Carlos Fernando Prada Quiroga is Biologist, MSc in genetics and evolution of Universidade Federal de Sao Carlos in Brazil. He has completed his PhD in genetics at the age of 32 years from Universidad Autónoma de Barcelona, Spain. He has had teaching experience in countries like 2 Brazil, Spain and Colombia. He is Associate Professor and director of Bacteriology and Clinical Laboratory research group. He has published more than 10 papers in reputed journals. His research focuses on the evolutionary origin of genomic rearrangements and its relationship to different human pathologies, using mainly bioinformatics tools.
Mitochondrial genome plays a variety of important roles, including the generation of ATP through respiration and oxidative phosphorylation (OXPHOS), production of reactive oxygen species (ROS), and initiation and execution of apoptosis. Therefore, variation in these genes can directly influence metabolic performance. The mitochondrial genome presents a gene structure relatively stable through the evolution of species with a relatively low rate of gene rearrangements compared to the nuclear genome. The recent increase in the number of sequenced mitochondrial genomes of hundreds of species, it has become clear that some groups of species the gene order is more conserved than in other groups. For example, mammals and birds have relatively stable gene orders, while in amphibians, reptiles and fish rearrangements are more common. Recent studies have found that the mammalian mitochondrial genome has a higher rate of rearrangements than expected. 244 mitochondrial events were identified: Inversions, deletions and non-homologous regions (non-HR: less than 10% nucleotide identity). Inversions were found at a frequency of 84.0 % (205/244). These inversions, classified as microinversions (<1Kb) are recurrent in tRNAs genes; deletion are detected on D-loop region and tRNAs; and non-HR located on control region. Based on the breakpoints (BP) of mitochondrial inversions, there were postulated fourteen fragile regions in the human mitochondrial genome. Over 250 pathogenic human mtDNA mutations have been characterized to show the cause a wide variety of diseases with heterogeneity of phenotypes and a variable age of onset. Most of deletions and duplications BP reported in the human mitochondrial genome match with the fragile regions previously postulated. These findings could indicate the important role of tRNA in the origin of rearrangements in the human mitochondrial genome and in mammals in general.
Laura Hill is a Senior Scientist in the Design and Development department at QIAGEN Manchester, UK. She has significant experience of companion diagnostic development, specialising in primer and multiplex assay design, feasibility studies and novel technologies. She previously worked at Life Technologies on the development and manufacture of primer mixes for DNA testing kits. She has a degree in Genetics from the University of Liverpool.
The Modaplex system is a fully automated platform that combines a real time PCR module with a capillary electrophoresis detection module. The combination of these two modules allows for a large number of targets to be detected in a single reaction. Multiplexing systems that allow such a broad range of targets to be detected are desirable in companion diagnostic assays; to reduce intrusive procedures to patients, by requiring less sample material, and to reduce laboratory processing time. To investigate how the Modaplex system can make use of these benefits, we have undertaken a feasibility study to combine the detection of 42 mutation targets and 2 control targets in a single reaction. Gene panels comprising of 13 KRAS mutation targets and 29 EGFR mutation targets are the focus of this multiplexing challenge. EGFR and KRAS are key molecules in the MAPK cellular signaling pathway. Since mutations in these genes alter cellular functions, determining the mutation status is a key requirement for personalised cancer therapies. The primer designs for the 44-plex assay have initially been tested using synthetic oligonucleotides and will be further challenged with the use of FFPE extracted clinical samples. This feasibility study will demonstrate the multiplexing capabilities of the Modaplex system to meet the growing market need for quicker, less intrusive mutation detection.