Download Final Project Outline Aaron Loewen 33334137 Name: Aaron

Final Project Outline
Aaron Loewen
33334137
Name:
Aaron Loewen
Topic:
Knockout of Eurl on chromosome 16 in Mus musculus (house mouse) to model Homo
sapiens sapiens (human) Eurl in chromosome 21.
Specific Question:
Is Eurl necessary for normal brain development in regard to brain size in Mus musculus?
How this is novel and original:
Eurl is a relatively new gene that has been only recently found to affect the brain in
humans. The human chromosome 21 is homologous to the mouse chromosome 16,
which allows us to use the mouse as a model organism for human brain development
when it comes to genes associated with chromosome 21. Knockout of the gene (as well
as the addition of another copy) has not been done in literature (according to PubMed)
so seeing the outcomes of the experiment will show us novel information on Eurl in
mice and therefore in humans by inference.
Impact:
With this proposed experiment we will be able to look into this new gene that has been
only recently described briefly in both humans and mice. Because the mouse
chromosome is homologous with the human one, we can use the model to infer
changes in humans based on the changes that occur in mice. This gene in humans may
be an important factor in brain development in the fetus if there is a change in human
brain size. As well, there would also be implications in the mechanism that Downs
Syndrome affects humans, as microcephaly is a symptom of said condition.
Hypothesis:
HA1 :
Knockout of Eurl will cause a change in expression levels of Eurl in Mus
musculus that increases the size of the brain of Mus musculus.
HA2 :
The addition of an extra Eurl will cause a change in expression levels of Eurl in
Mus musculus that decreases the size of the brain of Mus musculus.
Final Project Outline
HO :
Aaron Loewen
33334137
A change of Eurl will not cause a change in expression levels of Eurl in Mus
musculus, therefore not changing the size of the brain of Mus musculus.
Evidence:
There has been some correlation between Eurl expression and progenitor proliferation
and differentiation in neurons (Li et al. 2016). This relationship allows us to infer that it
may also affect brain development, particularly in brain size. The more Eurl there is, the
smaller the brain size should occur. Therefore adding another copy of it should reduce
brain size in theory.
The introduction of Downs Syndrome into this experiment stems from the fact the the
Eurl gene in humans is found in the region where Down’s Syndrome is likely to be
caused by (Li et al. 2016). Microcephaly has been found in Downs Syndrome patients,
and therefore would allow us to connect a decreased brain size to increased Eurl levels
(Tyler and Haydar 2013 and NCBI1 2016).
Removing the Eurl may also increase brain size in the same way.
Chromosome 16 in mice is orthologous/homologous to chromosome 21 in humans
(Gitton et al. 2002 and NCBI1 2016) which makes it a good model for brain development.
Brain size experiments have been done before with mice, which show that it can be a
strong model for human development (Tyler and Haydar 2013).
Prediction:
When Eurl is knocked out, we should see that expression of Eurl will decrease. When
Eurl is added, we should see an increase in expression. When knocked out, we should
see that head size will increase or remain the same. When an extra copy is added, we
should see that the head size will decrease (microcephaly).
Proposed Method *:
1. Obtain mouse embryonic stem cells.
2. Knockout Eurl in half of the embryos, creating a group that has the inactive Eurl and
a group that has the wildtype Eurl
- Knockout method is currently unknown
3. Add another copy of Eurl to the stem cells, replicating a Down’s group.
4. Allow mice to age to 30 days
Final Project Outline
Aaron Loewen
33334137
5. Measure Eurl expression using a gene reporter construct for the extra copy
variation
- Use a LacZ reporter for this to quantify product
- Do this at day 1, 6, 12, 18, 24, and 30.
o
6. Measure mice brains at day 30
Annotated Bibliography *:
Gitton, Y., Dahmane, N., Baik, S., Altaba, A.R., Neidhardt, L., Scholze, M., Herrmann, B.G., … & Yaspo, M.
(2002). A gene expression map of human chromosome 21 orthologues in the mouse. Nature,
420, 586-590.
Chromosome 21 in humans is orthologous with chromosome 16 in mice.
Li, S.S., Qu, Z., Haas, M., Ngo, L., Heo, Y.J., Kang, H.J., Britto, J.M., … & Heng, J.I. (2016). The HSA21 gene
EURL/C21ORF91 controls neurogenesis within the cerebral cortex and is implicated in the
pathogenesis of Down Syndrome. Scientific Reports, 6, 29514.
This article is what lead me into the Eurl gene, allowing me to see how it works with
neural cells and spine, as well as its wild-type expression levels.
https://www.ncbi.nlm.nih.gov/homologene/9696 NCBI 1
This link from NCBI shows homology within the Eurl genes from different species (Will
create citation for this site when I figure out how…).
Tyler, W.A. & Haydar, T.F. (2013). Multiplex Genetic Fate Mapping Reveals a Novel Route of Neocortical
Neurogenesis, Which Is Altered in the Ts65Dn Mouse Model of Down Syndrome.The Journal of
Neuroscience, 33(12), 5109-5119.
Gives us genetic information about Downs Syndrome.
http://www.nobelprize.org/nobel_prizes/medicine/laureates/2007/advanced.html
GENE KNOCKOUT IN MICE – THEORY AND METHOD
Final Project Outline
Aaron Loewen
33334137
Family / friends report info:
I will try to talk up its relationship with Downs Syndrome, as it is a fairly common issue
and any new info about it can help. No direct cure from this, but in terms of
mechanisms we can say that this could help lessen the effects of it. May also help with
microcephaly if other conditions affect this area (unknown at the moment but further
studies can look into it).
Finished info: I have finished what is here, as well as gathered some other info about Downs, Zika
(assists with proving microcephaly in mice is a good model for human brain
development) as well as other small bits of info that can be added later on.
Bioinformatics can also be used to create primers, so some knowledge of that will be
tagged onto a much more in-depth discussion of the methods used.
Feedback:
I was hoping to get feedback on whether I used sufficient controls in the experiment? As
well, any feedback on the method would be much appreciated.
* denotes unfinished sections