Epigenetics 2 Flashcards
First step of DNA methylation?
Dnmt3a/b methylate the DNA
Second step of DNA methylation?
Dnmt1 maintains the methylation
How is DNA demethylated passively?
Stop providing Dnmt1
How is DNA demethylated actively?
TET enzymes
How can TET1 be used to make IPS cells?
It can replace Oct4, and promote Oct4 demethylation and reactivation
How does bisulfite sequencing work?
Boil DNA w/ bisulfite
Causes methylated based to turn into Uracil
When its PCR the uracil turns into a T
Issue with bisulfite sequencing?
Couldnt differentiate between methylation and hydroxymethylation
How can nanopore sequencing be use to detect methylation?
methylated bases give a diff current to non-methylated bases
How many histones are in a nucleosome?
8
Where can histones be modified?
On their AA tails
What does H4K91 mean?
Fourth histone, AA 91
How can histone modifications be analysed?
Grow cells in culture
Cross link cells–>sticks protein to DNA
Fragment DNA
Attach DNA (that has protein stuck to it) to an antibody
Has specific antibodies for all the histone modifications
Digest protein and purify the DNA
Sequence the DNA
How is DNA fragmented?
Sonication, or with an enzyme
What kind of mark is acetylation of histones?
Active mark
What happens to the nucleosomes if they are deacetylated?
They can get closer together
What do HAT proteins do?
Acetylate histones
What do HDACs do?
Deacetylate histones
What is alpha ketoglutarate needed for?
Demethylation
What is acetylCoA?
Acetylation
What is energy generated from NADH and NAD used for?
Transfer of different marks
What can polycomb proteins do?
Form repressive complexes
Examples of roles carried out by lncRNA?
Can attach to a protein and bring it to DNA (to which it has also bound)
X inactivation
Gene silencing
What do CTCF proteins do?
Stop polymerases from continuing to act on DNA after the target gene has been passed
How does CTCF work?
It has zinc fingers that can hold the genome like a paperclip–> creates active and inactive compartments (boundaries)
