zakaj smo tu?
TRANSCRIPT
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Zakaj smo tu?
OECD Biotechnology Statistics 2009 (http://www.oecd.org/sti/42833898.pdf)
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Proteomika
“Kot da geni in mRNA kaj pomenijo...”
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Nature (2006) 441:840
DNA microarray
Single-cell proteomics (flow cytometry)
PROTEOMIKA
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“I think the problem, to be quite honest with you, is that you’ve never actually known what the question is.”
Life?
Universe?
Everything?
Douglas N. Adams
1952-2001
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Eksperimenti brez a priori hipoteze
Fishing expeditions vs. Postavljanje pravih vprašanj
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http://www.ibgc.u-bordeaux2.fr/YPM
15kD
180kD
pI M
w
6,8 4,2
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Merjenje razlik v izražanju proteinov: ‘Difference Gel Electrophoresis (DiGE)’
2-D Electrophoresis. Principles and Methods; GE Healthcare
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Masna spektromerija v proteomiki
Nature (2003) 422:198
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Masna spektromerija v proteomiki
Nature (2003) 422:198
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Nature (2013) 494:266
Proteom kot kvantitivna lastnost (fenotip)
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Post-translacijske modifikacije proteinov
Molecular Cell (2008) 31:449
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A plastic epigenome. DNA and histones are targets of multiple modifications that convey
flexibility to the genome.
P Sassone-Corsi Science 2013;339:148-150
Published by AAAS
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Nature Reviews Mol Cell Biol (2013) 14:211
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Primer analize ‘fosfoproteoma’
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Molecular Systems Biology (2012) 8:623
Science (1997) 278:680
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Acetilacija proteinov / ‘acetilom’
Molecular Cell (2013) 49:1
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Interakcije protein-DNA (ChIP-chip, ChIP-seq)
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ENCODE - NGS in proteomika
Molecular Systems Biology (2013) 9:640
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Cell (2013) 152:642
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Določanje prisotnosti/količine : določanje funkcije
- imunoprecipitacija s pomočjo TAP označevalca/MS (proteinski kompleksi)
- dvohibridni sistem kvasovke (Y2H, protein-protein fizične interakcije)
- proteinske mikromreže
Nature (2003) 422:208
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Test s komplementacijo fragmentov proteina (Protein-fragment complementation assay; PCA)
Membranski kvasni dvohibridni sistem
FRET/BRET ( Fluorescence/bioluminescence resonance-energy transfer )
Curr Opinion Biotech (2011) 22:50
Kvasni dvohibridni sistem
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Nature (2003) 422:208
Strategije Y2H
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Nature Methods (2011) 8:478
Y2H & NGS
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Nature (2002) 415:141
Analiza proteinskih kompleksov z afinitetno kromatografijo / MS
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Primer: Piruvat kinaza – lokalizacija in acetilacija
Mol Cell (20013) 52:340
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J Proteom Res (2009) 8:20
High content microscopy
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Pex11 represents an ancestral module from which multidomain nuclear receptors arose
TiBS (2000) 25:227
Nature Rev Mol Cell Biol (2010) 11:644
Pex11 is required for peroxisome proliferation
Fis1
Pex11
- the most abundant protein in the px membrane
- deletion prevents and over-expression induces px proliferation
- recruits, but does not necessarily binds to, (mt) fission machinery proteins
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MYTH Nature Protocols (2010) 5:1281-1293
How is the signal (from Pex11) transmitted?
cy
N-
C-
Pex11 Ftp1 (Pex34)
N-
C-
Fis1
N-
C-