A Novel Approach to mRNA Isolation using PNAs

DESCRIPTION (Applicant's abstract): Peptide nucleic acids (PNAs) have a number of features that suggest they would be ideal probes for purifying mRNA. In this revised format we have demonstrated the use of phosphono PNAs (pPNAs) in an RNA isolation procedure that improves the recovery of RNAs with secondary structure at the poly A region and RNAs with short poly A tails. We have developed a procedure for isolating mRNA (free of genomic DNA) from cells and tissue using pPNAs, and provided this kit to the research community. The goal of this revised Phase I study is to further develop the pPNA technology as to enable isolation of mRNA in a high-throughput format. Also by incorporating a novel PNA-like monomer on the base of trans-4-hyroxy-L-proline (HypNA) into the pPNA backbone we hope to be able to purify mRNA from plant Total RNA. This development of PNA technology would enable the HypNA-pPNA chimera to hybridize with plant mRNA molecules regardless of the presence of plant polyphenolics. In Phase II of this study, we would hope to further develop this technology such that it can be used in a novel approach to mRNA isolation from bacterial cells as well as to develop HypNA-pPNAs chimeras for array technology. PROPOSED COMMERCIAL APPLICATION: Any group that is trying to obtain a complete set of transcripts may find an application for pPNA technology. This may include those working on the Mammalian Gene Collection (MGC). We already supply the first two versions of the kit, Maxi and Midi mVADER, to the research community. Any future products generated from this proposal will also be provided to the research community. With the 96-well format this kit will allow high throughput analysis of gene expression for drug screening and functional genomics-applications. This kit would also facilitate those interested in generating mRNA for array profiles or for those interested in studying quantitative RT-PCR. We believe that the invading properties of pPNAs will enable capture of plant mRNA molecules even in the presence of lipopolysaccharides and polyphenolics. The successful application of pPNA technology to plant biotechnology would certainly increase the market potential of this product.