Molecular Cytogenetic Techniques for Studying Plants in the Genus Paris from Thailand

Abstract

Paris L. is a genus of herbaceous plants in the family Melanthiaceae. The genus comprises several highly valued medicinal species, such as Paris polyphylla Smith. investigated in the present study. Rhizomes of P. polyphylla have been used in traditional Chinese medicine namely Yunnan Baiyao for treatment of various symptoms, such as inflammation and injury. Due to its medicinal properties, intensive studies of these plants have been conducted with regarding both utilization and conservation of the resources. The present study reports the development of suitable methods for preparing high-quality chromosome spreads suitable for karyotype and fluorescent mapping of chromosome markers. The appropriate pretreatment for arresting metaphase in root tips was ice-water for 30-35 hours. Enzymatic squash method (Anamthawat, unpublished) was proven efficient as a technique of preparation of metaphase spreads. Enzyme K-15 (Pectinase: Cellulase, 300:800 units/10 ml) was used and the optimal time of digestion at 37°C was 12-15 minutes depending on root tips size. For fluorescent in situ hybridization (FISH) mapping of ribosomal genes (rDNA), the optimal temperature of denaturation was at 88-89°C for 10 minutes. The method developed in the present study will be used for the cytotaxonomical investigation of Paris taxa from Thailand.