. To obtain cDNA clones representing the remaining genes, we have generated an additional 157,835 5' ESTs from two previously existing and three new libraries. One new library is derived from adult testis, a tissue we previously did not exploit for gene discovery; two new cap-trapped normalized libraries are derived from 0-22-h embryos and adult heads. Taking advantage of the annotated D. melanogaster genome sequence, we clustered the ESTs by aligning them to the genome. Clusters that overlap genes not already represented by cDNA clones in the DGCr1 were analyzed further, and putative full-length clones were selected for inclusion in the new DGC. This second release of the DGC (DGCr2) contains 5061 additional clones, extending the collection to 10,910 cDNAs representing >70% of the predicted genes in Drosophila.

Animals↗

PubMed2002

A computer-based method of selecting clones for a full-length cDNA project: simultaneous collection of negligibly redundant and variant cDNAs.

We describe a computer-based method that selects representative clones for full-length sequencing in a full-length cDNA project. Our method classifies end sequences using two kinds of criteria, grouping, and clustering. Grouping places together variant cDNAs, family genes, and cDNAs with sequencing errors. Clustering separates those cDNA clones into distinct clusters. The full-length sequences of the clones selected by grouping are determined preferentially, and then the sequences selected by clustering are determined. Grouping reduced the number of rice cDNA clones for full-length sequencing to 21% and mouse cDNA clones to 25%. Rice full-length sequences selected by grouping showed a 1.07-fold redundancy. Mouse full-length sequences showed a 1.04-fold redundancy, which can be reduced by approximately 30% from the selection using our previous method. To estimate the coverage of unique genes, we used FANTOM (Functional Annotation of RIKEN Mouse cDNA Clones) clusters (). Grouping covered almost all unique genes (93% of FANTOM clusters), and clustering covered all genes. Therefore, our method is useful for the selection of appropriate representative clones for full-length sequencing, thereby greatly reducing the cost, labor, and time necessary for this process.

Animals↗