Biosystematics of selected Chara species (Charophyta) using amplified fragment length polymorphism

Species of the green algal genus Chara (Charophyta) frequently exhibit a great deal of morphological variability. For the first time, a polymerase chain reaction-based fingerprinting technique, amplified fragment length polymorphism (AFLP), was applied to detect genetic diversity in Chara. A total of 36 primer combinations were screened and the method was optimized for high sample throughput using three different fluorescence-labelled primers simultaneously. Six primer pairs were selected for further investigations and tested with a total of 33 samples of C. hispida, C. intermedia and C. tomentosa from several sampling sites. A total of 1265 different loci were detected. Ninety-nine percent of them showed polymorphisms. Although sample size was small, up to 91% intraspecific polymorphic loci were detected. The analysis included samples of C. intermedia and C. hispida growing in the same habitat, as well as samples from other countries. Although genetic similarity between samples of C. intermedia and C. hispida was high compared to C. tomentosa, clustering analysis revealed clear separation of samples according to morphological determination. Therefore, we disagree with R.D. Wood & K. Imahori (1965. A revision of the Characeae, vol. 1. Monograph of the Characeae. J. Cramer, Weinheim) who classified C. intermedia as a variety of C. hispida. Because of its high sensitivity, AFLP is an ideal tool used for further investigating the biosystematics of the genus Chara.