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analytical exam 2 > epigenetic > Flashcards

epigenetic Flashcards

1
Q

where does methylation take place one the DNA

A

at the cpg site (where a cytosine is followed by a guanine), it occurs on the cystestine on the lysine

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2
Q

where do the methyl groups for methylation come from?

A

deitry methyl donors

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3
Q

what is the cycle taht produced methyl

A

methionine metabolic pathway which is part of the one carbon pathway

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4
Q

how the methyl produced

A

methionine is converted to SAM. SAM donated its methyl group to make SAH which is then hydrolysed to homocytocine which is catabolised or remethylated to methionine

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5
Q

what is the enzyme involved in methylation

A

DNA methyltransferase

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6
Q

which DNMT are de novo and what does it mean

A

DNMT 3a and 3b and it means it can methylate unmethylated DNA

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7
Q

which DNMT are mainitance and what does this mean

A

DNMT1 and attached a methyl group to semi methylated DNA during replication

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8
Q

what is euchromatin

A

decondensed and transcriptionally active

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9
Q

what is heterochromatin

A

condensed and transcriptionally inactive

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10
Q

what does the nucleosome core consists of

A

3 copies of H2A,H2B, H3 and H4 making a histone Octamer, globular C terminals and N terminal tail

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11
Q

how many bp does a nucleosome consist of

A

146

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12
Q

what does it mean if nucleosome is acetylated

A

it is has a. ore open formation of DNA allowing for easier transcription

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13
Q

what does it mean if the nucleosome is deacteylated

A

less easy transcription

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14
Q

which enzyme deacetylates DNA

A

HDACs

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15
Q

which enzyme acetylates DNA

A

hats or KATs with actyl-coa as substrate

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16
Q

how many times can lysine be methylated

A

3

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17
Q

how many cytosines are methylated

A

3-4%

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18
Q

how does transcriptional repression happen

A

MeCP2 binds to methylated DNA and recreuits to HDCA1 for deatylation
also recuits DNMT which methylates the DNA

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19
Q

what enzyme add a methyl to lysine

A

KMT

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20
Q

how does DNA hypomethylation agent work?

A

it is a cytidine analogue. Methyl group cannot be added to the nitrogen on the 5 position. Also known to bind and insist DNMT

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21
Q

what is the example of tumour suppressor gene taht is switched off in cancer and what can it be switched back on by

A

P15INK4B inhibits cyclin dependant kinases and decitabine

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22
Q

what methylation is smoking related to

A

decreased methylation of coagulation factor II receptor like 3 gene- induces platelet activation

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23
Q

the methylation at what gene is thought to increase diabetes risk

A

5 gene loci

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24
Q

what is the relevance of epigenetic to modern medicine

A

-understanding disease medicine
-identifying biomarkers that might predict disease or inform us about prognosis
-using pigenetics to find new treatments

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25
Q

what is Mendels law

A

it states that in a diploid organisms there is an equal genetic contribution to offspring made by each parent

26
Q

what is genetic imprinting

A

mono-allelic expression

27
Q

what does maternally imprinted mean

A

maternally inactivated so paternally expressed

28
Q

what does paternally imprinted mean

A

paternally inactivated so maternally expressed

29
Q

why are genes imprinted

A

thought to be evolutionary as paternal genes tend to promote growth hand mothers tend to suppress

30
Q

where in the chromosome can you see loss of genes due to imprinting

A

15q11-13

31
Q

which imprinting disorder is maternal

A

anglemans

32
Q

why is DNA methylation easier to study

A

there are multiple methods, it is better studied and its highly stable over time regardless off how it is treated and collected

33
Q

what are genome studies

A

looking at a range of unspecific CpGs throughout the genome. Across Cpgs within report elements through out the genome

34
Q

what epigenome wide studies

A

detailed analysis of specific regions/cpGs across the genome

35
Q

what is gene/locus specific?

A

detailed analysis of specific region of genome

36
Q

wha are the 6 techniques used in methylation testing?

A

-restriction enzyme digest
-DNA sequencing
-immunoprecipitation
-fragment size analysis
-bisulfite modification
-DNA microarray

37
Q

how does cytosine extension assay work

A

HpaII restruction digests the unmethylated DNA
extension of the overhang using Taq polymerase and radio labelled cytosine

38
Q

What do the results of cytosine extension tell you

A

more radio active cytosine the lower methylation

39
Q

what are the advantages of cytosine extension

A

-cheap
-quanitifcation of methylation across the genome
-allows overall estimation of methylation between groups

40
Q

what are the disadvantages

A

-limited to CpGs in HpaII sites
-dont know where in the genome methylation differences are
-time consuming
-uses radio labelled cytosine

41
Q

what does MRE-Seq work?

A

cut with MSP1 and HPAII with one sample cut with just MSP1 and other just HPAII and then add adapters and then compare.

42
Q

what are the advantages of MRE-Seq

A

-quantification of methylation across genome
-allows overall estimation of methylation between groups
-sequencing method should allow for detection where changes are in genome

43
Q

what are disadvantages

A

-limited to CpGs in HpaII or other restriction sites
-expenisve

44
Q

how does MeDIP work

A

DNA is sonificated and then denatured. 5 methylcytidine Ab is used for immunoprecipation

45
Q

what two methods can you go onto to do with MeDIP

A

MeDIP seq and MEDIP-CIP

46
Q

What can MeDIP-Seq be used for

A

can compare healthy cells to cancer cells

46
Q

What can MeDIP-Seq be used for

A

can compare healthy cells to cancer cells

47
Q

what are the advantages of MeDIP

A
  • better knowledge of regions of the genome with methylation change
    -good genomic coverage
48
Q

what are the disadvantages of MeDIP

A
  • doesnt tell you the magnitude of methylation
    -cant resolve single CpG methylation changes
    -expensive
49
Q

what are the steps of covering cytosine to uracil in bisulphite modification

A

denature DNA and fragment it at 95 degrees and then do a conversion step by incubated it with sodium bisulphate at 65 degrees. Then do desulphitization which makes cytosine uracil.

50
Q

what is the differences between DNA with methylated and unmethylated with bisulphate modification

A

methyklateed has cytosine and unmethkated goes to uracil to thinene

51
Q

what are the advantages and disadvantages of whole genome bisulphite sequencing

A

-coverage of the whole genome
-exopensove
-lacks some precision

52
Q

what are the advantages/disadvantages of illumina infinitum methylation EPIC Bead chips

A
  • not always informative for distal regulators elements
    highly reproducible
    precise methylation quantification
53
Q

why measure methylation of specific genes

A

-interetested in a specific disease related gene
-hypotheiss driven research
-replication of findings
-biomarker of exposure
-predicitive disease biomarkers

54
Q

which methodology is most commonly used for gene specific methylation

A

bisulphite modification

55
Q

what are the advantages of methylation specific primers

A

relatively cheap
quick to develop
doesnt require specialist equipment

56
Q

what advantage does bisulphate have over MSP

A

a lot more qulaitive

57
Q

how do you do histone modification assays

A

Carry out DNA digestion by Mnase, then fragment DNA, then look for the specific histone modification that you want by using the antibody. Then fragments are pulled down again and sequenced.

58
Q

what is the histone code

A

post translational modification of key aa of the histone

59
Q

what does closed and open chromatin refer to

A

closed- trasncriptianallly silent
open- transcriptionally active

analytical exam 2 (6 decks)

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