Diagnostics Lec 8

Question 1

What is Duchenne muscular dystrophy

Answer 1

One of the most common and severe forms of muscular dystrophy

Question 2

who does DMD usually affect

Answer 2

boys in early childhood

Question 3

What does DMD increase over time

Answer 3

level of disability

Question 4

When can DMD become life threatening

Answer 4

if heart muscle or muscle for breath being affected

Question 5

What is the common lifespan with DMD

Answer 5

20s-30s

Question 6

What is the cause of DMD

Answer 6

Mutation in DMD gene in an X-linked recessive pattern

Question 7

DMD affects the protein dystrophin, what does the complete loss of dystrophin in DMD cause

Answer 7

muscular degenerative conditions

Question 8

What does defective or depleted dystrophin cause

Answer 8

another type of muscular dystrophy - Becker muscular dystrophy

Question 9

What is dystrophin

Answer 9

a very large protein encoded by 2.2Mb DNA, with 79 exons and 3685 amino acids

Question 10

How is DMD diagnosed

Answer 10

using multiplex ligation-dependent probe amplification

Question 11

What is multiplex ligation dependent probe amplification and how does it work

Answer 11

A multiplex PCR method, it detects changes in gene copy number

Question 12

What can multiplex ligation-dependent probe amplification be used for, why is this useful and what are the complications

Answer 12

Could be used to screen deletions or duplications of part of the DMD exons
Compliments sequence analysis which cannot detect copy number change
It is useful in analysis where large genes are involved
It is useful in the analysis of large genes where abnormalities can occur in many locations within that gene

Question 13

slide

Answer 13

5,6

Question 14

What are the working principles of MLPA

Answer 14

Pairs of target specific probes
Ligation
Universal primers
Fluorescent label
Capillary gel electrophoresis
Abundance of product proportional to bound probe which is proportional to number of target sequence probe bound to
Multiplex
Mobility modifier

Question 15

How does MLPA work

Answer 15

MLPA probes

Question 16

How do MLP probes typically work

Answer 16

specific to exon detection

typically measure 60-80 nucleotides

Question 17

How can MLPA probe ligated DNA fragment size be varied

Answer 17

via modification of mobility modifier

Question 18

What is analysed when looking for DMD

Answer 18

dystrophin gene
48 fragments analysed

Question 19

When analysing DMD results what do red and blue lines show

Answer 19

blue = test
red = control

Question 20

What are the features of a DMD female carrier

Answer 20

deletion of exon 43-50

heterozygous for dystrophin gene

Question 21

What are the features of a DMD affected male

Answer 21

deletion exon 46-50

mutation of single dystrophin gene