Henry Flashcards
(36 cards)
The Weissman model
Building blocks for cell types joined together in fertilised egg. The asymmetrical segregation of these to daughter cells resulted in cell types.
Intrinsic and extrinsic cues
Intrinsic from within the cell
extrinsic from outside environment
Types of extrinsic cues
Paracrine - Cell to cell
Autocrine - cell to self
Juxtacrine - only neighbouring cells
Signal transduction from cues (general pathway)
Receptor activates
Cascade of secondary messengers
Transcription factor activation
How can different TFs regulate one gene (shh)
regulation via different upstream enhancer sequences that different TFs can recognise
How do cells become distinct (in general)
They acquire different gene profiles to one another through TF activation or inactivation
What is more common during development, cell signalling to acquire fate or cytoplasmic inheritance
Cell signalling
cytoplasmic inheritance is more common at early stages
3 layers formed during gastrulation and tissues they form
Ectoderm - skin, CNS, Glia
Mesoderm - Skeleton and muscle
Endoderm - digestive tract and lungs
General progression of vertebrate development
Zygote —gastrulation—>Blastula –>pharyngula stage
details of pharyngula stage
highly conserved throughout vertebrates
formation of pharyngeal pouches, somite, notochord, neural tube, tail
What are the 4 general processes that occur during development
Pattern formation - establishment of body axes to allow positioning
Morphogenesis - establishment of shape
Differentiation - Gaining of specialisation whilst losing pluripotency
Growth - by cell proliferation, enlargement or accretion of ECM
Methods used in morphogenesis
Cell:
migration
death
adhesion
change of shape
methods of investigating gene expression
in situ hybridisation
reporter line gene fusions
RNAseq
In-situ hybridisation
DIG labelled RNA, complimentary to mRNA of gene of interest, injected into cell.
Anti-DIG maker with alkphos EZ added
Alkphos creates blue substrate
advantages of in-situ hybridisation
Anti-DIG marker commercially available so only RNA needs to be synthesised making this process cheap
Reporter line gene fusions
either replace a gene or use the same promoter sequence for GFP gene.
Shows when and where gene is expressed
GFP
protein from jellyfish that absorbs light of 475nm and radiates out at 510nm which is visible and blue. allows for high contrast imagery
RNAseq
provides quantitative information of all genes transcribed
- RNA from cell cytoplasmic extract is read
- No. of different length RNAs counted, average for each is found and up or downregulation can be deduced
- Volcano plot created
single cell RNAseq
allows specific transcriptome for cells in complex tissue to be read
allows detection of causes for complex diseases like autism in model organisms
methods of investigating Protein expression
Immunohistochemistry
Fusion protein constructs
Immunohistochemistry
Use of antibodies specific to epitopes on proteins
secondary antibody with alkphos or dye conjugated allows for visualisation of expression
Fusion protien constructs
Fluorescent gene added to 5’ end of gene of interest
allows for visualisation of expression and transport of gene product
3 Loss of function mutations
amorphic - null mutation = complete loss
hypomorphic - reduced expression
anti-morphic - competitive inhibition of gene
Gain of function mutation
hypermorphic- increased activity
