Abstract

Novell Chip-based Extraction of miRNA

Gregory Dame, Group Leader, University of Freiburg

MiRNAs are small (20 to 23 nucleotides in length) non-coding RNAs that regulate numerous essential cell functions. They operate by targeting messenger RNAs for cleavage or translational repression and it is known that their expression profiles classify human cancers. In this study we present the development of a fast on-chip miRNA extraction system. The chip-system is capable of extracting miRNAs from tumor cell cultures, fresh frozen cells and archive material samples by on-chip gel electrophoresis. Due to the size-limiting effect of this process, ribosomal RNA (rRNA), messenger RNA (mRNA) or genomic DNA are excluded. Both the duration and efficiency of the on-chip extraction process are compared to gold standard commercial extraction kits. On-chip lysis of e.g. breast cancer cells is achieved within only 30 - 45 seconds. MiRNAs are extracted from the crude lysate within 8 minutes (duration of commercial kit extraction: ~1h). Extracts are checked for purity by the Bioanalyzer 2100 capillary electrophoresis system. The excellent amplifiability of the extracted miRNAs is demonstrated by a RT-qPCR assay lacking preamplification steps. All on-chip processes are performed without the use of any toxic components.