L20?or 19? idk. L4 development - nuclear reprogramming

Question 1

totipotent

Answer 1

capabale of developing into entire organism
can differentiate into any cell type

Question 2

pluripotent

Answer 2

can differentiate into most

Question 3

what is the waddington landscape

Answer 3

potency decreases with time
the hill thing
kinda got disproved since now there’s ways of freezing cells at pluripotent stage or pushing them back from differentiated to pluripotent

Question 4

first experiment that showed cells are totipotent at 2 cell stage

Answer 4

hans driech
seperated sea urchin embryo at 2 cell stafe
formed 2 normal embryos (just slighlty smaller)
shows each had capacity to form full organism

Question 5

baby hair ligature experiments

Answer 5

tied embryo at 8 cell stage in middle with baby hair
from his sons head what
but not fully tied
one cell had nucleus the other didnt
the nucleated duplicated as usual until 16 cell stage
then one nucleus passed through the gap into the enucleated cell
and started duplicating
then both seperated fully
and formed 2 regular organisms

so shows that even at 16 cell stage, the cells individually hold enough info to form an organism
totipotent

Question 6

how did Briggs and Kings build on this experiement

Answer 6

able to clone frog embryos from blastula stage nuclei
see slides for full explanation

but found that totipotency stops at blastula stage
cuz couldnt get it to work after that

Question 7

how did john gurdon overcome the problem of totipotency getting stuck at blastula stage

Answer 7

tranplanted adult skin cells into enucleated cells
he realsied he had to ‘condition’ the cells into becoming totipotent again

so he did serial nuclear transfers
see slide

Question 8

how did he ensure he was acc nucelating the cells and it wasn’t becoming a new organism by accident (like the original nucelus not properly removed or somehintg)

Answer 8

albino nucleus put into pigmented enucleated eggs

got all albino frogs
that were all clones of each other

Question 9

dolly de sheep

Answer 9

first cloned mammal
similar mehtod to john gurdons method of cloning
and since her, many mammals been successfully cloned

Question 10

what cells can be pluripotent if isolated and grown in culture

Answer 10

epiblast cells
can give rise to somatic and germ cells

only cant do extraembryonic membranes

Question 11

how have ES cells been used to generate target knockouts in mice

Answer 11

need these cells to be cultured for a long time to generated the rare specific muations that you want

once u get it, must be extracted and can be grown to make chimeras

Question 12

what are induced pluripotency stem cells

Answer 12

cells from a differentiated stage forced back into pluripotency

iPS cells

Question 13

general method to make iPS cells

Answer 13

primary cell culture

put pluripotency genes (4 genes that induce pluripotency) in suitable vector

cells recieve the genes

culture and colonies will appear

can subculture the individual colonies
and freeze these iBS cells for further use

Question 14

2 big implications of iBS cells

Answer 14

• induce iBS cells from patients with disease
  then culture and study disease progression etc.
• induce iBS cells from patients for cell replacemtnt therapy
  (no immunological rejection cuz tranplanting with their own cells)

Question 15

what is the source of pluripotent embryonic stem cells

Answer 15

epiblast of pre-implantation embryos