LECTURE 5

Question 1

DNMT controlled reaction?

Answer 1

Methylation of cytosine

Question 2

What supplies the methyl group for the methylation of Cytosine

Answer 2

SAM-ch3

Question 3

Methylation causes

Answer 3

deactivation of genes and a decreased gene expression. this is due to a reduced Transcription factors binding.

Question 4

Methylated DNA is also?

Answer 4

more condensed =less accessible for Transcription factors

Question 5

Methylation heritability ?

Answer 5

yes heritable, when DNA synthesis occurs, methylation each strand (hemimethylated)DNMTI enzyme will come and methylate the other strand

Question 6

Requirement for methylation to occur

Answer 6

Short palindromic sequence

Question 7

function of NP65

Answer 7

ensures correct methylation on the unmethylated strand of the hemimethylated DNA

Question 8

Enzyme which controls new gene methylation?

Answer 8

DNMT3A and “3B

Question 9

TET1-3 enzymes enable

Answer 9

demethylaiton

Question 10

In meiosis methylation

characteristic

Answer 10

genome wide demethylation

paternal methylation rate is greater than the maternal

Question 11

Methylation during development

Answer 11

Determine cell fate

• As cells migrate from the neural layer they receive different methylation signals
• deffuenbaiation into different cell types

Question 12

theory to do with 5-Hydroxyl-methyl-cytosine?

Answer 12

Present in bacteria could it be to present in humans ?

Question 13

Method to test presence of of 5-hm-cytosine in mammals

Answer 13

1- Took cells and some from the brain (purkinje)
2- Extract the DNA
3- Into single nucleotides (digest)
4- Thin layer chromatography (to separate them)
5- A T G C and another spot

did the same with bacteria and received data in which the unknown peak and the peak in the bacteria known to be 5HM-C aligned

Question 14

What conclusion was lead to about why 5HMC is present in small volumes in the cell?

Answer 14

Intermediate In converting back into cytosine (5methylcytosine)

Question 15

Conclusion which lead scientists to believe that 5hmc has a function role as well as being an intermediate ?

Answer 15

Mutation in MeCP2 gene are known to cause rent syndrome

Question 16

MeCP2 function? Previously

Answer 16

Bind to 5-mC in order to promote chromatin condensing

Question 17

MeCP2 also known to ?

Answer 17

Bind to 5HMC to cause the chromosome to be less condense and activation of genes and their expression.

Question 18

Determining the locations of 5MC and 5HMC ?

Answer 18

Genetic map

Question 19

Why can’t we use standard sequencing approaches?

Answer 19

they are undetected. resemble cytosine in PCR Reactions

Question 20

Bisulphite sequencing?

Answer 20

Cytosine converted to uracil 
compare to original 
5mc not converted 
PCR 
- Sanger or illumina sequencing

Question 21

TAB-sequencing

Answer 21

Ability to distinguish between 5mc and 5hmc

• bisulfite sequencing
• Tet assisted: in which Glucose-Transferase adds a glucose group to HMC only
• TET enzyme does not work on glucose addition groups (5hmc) but does on mc to form carboxyl group.
• Carboxylated groups (5mc)are read as Thymines and glucose addition groups (hmc) are read as cytosine
• compare the bisulfites to the Tet assised, when there is a t instead of a C (in bisulfite only) presence of 5mc