developmental genes screening - (Feb/05/2007 )
Guys I am new to genetics forum, I as an undergrad started to work in a genetics lab recently-my first assignment my boss gave was to come up with ideas to do a genetic screen to identify genes involved involved in development (most of them will be lethal and so how will you overcome this pblm)--in flies
pls help me out
Thanks in advance
One possible approach is Morpholino antisense. This is the standard tool for gene knockdown in zebrafish labs, and it works in flies; see the figure by Nipam Patel in:
Ekker SC, Larson JD. Morphant technology in model developmental systems. Genesis. 2001 Jul;30(3):89-93. Figure 3 panel a-d.
This is available online:
Morpholinos are steric-blocking antisense that can be used at low doses to partially inhibit gene expression, useful to observe effects of a gene for which a knockout would be embryonic lethal. For a description see Wikipedia: Morpholinos which is accessible from this link:
For more information about Morpholinos see the Gene Tools website, www.gene-tools.com.
Morpholinos have not caught on in flies, but some recent work has been published:
Pilot F, Philippe JM, Lemmers C, Lecuit T. Spatial control of actin organization at adherens junctions by a synaptotagmin-like protein Btsz. Nature. 2006 Aug 3;442(7102):580-4. Epub 2006 Jul 9.
For a database of publications related to Morpholino oligos, see pubs.gene-tools.com
Using a Morpholino to knockdown a gene is much easier and faster than constructing a knockout. A Morpholino does not modulate the expression of hundreds of off-target genes. This off-target expression modulation is a known disadvantage of siRNA:
Short interfering RNAs can induce unexpected and divergent changes in the levels of untargeted proteins in mammalian cells. Scacheri PC, Rozenblatt-Rosen O, Caplen NJ, Wolfsberg TG, Umayam L, Lee JC, Hughes CM, Shanmugam KS, Bhattacharjee A, Meyerson M, Collins FS. Proc Natl Acad Sci U S A. 2004 Feb 17;101(7):1892-7. Epub 2004 Feb 09.
For screening, reduced prices are available for sets of 100 oligos.
wow, what a cool technique, Thanks Jon that was lot helpful, I am reading on it and I will use it in my next lab meeting, I will somehow persuade my boss and try this.
Thanks again Jon