Jb. Geller et Da. Powers, SITE-DIRECTED MUTAGENESIS WITH THE POLYMERASE CHAIN-REACTION FOR IDENTIFICATION OF SIBLING SPECIES OF MYTILUS, The Nautilus, 108, 1994, pp. 141-144
The large sample sizes needed for population and biogeographic studies
make sequencing impractical as a means of quantifying genotype freque
ncies. In a previous study, we identified sequence variation in the mi
tochondrial 16S ribosomal gene from mussels in a sibling species group
(Mytilus spp.) that discriminated mussels introduced to the west coas
t of North America (Mytilus galloprovincialis) from native mussels (My
tilus trossulus). In this study, we used site-directed mutagenesis by
the polymerase chain reaction (PCR) to transform a single diagnostic n
ucleotide substitution into a restriction site. By using a large (51 b
p) oligomer as a primer, a size shift in cut and uncut PCR products wa
s visible on an agarose gel. Thus, PCR followed by a restriction diges
tion allowed identification of species-specific haplotypes in hours ra
ther than days. This method is applicable to any study in which rapid
detection of genotypes is desired. We show the presence of M. trossulu
s haplotypes in Monterey Bay, California, extending its southern range
. We also present evidence for heteroplasmic individuals that contain
mitochondrial haplotypes indicative of both species.