A comparison of different protocols for RAPD analysis of Littorina

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Abstract

Randomly amplified polymorphic DNA (RAPD) is a fast and useful method of genome marking that is useful for studies of, for example, parentage, mating patterns, taxonomy of sibling species and intra-specific population genetic structures. Here we compare three different procedures for extracting high molecular weight genomic DNA; phenol-chloroform, hexadecyltrimethyl ammonium-bromide (CTAB) and Chelex 100. Double phenol-chloroform and CTAB extractions both generated high amounts of high quality DNA while Chelex 100 failed to do so. We also compared PCR-amplification with different concentrations of template DNA and found that 1–2 ng per 25 μ1 of amplification cocktail gave the best results. Amplifying DNA prepared by the three extraction methods revealed that DNA extracted with double phenol-chloroform gave the clearest bands. The double phenol-chloroform extraction seems thus the most suitable extraction method for RAPD in Littorina, however Chelex may be the only method useful for extracting DNA from very small individuals, for example, pre-hatching stages.