Optimizing DNA extraction protocols for bryophytes: Insights from Orthotrichaceae.

Premise: Extracting high-quality DNA from challenging plant tissues can be hindered by high levels of phenolics, carbohydrates, and other compounds that bind to or coprecipitate with DNA. Bryophytes, due to their small size, tendency to intermix, biochemical richness, and capacity to absorb environmental pollutants and heavy metals, pose unique challenges that further complicate proper DNA isolation. Methods and Results: We compared cetyltrimethylammonium bromide (CTAB)-based extraction protocols and evaluated step-by-step modifications aimed at maximizing DNA yield and purity for Orthotrichaceae, one of the most diverse families of bryophytes. The final protocol was also evaluated on representatives of other bryophyte families as a preliminary exploration of its broader applicability. Conclusions: We present an optimized, rapid, and efficient DNA extraction protocol that yields high-quality DNA suitable for high-throughput sequencing and molecular analyses. The presented extraction protocol is likely to work well for other bryophyte taxa and may be of value for molecular analyses involving recalcitrant samples.