Abstract
Genome sequencing requires insertion of random fragments of the sequenced organism’s DNA into a unicellular host, most often Escherichia coli bacteria. This manipulation was found in the past to be analogous to naturally occurring horizontal gene transfer, and moreover has proved valuable to understanding toxicity of foreign genetic elements to E. coli. Sequencing of the Caenorhabditis elegans genome was similarly achieved via DNA transformation into E. coli. However, numerous attempts have proven a significant percentage of the genome unclonable using bacteria, although clonable via yeast. We examined the genomic segments that were not clonable in bacteria but were clonable in yeast, and observed that, in line with previous hypotheses, such sequences are more repetitive on average compared with the entire C. elegans genome. In addition, we found that these gap-sequences encode significantly more for DNA transposons. Surprisingly, we discovered that although the vast majority of the C. elegans genome is clonable in bacteria (77.5%), almost all the thousands of sequences that encode for PIWI-interacting small RNAs, or 21U-RNAs (91.6%) were only clonable in yeast. These results might help understanding why most piRNAs in C. elegans are physically clustered on particular loci on chromosome IV. In worms and in a large number of other organisms, piRNAs serve to distinguish “Self” from “Non-Self” sequences, and thus to protect the integrity of the genome against foreign genetic elements, such as transposons. We discuss the possible implications of these discoveries.
Highlights
During the 1990s and early 2000s, there was a race to sequence the human genome (Collins et al, 2003)
As part of sequencing the C. elegans genome, the whole genome of the worm was randomly broken into overlapping fragments, which were transformed into Escherichia coli bacteria through the use of very large cloning vectors termed cosmids and fosmids (Coulson et al, 1986; Kim et al, 1992; Perkins et al, 2005)
Even though there have been numerous attempts over the past 20 years at filling the gaps using cosmids and fosmids [including a consortium dedicated to creating a library of fosmids that would cover the entire genome (Perkins et al, 2005)], none have been successful in covering these gaps
Summary
During the 1990s and early 2000s, there was a race to sequence the human genome (Collins et al, 2003). An unexpected result of the development of the different DNA cloning techniques was the accumulation of negative results, failed cloning attempts that allow gaining insight regarding barriers for genomic information transfer between organisms. Even though there have been numerous attempts over the past 20 years at filling the gaps using cosmids and fosmids [including a consortium dedicated to creating a library of fosmids that would cover the entire genome (Perkins et al, 2005)], none have been successful in covering these gaps. About 20% of the genome could not be cloned in bacteria in spite of these repeated efforts. Throughout this manuscript we will refer to such sequences as “gap sequences.”
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