Paramecium tetraurelia

The DNA from the macronucleus of Paramecium tetraurelia is being sequenced at Genoscope using a whole genome shotgun sequencing strategy. Several libraries with different insert sizes (3 kb, 5 kb and 10 kb) have been constructed from macronuclear DNA. This comes from purified macronuclei from stock strain d4-2. The macronuclei are easy to obtain by simple centrifugation due to their size and density. The cloning of the large inserts (10 kb), which provide large scale clone links during the assembly, required a special development effort because of the high A + T content of the Paramecium genome (70-75%, and up to 85% in the intergenic regions).

The ensemble of the sequences read will represent a 10X coverage of the haploid macronuclear genome, for which the size is estimated at about 100 Mb. These reads are assembled with an assembler developed at Genoscope. The elimination of repeat sequences during the formation of the macronucleus and the apparent absence of duplicated regions will facilitate the assembly, for which no mapping data is available. Problems may occur with the chromosome ends: each of the chromosomes of the macronucleus (there are about 350 chromosomes) is present in a thousand copies, which may differ slightly at the points where the telomeres are added. These problems should be easy to resolve.

The assembly is unlikely to lead to a number of contigs, or even of scaffolds, which would be equivalent to the number of chromosomes in the macronucleus. Nevertheless, there should be only a few gaps with sequencing at 10X coverage. The sequencing of the micronucleus of Paramecium, which is in project phase, should facilitate ordering of the sequences of the macronucleus.

Contacts:
Patrick Wincker (Genoscope) - Linda Sperling, Jean Cohen (CGM)