Analysis of rotifer ribosomal gene structure using the polymerase chain reaction (PCR)

Abstract

We have used the polymerase chain reaction (PCR) to selectively amplify 18S ribosomal genes in rotifer taxa from major planktonic clades . In each case, we obtained an amplified product of between 1 .8 and 2 .0 kilobase pairs . We analyzed the PCR products using 6-and 4-base cutting restriction enzymes, comparing fragment mobilities . For example, Brachionus plicatilis (BSL strain) 18S genes have no restriction sites for Hind III or Bam HI and only a single site for Eco RI (all 6-base cutters) . The 4-base cutter Msp I, on the other hand, has at least 4 enzymatic sites, producing fragments between approximately 110 and 460 base pairs in length . Results of this type can be used to differentiate among species and species groups within the Rotifera and can be used as the basis for construction of a broad molecular phylogeny of the group .

Using amplified fragment length polymorphisms (AFLP) to study genetic variability in several freshwater rotifer species

“…Total genomic DNAs were extracted from 50-200 individuals from each rotifer species by the method of Walsh & Starkweather (1993). DNA concentrations were visually estimated on agarose gels by comparison with standard k phage digested by HindIII.…”