Lecture 28: Genetics and the nervous system 3 – molecular neuroscience

Question 1

Will neurones regrow after damage?

Answer 1

no

Question 2

what’s a negative about spinal cord regeneration therapy?

Answer 2

Huge financial burden

Significant loss to quality of life

Question 3

what are the 5 genetic engineering methods?

Answer 3

CRISPR/Cas9, Viral vectors, DNA microinjection, Electroporation, Nanoparticle / lipid transfection

Question 4

What does PCR stand for?

Answer 4

Polymerase Chain Reaction

Question 5

when was PCR developed?

Answer 5

1985

Question 6

what is PCR used for?

Answer 6

used to create copies of a specific gene sequence

molecular photocopying

Question 7

what do you need for PCR?

Answer 7

DNA polymerase
DNA / PCR primers
dNTPs
dNTPs
Buffers
Thermocycler / Thermal cycler

Question 8

what are the three steps of PCR?

Answer 8

denaturation
annealing
elongation

Question 9

what happens in step 1 of PCR?

Answer 9

splitting DNA into two strands, no more hydrogen bonds holding them together

Question 10

How lögn are PCR primers?

Answer 10

Usually 18-24bp in length

Question 11

primers work in ___

Answer 11

pairs

Question 12

what does the forward primer align with?

Answer 12

Anti sense strand

Question 13

How does reverse primer move?

Answer 13

3’ to 5’

Question 14

what happens in step 2, annealing for PCR?

Answer 14

temp reduced

hydrogen bonds formed between primers/dNTPs and template DNA

Question 15

what happens in elongation?

Answer 15

DNA polymerase joins the backbone of the nucleotides together (3’5’)

Question 16

what temp is needed for elongation?

Answer 16

70 degrees

Question 17

How many cycles of PCR are done?

Answer 17

20-40

Question 18

what happens during the 1st cycle of PCR?

Answer 18

DNA polymerase keeps going and going until there is no more template strand

Question 19

what happens during the end cycle of PCR?

Answer 19

this is where your DNA containing only your target sequence is formed

Question 20

what happens during every cycle?

Answer 20

the number of DNA molecules doubles

Question 21

what happens after the 3rd cycle of PCR?

Answer 21

double the number of DNA molecules that only contain the sequence between your primers

Question 22

what do you need to apply to see a PCR product?

Answer 22

DNA-binding dye

- need UV light

Question 23

How do you separate PCR product from the template DNA?

Answer 23

Gel electrophoresis

Question 24

Gel electrophoresis separates DNA fragments based on?

Answer 24

size

Question 25

in gel electrophoresis where is the sample loaded?

Answer 25

into an agarose gel matrix

Question 26

in gel electrophoresis what cause stem DNA to migrate?

Answer 26

An electric field

Question 27

DNA moves towards what and why?

Answer 27

DNA is negatively charged so will move towards the positive electrode

Question 28

what shape is formed from gel eletrophoresis?

Answer 28

ladder

Question 29

what does the product of electrophoresis allow us to do?

Answer 29

measure how big your PCR fragment is

by using distance on ladder

Question 30

what is PCR?

Answer 30

molecular technique used to replicate DNA fragments of interest

Question 31

what is DNA cloning?

Answer 31

Process of making multiple, identical copies of a DNA fragment

Question 32

what is DNA cloning used for?

Answer 32

Separate out copies of genes with SNPs
Add promotor sequences
Make huge numbers of copies of a gene
Express proteins