5/ developmental genetics Flashcards
1
Q
vertebrate gene nomenclature: homo sapiens, mus musculus, gallus gallus, xenopus laevis, danio rerio. gene symbol/protein
A
- hs: SHH/SHH
- mm: Shh/SHH
- gg: SHH/SHH
- xl: shh/Shh
- dr: shh/Shh
2
Q
how do we make random mutations?
A
- radiation - x rays, breaks DNA in half or modifies sequence
- chemical - base modifiers, very toxic hence mutations, random through genome
- can be point mutations (single base pair), deletion, insertion, translocation
3
Q
method of forward genetics (phenotype –> gene)
A
- cause high levels of background mutations in model organism - mutagenesis
- look for interesting phenotypes
4
Q
method of reverse genetics (gene –> phenotype)
A
- start with gene of interest and see what a mutation in it looks like
- gene knock out: completely remove gene to determine its function
- knock in/gene replacement: add gene of interest to a different location, makes small change to endogenous gene
5
Q
brief notes on CRISPR
A
- can be used for know out and knock in
- can be used in any organism
6
Q
3 ways mutations affect genes
A
- changes in regulatory sequences in the DNA that affects transcription (eg enhancers, TF can’t bind as well, less protein)
- changes in non-coding sequence of the transcript - could affect RNA splicing, stability or translation
- changes in coding sequence: affect folding of protein or truncate it. missense single amino acid substituted, nonsense stop codon
7
Q
how do normal TFs work?
A
- bind to DNA, dimerization, conformational change, transcription activation
- TFs have a DNA binding domain and dimerization domain - need 2 versions of a TF to activate
8
Q
amorphic TF mutation
A
- eg missense mutation that inactivates DNA binding domain
- +/- normally enough gene product from 1 wild type copy, haplosufficient. dimerization can still occur
- -/- no transcriptional activation
- recessive
9
Q
note on number of lethal genes
A
most people carry between 10-20 lethal genes but haplosufficiency means one gene is enough
10
Q
antimorphic (dominant negative) TF mutation
A
- eg missense that destroys dimerization domain
- +/- mutant binds DNA but doesn’t dimerize w WT, no conformational change, no transcription (just enough function to mess it up). only transcription when 2 WT together
- -/- completely inactive
- DOMINANT
11
Q
hypomorphic (weakened) TF mutation
A
- eg weakened DNA binding domain
- +/- normally fine. mutant may also be able to dimerize w wild type
- -/- mild phenotype, less severe than amorphic. dimer forms but DNA often falls off
- recessive
12
Q
hypermorphic (overactive) TF mutation
A
- missense resulting in activation w/o dimerisation
- +/- mutant form binds DNA and is always active - constitutively active
- -/- same
- DOMINANT
13
Q
whats the word for types of phenotypes produced by mutations
A
Muller’s Morphs
14
Q
genetic pathway
A
- series of things that need to happen to produce a phenotype
- eg enzyme A activates enzyme B which activates C which activates enzyme D which produces melanin in fish
- an amorphic mutation in any of the enzymes would cause an albino fish
- different mutations in different genes that result in the same phenotype suggest the genes function in the same pathway
15
Q
Green fluorescent protein
A
- able to identify the single gene that jellyfish used to fluoresce and inject it into other animals (now many others)
- laser powered blue light absorbed by GFP, which becomes excited and emits light at a different wavelength
- looks like its glowing in the dark
16
Q
generating GFP transgenic line
A
- genetically engineer GFP onto end of last exon by gene fusion/replace the gene by reporter construct
- put back into animal
17
Q
uses for GFP transgenic lines
A
- to follow expression of gene or follow behaviour of cells in vivo
- follow subcellular localisation of a protein
