Advanced Molecular Techniques

Question 1

How is a DNA microarray made?

Answer 1

Robot takes a set of fountain pens and dips them in each ink wells in a 96-well plate, containing the DNA. Each well represents a different human gene. The pens print DNA ink in array on microscope slide ~30,000 spots. Finally producing an array that represents about thirty thousand different human genes

Question 2

What is conditional gene expression?

Answer 2

Looking at mRNA hybridised on a micro array slide

Question 3

When would Array comparative genome hybridisation be used?

Answer 3

uses all DNA to compare genes present – allows analysis of deletions, duplications in genome

Question 4

What does RT-PCR stand for and what is it?

Answer 4

reverse-transcriptase polymerase chain reaction: used to clone RNA expressed genes by reverse transcribing the RNA of interest into its DNA complement through the use of reverse transcriptase. Subsequently, then amplified using PCR. Primer to poly A tail

Question 5

What is the difference between DNA fingerprinting and DNA profiling?

Answer 5

DNA fingerprinting –> from introns - uses repeat sequences = mini satellites, 30-60bp, highly variable, one from mum one from dad, run on PCR, compare bands.

DNA profiling (modern) –> short tandem repeats (STRs) = similar between closely related humans, but are so variable that unrelated individuals are extremely unlikely to have the same. Look at 16 STR regions = 32 diff bands, run on PCR, plot graph = peaks = number of bases in repeats

Question 6

What is karyotyping?

Answer 6

preparation of metaphase cells, identify and evaluate the size, shape, and number of chromosomes

Question 7

Describe FISH

Answer 7

FISH - fluorescent in situ hybridisation –> fluorescent probes that bind to only those parts of the chromosome with a high degree of sequence complementarity – identify where gene is on chromosome , control probe, can do chromosome painting = allows identification of each chromosome, translocations