Samenvatting

Bryophytes are small, haploid, non-vascular plants, which can be divided in three distinct phyla; the mosses (Musci), the liverworts (Hepaticae) and the hornworts (Anthocerotae) (Gradstein & Melick, 1996), (Touw, Kruijer, & Stech, 2016). They are valuable indicators of environmental chance, due to the fact that water easily enters the plant body carrying harmful contaminants and pollutions and because they occur in almost all environments (Gignac, 2001). Species from the liverwort phylum are the objective of this research, and it is tried to verify whether the altitudinal gradients of mount Gede in the Gede-Pangrango National Park (West-Java) influence genetic diversity. This is done by combining morphological taxonomy with molecular analysis using DNA barcoding, a technique that uses standardized parts of an organisms genome for species identification and phylogeny (Stech & Quandt, 2010). The marker regions trnL-F, part of the chloroplast DNA, and ITS, part of the nuclear ribosomal DNA, are commonly sequenced when assessing plant phylogeny (Quandt & Stech, 2004).
The taxa under investigation are various Bazzania species and the Heteroscyphus coalitus, both collected in 2017 on the aforementioned mountain. Initially, it was needed to justify the use of column-based, automatic DNA extraction over DNA extraction with CTAB-lysis, which is especially designed for plant material. Then, PCR amplification of the genetic markers was optimized prior to high-throughput processing and subsequent Sanger sequencing. Afterwards, sequences were edited and aligned, and then assessed for their molecular phylogeny by using the statistical approach of Bayesian Inference, incorporating the a Metropolis-coupled Markov chain Monte Carlo (MCMCMC) analysis. The ensuing phylogenetic trees were used to gain insight on intraspecific variability.
High-throughput DNA extraction was justified, and basic PCR reaction mixtures were found to be optimal for either trnL-F or ITS marker amplification. It was established that the combination of morpho-molecular analysis could be used to identify yet unidentified Bazzania species, but should be adjusted in order to analyse Heteroscyphus coalitus, as the current method resulted in a low PCR yield. The technique used allowed for the identification of previously unidentified Bazzania species, and could so help visualise the distribution of the Bazzania species over the altitudinal gradient on mount Gede. Also, the method used visualised intraspecific variation on some Bazzania species, which could subsequently be linked to their altitudinal distribution. Recommendations for future research focussed on the expanding of the data-set, the incorporation of additional analysis of phylogenetic relationships and the use of other techniques that could aid defining the influence of environmental factors of liverwort growth.