L17 Genomics in Drug Discovery RP

Question 1

what is a genome?

Answer 1

an organisms complete set of genes and chromosomes.

Question 2

what is genomics, and what can we achieve by understanding it?

Answer 2

the process of mapping, squencing and analysing genomes. Through understanding genomics, we can identify genes involved in a disease (novel genes).

Cloning genes to identify them may be useful when predicting what they do. This is ideal as ophan drugs can be matched with new treatment indications. e.g Albuterol - doesnt work on mexicans due to polymorphism on B2 receptors

Question 3

how many base pairs are there in the body?

Answer 3

3 billion base pairs in the body. With 2% encoding genes. Meaning there is only a 2% chance to obtain a mutated gene that codes for something like a protein.

Question 4

A gene is

Answer 4

a gene is a section of a chromosome in the nucleus of the cell

Question 5

what is the use of mRNA in genomics?

Answer 5

used to measure expression. mRNA is formed from DNA via RNA Polymerase

Question 6

what enzyme converts mRNA-> DNA?

Answer 6

Reverse Transcriptase

Question 7

what is splicing

Answer 7

process where useless genes are removed and the exons (useful genes) are joined together. mRNA splicing occurs to rearrgange the order of exons which is also how different varieties of antibiodies can be made.

Question 8

The process where useless genes are removed and the exons (useful genes) are joined together is known as

Answer 8

splicing

Question 9

what are DNA microarrays?

Answer 9

They’re able to measure the expression of 50,000 mRNA transcripts and 100,00 polymorphisms per experiment.

The appropriate oligonucleotide is matched with the opposite sequence of the mRNA strand.

Question 10

They’re able to measure the expression of 50,000 mRNA transcripts and 100,00 polymorphisms per experiment.

The appropriate oligonucleotide is matched with the opposite sequence of the mRNA strand.

Answer 10

DNA Microarrays

Question 11

what is reverse transcriptase PCR used to produce?

Answer 11

several copies of DNA from a single mRNA strand.

Question 12

several copies of DNA from a single mRNA strand is made by a process called

Answer 12

Reverse Transcriptase Polymerase Chain Reaction (PCR)

Question 13

Explain Microarray Expression Analysis

Answer 13

1) In this experimental setup, the cDNA derived from the mRNA of known genes is immobilized.
2) The probe (sample) has genes from both the normal as well as the diseased tissues.
3 The probe will hybridise onto the cDNA which bears a fluorescent marker.
4) This expression pattern is then compared to the expression pattern of a gene responsible for a disease.

Question 14

what does cDNA allow us to do?

Answer 14

cDNA allows us to compare healthy mRNA with faulty mRNA,

Healthy mRNA is the control and the regulation of specific genes are determined.

RT PCR is conducted for both healthy mRNA and faulty mRNA

Question 15

give me a scenario of cDNA fluorescent markers in use

Answer 15

so Green fluorescence can signifty healthy. and Red can signify faulty.

Question 16

Advantages of Genomics

Answer 16

It is fast as the expression levels for over 10,000 genes can be obtiained from the experiment.

-> It is comprehensive, as it deals with the entire human genome on two chips.

• > Flexible as can identify any SNP and genome
• > Used to identify parent groups that the drug would work in
• > It can be used to revive drugs that were thought to be never used

Question 17

Disadvantages of Genomics

Answer 17

1) Expensive process
2) Expression of mRNA may not reflect what occurs at the protein level
3) Data analysis process is very complex
4) Huge number of redundant data points
5) Can only use for genetic diseases

Question 18

In genomics, how do we identify significant genes?

Answer 18

Can be identified using stats. This can be achieved in three ways:

• Ranking can be used to identify the significant genes, by ranking the genes from highest -> lowest and taking the top 1%
• A Threshold can also be used. You can select which genes may have a 2X effect on a disease etc
• Inferrential and Descriptive statistics:

Inferrential Identifies significantly regulated genes, while descriptive indentifies co-regulated genes

Question 19

in gene identification what is the issue in ranking genes?

Answer 19

• It is likely that most crucial genes may be missed out, which could be towards the bottom
• The Top 1% may not be important.

Question 20

Describe the main tool used in statistical analysis in genomics

Answer 20

Inferential Stats: T-tests are done to identify the right order of genes to observe and the ones to cut off. This is achieved by conducting t-tests for every gene and accept these with a significance value of <0.05 (less than 0.05) However, some genes provide a false positive. The P value is implemented to correct this.

Question 21

How do you calculate the P value. And when the P value is calculated what does this even mean?

Answer 21

Original p value(0.05)/ square root of number of all samples. In the cas ethat a few genes gave a value lower than the p value, it needs to be confirmed that genes influence the changes that are expected as the values are lower than the P values, meaning thart the genes are signifnicant.

Question 22

What is the next step after identifying useful genes?

Answer 22

cloning the useful genes by RT PCR and then western blotting to confirm that those genes influence protein expression

Question 23

what is cDNA

Answer 23

cDNA is known to be synthesized from mRNA. It is synthesized in a reaction that is catalyzed by the reverse transcriptase and DNA polymerase enzymes.

Question 24

Compare and Contrast the Genomic and Traditional Drug discovery approach

Answer 24

Genomics is is a Genome wide screen approach that interrogates over 18,000 genes, and allows you to generate thousands of data points per experiment

By Contrast, Traditional DD looks Biological function, Addreses one Target or pathway and generates 1 data point per experiment.

Question 25

Compare and Contrast HTS with Traditional DD and Genomics

Answer 25

Genomics uses Multi Arrays, and you can generate up to 48 million data points.

Traditional DD produces 96 data points per experiment, using 96 wells for HTS

Question 26

What was the initial aim of the Human Genome Project?

Answer 26

Identify role of gene products, how they are controlled, how they interact,
and their involvement in human disease

Question 27

what was the findings of the HGP?

Answer 27

Identity of 90% of genes are known
Structure and function of 75% of these are known
40 genes obtained directly from bacteria
40% of DNA is repetitive

Question 28

what did we find out about SNPs?

Answer 28

– SNPs define individuality, most are neutral, many are disease causing