Open Challenge Question 1 You have been approached by a researcher studying colo
ID: 273505 • Letter: O
Question
Open Challenge Question 1
You have been approached by a researcher studying colon cancer who suspects that
epigenetic modifications are contributing to the tumor phenotype. They are seeking your
advice on what experiments they should run to determine if disruption to the epigenetic
state is playing a role in this cancer.
Based on what you learnt from exploring the datasets using RNAseq and ChIP- seq , what experiments
would you advise that this researcher perform? Specifically, make reference to
experimental data you think would have improved your understanding of, or allowed
further analysis of, the breast cancer phenotype in this experiment.
Explanation / Answer
In the early years of the molecular biology revolution, cancer research was mainly focused on genetic changes (ie, those that altered DNA sequences). Although this has been extremely useful as our understanding of the pathogenesis and biology of cancer has grown and matured, there is another realm in tumor development that does not involve changing the sequence of cellular DNA. This field is called “epigenetics” and broadly encompasses changes in the methylation of cytosines in DNA, changes in histone and chromatin structure, and alterations in the expression of microRNAs, which control the stability of many messenger RNAs and serve as “master regulators” of gene expression.
Cancer epigenetics is the study of epigenetic modifications to the DNA of cancer cells that do not involve a change in the nucleotide sequence.
Popular approaches for measuring CpG methylation in cells include:
The HpaII tiny fragment Enrichment by Ligation-mediated PCR Assay (HELP Assay) is one of several techniques used for determining whether DNA has been methylated. The technique can be adapted to examine DNA methylation within and around individual genes, or it can be expanded to examine methylation in an entire genome.
The technique relies upon the properties of two restriction enzymes: HpaII and MspI. The HELP assay compares representations generated by HpaII and by MspI digestion of the genome followed by ligation-mediated PCR. HpaII only digests 5'-CCGG-3' sites when the cytosine in the central CG dinucleotide is unmethylated, the HpaII representation is enriched for the hypomethylated fraction of the genome
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