The MicrobrewRNA

The MicrobrewRNA is a unique analysis service for relative quantitation of endogenous microRNA expression.

Introduction:

The recent discovery of microRNAs and subsequent identification of their pivotal role in gene silencing has led to the recognition of a new level of eukaryotic gene regulation ¹ . Abberant microRNA expression (which has been observed in a number of human diseases) can lead to translational repression or degradation of target mRNAs.

To assist researchers the MicrobrewRNA service offers to;

• Isolate RNA from cells or tissue and provide relative quantitation* of microRNA and mRNA expression levels.

• Verify and quantitate microRNA expression data obtained from Northern Blots or microRNA arrays.

• Quantitate microRNA levels resulting from transfected expression plasmids.

• Identify those mRNAs from computationally predicted microRNA targets subject to degradation.

Background:

The service uses a method, developed by researchers at the Ordway Research Institute and Wadsworth Centre ² .

This anchored oligo(dT) based RT-PCR approach reverse transcribes all microRNAs and mRNAs in a sample. Subsequently, the levels of miRNAs and mRNAs can be measured in cell lines and tissues, allowing relative quantitation. An inverse relationship to the expression levels of the miRNA c onfirms those predicted mRNA targets subject to degradation .

For further details of the MicrobrewRNA service, please contact gbrock@ordwayresearch.org or ghurteau@ordwayresearch.org

* Relative quantitation determines the changes in steady-state microRNA levels in samples and expresses it relative to the levels of an internal control RNA or miRNA.

References:

1. Ambros V. microRNAs: tiny regulators with great potential. Cell 2001; 107:823-6.

2. Hurteau GJ, Spivack SD, Brock GJ. Potential mRNA Degradation Targets of hsa-miR-200c, Identified Using Informatics and qRT-PCR. Cell Cycle 2006; 5.

The method is outlined below (click individual images to enlarge)

The MicrobrewRNA currently has preliminary data on several miRNAs and associated mRNA targets.

For further information contact:

gbrock@ordwayresearch.org

ghurteau@ordwayresearch.org

Additional information.

The associated report details analysis of a variably expressed microRNA (miR-200c) identified in cell lines. A combination of Affymetrix expression data and computational prediction programs was used to select twenty potential mRNAs. Expression analysis revealed four mRNAs with an inverse relationship to miR-200c. Finally, inhibition of miR-200c using an anti-miRNA 2’-0-Methyl oligonucleotide (AMO) resulted in an increase in expression of one of the targets, the transcription factor TCF8 2 .


Background reading.  

The URT method is covered by , USPTO Pat. Pend. #10/342,684.
Lim LP, Lau NC, Garrett-Engele P, Grimson A, Schelter JM, Castle J, Bartel DP, Linsley PS, Johnson JM. Microarray analysis shows that some microRNAs downregulate large numbers of target mRNAs. Nature 2005; 433:769-73.
Bagga S, Bracht J, Hunter S, Massirer K, Holtz J, Eachus R, Pasquinelli AE. Regulation by let-7 and lin-4 miRNAs results in target mRNA degradation. Cell 2005; 122:553-63.
Ambros V. microRNAs: tiny regulators with great potential. Cell 2001; 107:823-6.
Lagos-Quintana M, Rauhut R, Meyer J, Borkhardt A, Tuschl T. New microRNAs from mouse and human. RNA 2003; 9:175-9.