Full Length Research Paper

A simple method of DNA extraction from coffee seeds suitable for PCR analysis

Manoj Kumar Mishra^{*1, 2}, N. Sandhya Rani¹, A. S. Ram¹, H. L. Sreenath¹ and Jayarama¹

¹Central Coffee Research Institute, Coffee Research Station --577117, Dist – Chikmagalur, Karnataka, India.

²Laboratory of Genetics, Departimento di Biologia Universita di Trieste, P.le Valmaura 9,  34100 Trieste, Italy.

*Corresponding author. E-mail: manoj133@yahoo.com. Tel: + 39.040812237, Fax: + 39.040810860.

Accepted 19 April, 2007

High quality genomic DNA was successfully extracted from coffee seeds using a simple protocol devoid of liquid nitrogen or freeze-drying and proteinase K. The isolated DNA was quantified using spectrophotometer and using agarose gel electrophoresis. The DNA was free from polysaccharides, polyphenols, RNA and other contaminants. The quantity of DNA ranged from 180 to 630 μg/g of seed powder. Quality of DNA was confirmed by digestion using EcoRI, HindIII and PstI restriction endonucleases and complete digestion was observed. PCR with random decamer primers and consensus primers of mitochondria and chloroplast DNA and PCR-RFLP revealed the suitability of the DNA for PCR based marker techniques including diagnostics.

Key words: Coffee seed, DNA extraction, RAPD, PCR-RFLP, molecular diagnostics.