Final Report Summary - GENSENSOR-NANOPARTS (Nano-biotechnical components of an advanced bioanalytical microarray system)
The project aimed to contribute to the further development of the deoxyribonucleic acid (DNA) microarray field, which is scientifically and commercially an important part of the genomic bioscience revolution ongoing for the 15 years preceding the project. The main objective of the GENSENSOR-NANOPARTS project was to fill the gap in DNA analytics still existing between extracting nucleic acids from the biological material and analysing it by using microarrays. It is of general importance for biological research, clinical diagnostics and industrial applications, irrespective whether the microarrays are used for gene expression analysis or for identifying and genotyping organisms.
The work performed can be summarised by describing the end results, which consist of the following achievements:
1. unique DNA sequences highly specific for each of the following five microorganisms were defined in silico at the whole genome level: Salmonella enterica ssp., Enterica serovar Typhimurium LT2, Salmonella enterica ssp., enterica serovar Typhi, Listeria monocytogenes Li 2, Staphylococcus aureus ssp. Mu50, and Staphylococcus epidermidis;
2. polymerase chain reaction (PCR) primers, DNA captures for microarrays, locked nucleic acid (LNA) captures for nano-beads and target DNA sequences were positively selected experimentally and best LNA-DNA pairs were used as model system for selective extraction experiments;
3. different types of LNA-coupled magnetic nano-beads were developed and used to "fish" target DNA of the model organism mentioned above;
4. different extraction media like buffer, milk, cell lysate and mixtures containing chaotropic agents to free the nucleic acids from other components of the biological material to be extracted are demonstrated to allow the sequence selective extraction.
Experimental difficulties in using the PCR as extraction assay in the presence of an excess of nano-beads lead to the conception of a new padlock-based DNA-analytical method and to its experimental realisation (not yet finished).
These achievements correspond to the essentials of the objectives of the GENSENSOR-NANOPARTS project, which was therefore considered to be fulfilled successfully.
The work performed can be summarised by describing the end results, which consist of the following achievements:
1. unique DNA sequences highly specific for each of the following five microorganisms were defined in silico at the whole genome level: Salmonella enterica ssp., Enterica serovar Typhimurium LT2, Salmonella enterica ssp., enterica serovar Typhi, Listeria monocytogenes Li 2, Staphylococcus aureus ssp. Mu50, and Staphylococcus epidermidis;
2. polymerase chain reaction (PCR) primers, DNA captures for microarrays, locked nucleic acid (LNA) captures for nano-beads and target DNA sequences were positively selected experimentally and best LNA-DNA pairs were used as model system for selective extraction experiments;
3. different types of LNA-coupled magnetic nano-beads were developed and used to "fish" target DNA of the model organism mentioned above;
4. different extraction media like buffer, milk, cell lysate and mixtures containing chaotropic agents to free the nucleic acids from other components of the biological material to be extracted are demonstrated to allow the sequence selective extraction.
Experimental difficulties in using the PCR as extraction assay in the presence of an excess of nano-beads lead to the conception of a new padlock-based DNA-analytical method and to its experimental realisation (not yet finished).
These achievements correspond to the essentials of the objectives of the GENSENSOR-NANOPARTS project, which was therefore considered to be fulfilled successfully.
