Direct amplification and NaOH extraction: two rapid and simple methods for preparing bryophyte DNA for polymerase chain reaction (PCR)

Abstract

PCR (polymerase chain reaction) has become one of the most important techniques used in molecular systematics. Generally, the methods applied to isolate DNA for PCR amplification depend on multiple steps to isolate and clean the final product. This involves considerable effort and time when many samples have to be processed and poisonous organic solvents are often needed (phenol, chloroform etc.). In this contribution, two very rapid and simple techniques intended for use in higher plants are shown to be useful in bryophyte molecular biology: direct amplification from plant tissues and the NaOH extraction method. In 15 of the 17 investigated bryophytes (two hepatics and 15 mosses) the trn LUAA intron of the chloroplast DNA was successfully amplified by the direct approach, while the NaOH extraction method gave amplifiable DNA in all 17 species. DNA amplified by both methods was successfully used in cycle sequencing.