PhD ThesisDespite being a model organism to study settlement in marine invertebrates, little is
known about the genetics of the barnacle Balanus amphitrite. To fill this gap, cDNA
libraries representative of different developmental stages were generated and sequenced.
Nearly 14,000 genes were annotated, which may represent 2/3 of the species’ total
protein coding regions. The database that was created to allow public access to this
genetic information will profoundly benefit future research aiming to understand the
molecular regulation of development and settlement in this species.
Furthermore, a quantitative real-time PCR assay to study gene expression in B.
amphitrite was designed and validated. Eleven genes were studied for their ability to
normalize qRT-PCR data. Total RNA extracted from seven developmental stages was
reverse transcribed and the expression stability of the selected genes was compared. It
was found that transcripts encoding cytochrome b and NADH dehydrogenase subunit 1
were expressed most stably, and their use to normalize gene expression data is
recommended.
Conflicting evidence exists on the role of bacteria in B. amphitrite settlement. However,
there is a paucity of information on the microbial community naturally associated with
this barnacle. In order to reveal the existence of stable associations, a 16S rRNA-based,
taxon-specific qPCR assay was developed to monitor the preponderance of 5 bacterial
phyla and classes. Furthermore, attempts to profile these qPCR products by DGGE were
made. This new method was applied to characterise the bacterial communities
associated with different B. amphitrite developmental stages and body parts. It was
found that the structure of these communities changed throughout the barnacle life cycle
in a highly reproducible manner. Furthermore, bacteria isolated from the barnacle shell
were capable of inducing settlement of conspecific larvae. The analysis of these
communities at a lower taxonomic level should confirm if any of these ecologically
important bacteria are vertically transmitted.Natural Environment Research Council, UK:
The Boehringer Ingelheim Fonds:
The European Community, The Marine Genomics Network:
The Office of Naval Research,USA:
The Prime Minister Initiative, UK