Investigation of sex-linked DNA regions in Cycas species using amplified fragment length polymorphism markers

Abstract

Cycads are dioecious plant species in which the male and female organisms can be visually identified only by their reproductive organs or cones. However, these plants must grow for several years before producing their first cones. DNA markers constitute an efficient tool that facilitates sex differentiation of cycads at an early age. Therefore, this research used amplified fragment length polymorphism (AFLP) markers to scan the male and female genomes of four Cycas species (C. chamaoensis, C. clivicola, C. edentata and C. siamensis) for polymorphic regions and their later utilization as specific markers. The DNA fingerprints of the male and female groups of each species were compared using 64 combinations of AFLP primers. Three primer pairs did not yield any DNA bands. Sixty-one primer pairs yielded 6–58 DNA bands, with an average of 30 bands per primer pair. In total, 302 polymorphic bands were obtained with 6.06%, 3.62%, 3.54% and 2.92% from C. edentata, C. siamensis, C. chamaoensis and C. clivicola, respectively. Next, 54 primer pairs designed from sequences of 141 selected polymorphic bands were tested on pooled and individual DNA. Though 35 primer pairs yielded a single amplified product, none could differentiate between male and female plants.