Lecture 28: Genetics and the nervous system 3 – molecular neuroscience Flashcards
Will neurones regrow after damage?
no
what’s a negative about spinal cord regeneration therapy?
Huge financial burden
Significant loss to quality of life
what are the 5 genetic engineering methods?
CRISPR/Cas9, Viral vectors, DNA microinjection, Electroporation, Nanoparticle / lipid transfection
What does PCR stand for?
Polymerase Chain Reaction
when was PCR developed?
1985
what is PCR used for?
used to create copies of a specific gene sequence
molecular photocopying
what do you need for PCR?
DNA polymerase DNA / PCR primers dNTPs dNTPs Buffers Thermocycler / Thermal cycler
what are the three steps of PCR?
denaturation
annealing
elongation
what happens in step 1 of PCR?
splitting DNA into two strands, no more hydrogen bonds holding them together
How lögn are PCR primers?
Usually 18-24bp in length
primers work in ___
pairs
what does the forward primer align with?
Anti sense strand
How does reverse primer move?
3’ to 5’
what happens in step 2, annealing for PCR?
temp reduced
hydrogen bonds formed between primers/dNTPs and template DNA
what happens in elongation?
DNA polymerase joins the backbone of the nucleotides together (3’5’)
what temp is needed for elongation?
70 degrees
How many cycles of PCR are done?
20-40
what happens during the 1st cycle of PCR?
DNA polymerase keeps going and going until there is no more template strand
what happens during the end cycle of PCR?
this is where your DNA containing only your target sequence is formed
what happens during every cycle?
the number of DNA molecules doubles
what happens after the 3rd cycle of PCR?
double the number of DNA molecules that only contain the sequence between your primers
what do you need to apply to see a PCR product?
DNA-binding dye
- need UV light
How do you separate PCR product from the template DNA?
Gel electrophoresis
Gel electrophoresis separates DNA fragments based on?
size
in gel electrophoresis where is the sample loaded?
into an agarose gel matrix
in gel electrophoresis what cause stem DNA to migrate?
An electric field
DNA moves towards what and why?
DNA is negatively charged so will move towards the positive electrode
what shape is formed from gel eletrophoresis?
ladder
what does the product of electrophoresis allow us to do?
measure how big your PCR fragment is
by using distance on ladder
what is PCR?
molecular technique used to replicate DNA fragments of interest
what is DNA cloning?
Process of making multiple, identical copies of a DNA fragment
what is DNA cloning used for?
Separate out copies of genes with SNPs
Add promotor sequences
Make huge numbers of copies of a gene
Express proteins
