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spontaneously shorten and possibly rearrange, complicating their use for mapping
mutations. In addition, mosaicism can complicate the interpretation of comple-
mentation experiments.
III. Characterization of Chromosomal Rearrangements
The majority of genomic rearrangements currently used in C. elegans research
have been generated and captured in broad mutagenesis screens ( Girard et al., 2007 ).
A consequence of this is that mutated genomes carry a significant number of single
nucleotide differences. Not all of these mutational events will be eliminated by
outcrossing and recombination. In many cases, accrued mutations will be retained
because of suppressed crossing over in the region of the genomic rearrangement.
Thus, the more fully characterized the rearrangement is the better it is for experi-
mental interpretation. Many rearrangements have not yet been characterized at the
molecular level. Analysis has been limited to general identification of the nature and
extent of the rearrangement using relatively low-resolution labor-intensive genetic
techniques. High-resolution characterization at the nucleotide level is now a prac-
tical option and a desirable approach to prevent problems arising from inconsistent
or incorrect results.
A. Restriction Enzyme Digestion of Single Nucleotide Polymorphisms (snip-SNP)
Single nucleotide differences between strains (SNPs) provide a high-resolution
mapping tool and a significant advance over traditional mapping methods
( Wicks et al., 2001 ). SNP mapping relies upon the availability of small changes at
the genomic level between two closely related, yet distinct isolates of the same
species. In C. elegans there is a sufficiently distant, though still compatible isolate,
CB4856 (Hawaiian) ( Flibotte et al., 2009; Jakubowski and Kornfeld, 1999; Koch
et al., 2000; Swan et al., 2002; Wicks et al., 2001 ). Several million years of
evolutionary drift between the wild-types Bristol N2 and Hawaiian CB4856 have
resulted in a large number of base pair changes at intervals of 1000 bp on average.
Sequence alterations of various types are substrate for this type of analysis,
including single-nucleotide changes, small deletions or insertions. Small deletions
and insertions can be detected using a simple PCR assay. Nucleotide changes that
alter a restriction site result in fragment length difference that can readily be
observed by gel electrophoresis (snip-SNP).
Using SNPs to map deletions is relatively straightforward and has been applied to
several deficiency strains ( Kadandale et al., 2005; Zhao et al., 2008 ). A strain
carrying the deletion to be characterized is mated to the Hawaiian isolate.
Suitable snip-SNPs are then analyzed from DNA extracted from the resulting het-
erozygous progeny ( Fig. 7 ). The deleted region of the genome will effectively be
homozygous for the Hawaiian snip-SNPs while the rest of the genome will show a
heterozygous pattern.
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