Wanted

PhD student wanted. Scholarship from the "Young researcher" program is available:

• Applications for Scholarship: 04.jun.2010 do 09.jul.2010
• Start: October 2010
• Duration: 3.5 year studentship
• Mentor: dr. Aleš Snoj
• Provisional title of PhD: Genetic background responsible for rock-boring mechanisms in date mussel (Lithophaga lithophaga)
• Detailed description of research work and objectives are available on request (ales.snoj@bf.uni-lj.si)

The objective of the work-package is to define the genetic background responsible for rock-boring mechanisms in date mussel (Lithophaga lithophaga) and assess the possibilities for regulation of the involved genes.

Calcareous rock boring of date mussel is an energy demanding action. Restricted water circulation caused by specific microhabitat of the species provides only a limited access of nutrition needed for date mussel growth. These factors are probably one of the main reasons causing exceptionally slow growth of date mussels, which need approx. ten times more time for reaching the same body size as mussel shell (Mytilus edulis). Understanding the mechanisms responsible for rock boring would enable the recognition and identification of specific factors – either those referring to date mussel physiology, environmental ones (i.e., influence of substrate) or interaction of both – having influence on development of the date mussel and its life history and growth capacity. Although the boring mechanisms of the date mussel have already been studied, the actual mechanism is still debated. It was suggested the boring mechanism is based on chemical decomposition of CaCO3, which could be attributed to some Ca-binding lipoproteins secreted from pallial gland acting as chelating agents. According to another hypothesis, pH decrease due to excretion of metabolic CO₂ should be capable of CaCO₃ dissolution. In either case, boring mechanism relays upon specific genetic background, which may be determined by the existence of specific genes unique to rock boring organisms or specific level of gene expression varying between boring and non-boring organisms. Accordingly, genes being involved in rock boring process (i.e., candidate genes) are expected to be found either in pallial gland or in metabolically active tissue (i.e., pallium). The elucidation of the mechanisms responsible for rock boring will provide the insights into how these mussels adapt to harsh growing environment and offer possibilities for novel commercial production strategies.

To overcome the substantial lack of information, the identification of the candidate genes will be performed by comparison analysis of DNA expression profiles in date mussel and in its phylogenetically related non-boring co-existing equivalent (e.g., M. edulis). To determine differentially expressed genes between L. lithophaga and other comparing equivalents, a suppression subtractive hybridization cDNA library will be performed, using the total mRNA from pallial gland and metabolically active tissue of L. lithophaga and M. edulis. To reduce the number of false positive clones mirror orientation selection (MOS) of the subtracted cDNA library will be performed. In addition, phenotypic differences between the studied mussles will be analysed also at the level of DNA transcription, comparing transcription profiles of different tissues using a MytArray 1.0, a mussel cDNA microarray developed for Mytilus sp.. Differential expression and expression profiles of the ESTs obtained from the subtracted library will be confirmed by qRT-PCR. To obtain full-length cDNA sequences of differentially expressed genes, the 5’ and 3’ rapid amplification of cDNA ends (RACE) system will be used. Exon-intron position will be predicted by comparative approach and primers will be designed to amplify corresponding genomic DNA and to determine the sequence of the entire candidate genes including promoter region. Level of expression of the studied genes will be monitored during the life cycle of the date mussel and the results will be compared and paralleled with corresponding morphological and physiological changes during ontogeny of the species individuals.

Collectively, these investigations are significant as they should reveal novel genes that determine the specific physiological properties of rock boring date mussel and could provide novel strategies for commercial production of date mussels.

As of 2007/08 it is possible for non slovenian citizens to join young researcher program. For practical information contact daliborka.dusanic@bf.uni-lj.si, who joined our department in 2007.

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