Worm Breeder's Gazette 13(5): 18 (February 1, 1995)

These abstracts should not be cited in bibliographies. Material contained herein should be treated as personal communication and should be cited as such only with the consent of the author.

PCR Fingerprinting in C. elegans and (More or Less) Related Species.

Mark Benecke, Einhard Schierenberg

Zoologisches Institut der Universitat, D-50923 Koln

We are interested in phylogenetic relationships among
lower invertebrates. PCR-based technology of DNA fingerprinting(1)
appears to be promising, relatively cheap and time/energy
saving (compared to sequencing or multilocus fingerprinting).
As a pilot experiment four samples of genomic DNA from N2,
a C. elegans isolate from Australia, Rhabditis tripatitas
and Dugesia gonocephala (Turbellaria) were diluted up
to 0,1 ng/microL. DNA was subjected to PCR using several
short synthetic oligonucleotides and two different taq
polymerases. The amplification protocol differs from
the standard PCR conditions (2) in that only a single oligonucleotide
primer is employed per PCR reaction and no prior knowledge
of the genome is required. Short primers, like short oligonucleotides
in multilocus fingerprinting(2), detect several DNA
sites. Priming sites which are located close to one another
in inverted orientation are the targets for RAPD (Random
Amplified Polymorphic DNA) analysis. The technique essentially
scans a genome for these small inverted repeats and amplifies
intervening DNA segments of variable length. For any given
template-primer-taq polymerase combination (TPPC),
the amplification products are reproducible-'. Amplification
products were resolved for 4 hours on I . 8% agarose gels
containing EtBr.
Our RAPD pattems indicate that laboratory strains due
to inbreeding (2) constitute essentially homogeneous
worm populations without visible differences (with this
techniques) between individual worms. The closer strains
are related to each other, the better their RAPD pattems
should resemble each other. With the two primers tested
the RAPD patterns of the two C. elegans strains appear to
differ in only one band, while those of the two other species
differ to a much higher extent. This finding is similar
to that with our multilocus fingerprints. Depending on
the chosen TPPC (mentioned above) we find different banding
patterns. These probably reflect variations in the informational
content(2). The observation that two randomly chosen
primers (out of hundreds) allow amplification of our worm
DNA is consistent with the supposition that about 80% of
all RAPD primers are suitable for this type of analysis
in worms(4).
Our first results suggest that we can distinguish strains
by their defined characteristics visualized by the fingerprint
technique. The results of our dilution series indicate
that DNA of one C. elegans individual is sufficient for
several RAPD experiments. In conjugation with analysis
of developmental features (5) we plan to use this assay
to analyze phylogenetic relation ships among nematodes.
In addition, we would like to explore to which extent the
RAPD technology 1 is suitable for comparative studies
on higher taxonomic levels.