MicroRNA Knockdowns with Morpholinos

Gene Tools announces a new paper describing techniques for miRNA activity knockdown, maturation knockdown and specificity controls using Morpholino oligos.

MicroRNAs regulate the expression of genes, often large groups of genes. They are important in embryonic development, in cardiac function, in carcinogenesis and in many other processes. The recent discovery of miRNAs has altered our understanding of gene regulation, with many new reports of miRNA activity appearing in the literature. Experimental manipulation of miRNA activity will become a standard tool of molecular biologists, but currently these techniques are in development and still improving.

MicroRNAs mature through several steps. A primary miRNA is transcribed from DNA and folds into a stem-loop. The stem-loop is cleaved from the rest of the transcript by the nucleolytic enzyme Drosha, then the loop is cleaved from the stem by the nucleolytic enzyme Dicer. The double stranded stem interacts with the protein Argonaute, which cleaves and releases one of the strands, forming the miRISC complex with the other strand, now called the guide strand. It is the miRISC complex which interacts with mRNAs, altering their expression.

Knockdowns of miRNA activity have generally used oligos targeting the miRNA guide strand. A Morpholino oligo targeting an miRNA guide strand can interfere with the activity of the miRNA. It is difficult to control for the specificity of the knockdown when using this technique alone. However, Morpholinos targeting the nucleolytic processing sites of an immature miRNA can prevent maturation of the miRNA. This allows sets of nonoverlapping Morpholino oligos targeting a primary miRNA to be used as specificity controls; if two non-overlapping oligos targeting the same miRNA produce the same phenotype, this supports the hypothesis that the phenotype is due to knocking down the activity of the targeted miRNA and not due to an off-target effect. These techniques are explored in the following paper:

Kloosterman WP, Lagendijk AK, Ketting RF, Moulton JD, Plasterk RHA. Targeted inhibition of miRNA maturation with morpholinos reveals a role for miR-375 in pancreatic islet development. PLoS Biol. 2007;5(8): e203.

Available online: http://biology.plosjournals.org/perlserv/?request=get-pdf&file=10.1371_journal.pbio.0050203-L.pdf

Morpholino oligos are available from Gene Tools LLC: www.gene-tools.com

GENE TOOLS, LLC is a biotech company that makes custom-sequence Morpholino oligos for biological and medical research. GENE TOOLS, founded in 1997, is a spinoff of AVI Biopharma Inc. focused on research and diagnostic applications of Morpholino technology. Morpholinos from Gene Tools have been commercially available since 2000 and there are over 1700 publications in the research literature using Morpholinos (pubs.gene-tools.com).

Our primary products, Morpholino antisense oligos, are synthetic molecules. A specific Morpholino blocks the activity of a selected gene. The sequence in which the subunits of the Morpholino are linked determines which gene will be blocked. At GENE TOOLS we custom-synthesize each Morpholino in the appropriate sequence to shut down the activity of the customer’s gene of interest. We usually synthesize over 200 of these Morpholinos each week.

GENE TOOLS’ customers are scientists at universities, research institutes and pharmaceutical companies. Morpholino oligos are used by:
• developmental biologists studying embryo development by injecting Morpholinos into eggs or zygotes of zebrafish, frog, ascidians, sea urchins or other organisms;
• cell biologists determining the function of newly sequenced genes; and
• medical researchers studying cellular mechanisms underlying cancer and other diseases.

For more information on Morpholino oligos, see the Gene Tools website (www.gene-tools.com) and the Wikipedia Morpholino article (http://en.wikipedia.org/wiki/Morpholino).

Gene Tools, LLC
One Summerton Way
Philomath, OR 97370 USA

Press contact: Jon D. Moulton, Ph.D. (541) 929-7840 x1201

This release was published on openPR.