L25- Interactive Sickle Cell diagnosis

Question 1

case study story

Answer 1

• Healthy couple, Ken and Carol, originally from Kenya.
• First baby boy, Keith, born in Kenya, healthy.
• Family moved to the UK for postgraduate studies, while Carol pregnant again.
• Second baby, Celia, born in the UK.
• At 3 months Celia developed problems and was referred to a paediatrician.

Question 2

examination report for baby celia

Answer 2

• Painful swelling of fingers
• Pale complexion
• Enlarged spleen
• Low haemoglobin levels of 7 g/dl
• Reticulocyte count 12%
• High reticulocytes

Question 3

High reticulocytes due to

Answer 3

body trying to replace RBC

Question 4

examiantion of blood film from baby celia

Answer 4

Sickled erythrocytes

Question 5

sickled erythrocytes due to

Answer 5

mutation B suninit of haemoglobin

• makes them stick together as polymers
• cells become less elastic and more likely to burst

Question 6

family tree

Answer 6

Question 7

diamond

Answer 7

unknown sex

Question 8

conclusions from family tree

Answer 8

• Parents must be carriers (heterozygous)-
• 1/4 chance that the unborn child has sickle cell
• 3/4 chance will be unaffected

Question 9

Methods use to anlysis Celias DNA first

Answer 9

PCR

Question 10

PCR

Answer 10

Can use very little amount of DNA using very specific DNA primers for the HBB gene

Question 11

primers define the

Answer 11

amplification area- primers incorporated in final product

Question 12

technique used with PCR

Answer 12

restriction analysis, gel electrophoresis, DNA sequencing, Southern blotting

Question 13

HBB open reading frame (ORF)

Answer 13

part of the gene translated as a protein- after rmeoval of introns

Question 14

where is the HBB gene found

Answer 14

Chromosome 11, location 11p 15.6

Question 15

what type of mutation is sickle cell disease caused by

Answer 15

single based substitution

A –> T

glutamate –> valine (Glu6Val)

Question 16

substiution of A –> T destroys the

Answer 16

MstII restriction site

MstII recognises 5’CCTNAGG

Question 17

glutamate –> valine (Glu6Val) meaning

Answer 17

(methionie (ATG) at the beginning of the ORF does not count as part of the gene - so ignore then count in 6 to find the Hbb mutation

Question 18

Answer 18

Answer: A and C

A- would PCR the whole of the Hbb gene

C- straddles the restriction site so could be used

Question 19

Answer 19

Answer: C

e. g. 18bp primer at either end of the gene
- Primers will be incorporated

Question 20

Answer 20

B

Question 21

Answer 21

C

Question 22

Answer 22

Answer: D

RBC do not have DNA

Question 23

Answer 23

Answer: C
We would need to use reverse transcriptase to produce cDNA so that Taq polymerase could start synthesis