E Presentation Morpholino Oligos

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Information about E Presentation Morpholino Oligos
Technology

Published on May 22, 2009

Author: sol777

Source: slideshare.net

Description

Morpholino Oligos are an interesting technique to silence Genes. Learn more about their usage and potential.

Morpholino Oligos: Making Sense of Antisense? g Based on the review of Janet Heasman Division of Developmental Biology, Children’s Hospital Medical Center, Cincinnati, Ohio Developmental Biology 243, 209–214 (2002) 08.12.2008 Genie Genetique – Sebastian König and Sebastian Olényi

Overview • Do nreg lating gene e pression helps Downregulating expression understanding their functions • Most of the experiments are done to understand embryonic development • Morpholinos are synthetic, usually 25 bases long, with a diff ith different sugar-structure t t t • Typical model organisms: zebrafish, African clawed frog, frog sea urchin, chick and mice. urchin mice Genie Genetique – Sebastian König and  08.12.2008 Sebastian Olényi

Structures Genie Genetique – Sebastian König and  08.12.2008 Sebastian Olényi

Principle “DGI reliably  achieves >90%  knockdown of  gene expression” Genie Genetique – Sebastian König and  08.12.2008 Sebastian Olényi

Advantages • E Equal or hi h efficacy and specificity l higher ffi d ifi it greatly superior to any other antisense structural type t t lt • Morpholinos do not activate the complement cascade or the interferon system and are not degraded • Excellent solubility and stability • Working at high concentrations is p g g possible • No need for RNase H to ensure their activity Genie Genetique – Sebastian König and  08.12.2008 Sebastian Olényi

Disadvantages • Completely artificial molecule, so no vectors possible p • Delivery usually only via microinjection or electroporation • Not always high effects, uncertain amount of downregulation • Side-effects for high dosage oligo- dosage, dependent Genie Genetique – Sebastian König and  08.12.2008 Sebastian Olényi

Application • 25 25mers, Little self-complementarity (l Littl lf l t it (less th 4 than contiguous intrastrand base pairs) • Less than 36% guanine content • No runs of more than triplets of GGG • Concentrations between 1 and 12µM • 2-4 days effectiveness • Controls with Co t o s t – mRNA rescue experiments – Two different morpholinos directed to different gene parts with same effect t ith ff t – Other over- and/or underexpression studies Genie Genetique – Sebastian König and  08.12.2008 Sebastian Olényi

Genie Genetique – Sebastian König and  08.12.2008 Sebastian Olényi

Thank you! References http://www.sanger.ac.uk/Info/Press/gfx/zebrafish.jpg http://www.columbia.edu/itc/cerc/danoff‐burg/invasion_bio/inv_spp_summ/xenopuslaevis.jpg http://www.alaskaunderseatours.com/photos/Red‐Sea‐Urchin.jpg http://www.gene‐tools.com/products_and_applications  http://www.bentham.org/ctmc/openaccessarticles/ctmc7‐7/0003R.pdf Genie Genetique – Sebastian König and  08.12.2008 Sebastian Olényi

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