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C. elegans: Forward Genetic Screen

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by Cullen Lightsey on 19 April 2013

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Transcript of C. elegans: Forward Genetic Screen

Cullen Lightsey, Stephanie Holt,
Mireya Duran, Huy Nguyen C. elegans
Forward Genetic Screen Conclusion Citations Results Forward genetic screens can help to understand mutagens and the mutations they produce
Studying C. elegans genes related to movement can carry over to human studies
Parkinson's Disease
Wilson's Disease
Ultimately, understanding the genetic life cycles of C. elegans will build our understanding of more complex life cycles and lead to better health care. LeDoux, Mark. Animal Models of Movement Disorders. Burlington: Elsevier Academic Press, 2005. Print.
Brenner, S. The genetics of Caenorhabditis elegans. Genetics 77, 71–94 (1974)
The C. elegans Sequencing Consortium. Genome sequence of the nematode C. elegans: a platform for investigating biology. Science 282, 2012–2018 (1998).
Kaletta, Titus, and Michael O. Hengartner. "Finding Function in Novel Targets: C. Elegans as a Model Organism." Nature Reviews Drug Discovery 5.5 (2006): 387-99. Print. Why C. elegans? Methods P0 generation mutanized with ethylmethane sulfonate (EMS)
5 Adults picked and placed separately on growth agar plates
After 12 hours, P0's laid eggs and were transferred to new plates
After another 12 hours, P0's were killed, isolating the F1 generation
F1's allowed to grow for 1 day then 5-7 from each plate transferred to new plate
After F1's laid eggs, all were removed and killed, isolating the F2 generation
F2 generation grew and was screened for mutation Our F2 and F3 generations both showed uncoordinated movement indicating a unc-like mutation
All worms in the F3 generations showed signs of unc mutation, suggesting that it is a recessive, inherited mutation. More detailed studies are needed to verify this hypothesis. Errors During our experiment, one P0 adult failed to produce any offspring when transferred to the second plate. We continued with our other nine P0 adult's offspring.
We also grew mold on one F1 plate. Results They are easy to culture

They have a fast life cycle

They reproduce rapidly

They are small and allow large scale in vivo study

Hermaphroditic

They share many genes and disease pathway with human.
unc-119 of C. elegans and HRG4/HsUNC-119 of human
DmUNC-119 genes of Drosophila Background Information - Sydney Brenner was a biologist from South Africa

- Studied C. Elegans using a forward genetic screen
-Study mutant phenotypes to understand key biological
processes
- Discover the molecular properties of the phenotype to find
what genes regulate these processes

- Identified over 600 mutants and their associated genetic pathways

- Won the Nobel Prize in 2002 for his work Citations Continue Maduro, M., and D. Pilgrim, SA. "Genes That Act Downstream of DAF-16 to Influence the Lifespan of Caenorhabditis Elegans." PubMed 141(3) (1995): n. pag. Print.
Maduro, Morris, Michael Gordon, Roger Jacobs, and David Pilgrim. "The Unc-119 Family of Neural Proteins Is Functionally Conserved Between Humans, Drosophila and C. Elegans." Journal of Neurogenetics 13.4 (2000): 191-212. Print
"Genes and Mapped Phenotypes." National Center for Biotechnology Information. U.S. National Library of Medicine, n.d. Web. 04 Apr. 2013
Anita G Fernandez, Bastiaan O R Bargmann,Emily K Mis, Mark L Edgley, Kenneth D Birnbaum, Fabio Piano. " High-throughput fluorescence-based isolation of live C. elegans larvae." Nature 141(3) (2012): pag1502–1510. Print Question Time! Materials - 20 Large NGM plates seeded with OP50 E.Coli
- 20 Small NGM plates seeded with OP50 E.Coli
- A work pick made from a glass pipette and
32-gauge platinum wire
- A dissecting microscope
- A Bunsen burner
- Parafilm
- N2 wild type C. elegans treated with
EMS mutagen C. Elegans Life Cycle Common Mutations of C. Elegans Roller (Rol): Rotates along long axis, circular movement patterns
Dumpy (Dpy): Shorter and stouter worms than N2
Uncoordinated (Unc): Deviations in self propelled movement from N2
Protruding Vulva (Pvl): Protrusion at vulva due to incomplete morphogenesis
Lineage Variant (Lin): Particular cells exhibit variations in development
Egg Laying Defect (Egl): Eggs laid at a slower rate, at an inappropriate time, no eggs, etc.
Blister (Bli): Fluid filled blisters along the cuticle
Long (Lon): Worms are longer and thinner
Sterile (Glp): Unable to reproduce, defective gametes
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