Advanced Molecular Techniques

Question 1

Give examples of analysis of DNA at the nucleotide level.

Answer 1

DNA sequencing

PCR + restriction analysis/DNA sequencing

Question 2

Give examples of analysis of DNA at the gene level.

Answer 2

Southern hybridisation
Northern hybridisation
RT-PCR
Microarray
DNA fingerprinting/DNA profiling

Question 3

Give examples of analysis of DNA at the chromosome level.

Answer 3

Karyotyping

FISH/Chromosome painting

Question 4

What are some ethical problems regarding DNA sequencing.

Answer 4

Who would be interested in your genome information
Can the knowledge help prevent illness later in life
Does it open up areas for discrimination for example by insurance companies
Who owns ‘your’ DNA sequence

Question 5

What is RT-PCR or Reverse transcriptase PCR?

Answer 5

It works very similar to normal PCR. However it is used to look at RNA instead of DNA. A DNA copy is made from the mRNA and then continuously amplified.

Question 6

Give an example of how Microarray can work.

Answer 6

Two conditional gene expressions are looked at:
Cancer cells and normal cells.
RNA is isolated and the mRNA is coded to cDNA which is single stranded DNA done by reverse transcriptase.
The different DNAs are then labeled by two different fluorochromes and combined and hybridised to microarray. Here we can compare the two conditions.

Question 7

What is DNA fingerprinting/profiling? How does it work?

Answer 7

DNA fingerprinting is not looking at genes. Its looking at non-coding areas. The amounts of repeats are looked at in the non-coding areas. The non-coding areas are then spliced off and put into DNA gel electrophoresis.