THE INFRASTRUCTURE FOR RESEARCH ON NON MAMMALIAN MODEL ORGANISMS

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23 Οκτ 2013 (πριν από 3 χρόνια και 9 μήνες)

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THE INFRASTRUCTURE FOR
RESEARCH ON NON MAMMALIAN
MODEL ORGANISMS

Danio

rerio

Drosophila melanogaster

Univeristy

of Tampere


Mika
Rämet


Howard Jacobs


Dan
Hultmark




Model organisms:


Zebrafish


Drosophila



Budget (2010
-
2012)


340.000


(zebrafish)


120.000


(Drosophila)

University of Helsinki



Osamu
Shimmi


Ville
Hietakangas


(Institute of Biotechnology)


Tapio

Heino


(Department of Biosciences)



Model organism:


Drosophila



Budget (2010
-
2012)


15.000


Members

Provided services from fly groups


a.
Maintenance of the fly lines

b.
Hands
-
on guidance to work on mutant and RNAi fly
lines

c.
Annual training course

The Drosophila units in
Viikki

Biocenter


Fly maintenance:

Separate room for fly work




Media kitchen for making the fly food




Fly incubators (25oC, 18oC, 29oC)




18oC room for fly stock maintenance

Research apparatus:

Work station for fly
anaesthesia

(CO2)

Fluorescence stereomicroscope

Equipments to generate transgenic flies


Research fields:

Developmental biology, signaling




(
Shimmi
), growth and metabolism




(
Hietakangas
), Neurobiology (
Heino
)

Why is Drosophila research useful?

1.
Genome wide approaches, e.g. microarray analysis, high
throughput screening, are commonly used in biomedical
research
---

a large number of ‘candidate genes’

2.
Challenging issue is to understand in vivo function of
‘candidate genes’

3.
Knock
-
out approach
---

time consuming for evaluating a
large number of ‘candidate genes’

4.
Knock
-
down approaches (RNAi,
morpholino
) are useful
--
-

tissue specific, stage specific system of gene regulation
desired

5.
Many
genes required for
growth and development are
highly conserved between mammals and Drosophila

i
n vivo RNAi approach in Drosophila

*Two stock centers provide Drosophila RNAi lines covering
over 95% of Drosophila genome

Gal4 drivers:

tissue specific,
e.g

neuronal cells, wing
imaginal disc,

Inducible (HS driver),

Over 500 drivers are
available

UAS
-
RNAi;
available at
the stock
centers

Results available in 10 days

Signaling cascades in Drosophila wing
development

Hh

DPP/BMP

EGF

Notch

DPP/BMP

L4

L5

L2

L3

PCV

*Loss of these gene functions are embryonic lethal

MS1096 (larval wing disc) >tkv

(BMP type I receptor)

shv (pupal wing) >tkv

(BMP type I receptor)

MS1096 >net

(downstream of Notch)

En (posterior wing disc) >Rolled

(MAPK, downstream of EGFR)

MS1096 >
smo

(
Hh

signaling)

i
n vivo RNAi in Drosophila wing

Application of in vivo Drosophila RNAi

RNAi clone of BMP type I receptor in wing imaginal disc

Anti
-
phosphoMad

(Readout of signal)

RNAi clone

Merged image

*Experiments (fly crosses, dissection and
Ab

staining) can be
done for 6
-
7 days

in vivo
RNAi screening

MS1096 Gal4 driver > RNAi lines

---

tested ~2000 lines so far

One of the hits,
lolal

(transcriptional factor
containing a BTB/POZ domain) is a postulated
component that genetically interacts with dpp
by a previous genetic screening (Manuscript in
preparation
)

MS1096 >
lolal

Drosophila in vivo RNAi is a suitable approach for
primary characterization of ‘candidate genes’ in the
post
-
genome era


1)
We plan to organize the training course for non
-
Drosophila
researcher; in vivo RNAi

2)
We plan to set up a web site providing a practical
information for in vivo RNAi