Lecture 3: Clinical Applications of DNA Technology Flashcards
What is the best way to describe the mode of inheritance for Fragile X syndrome
X-linked dominance with reduced penetration
Where does the risk for a full mutation for Fragile X come from (who had the premutation?)
carrier mother
premutation grandfather
Which restriction enzyme - EagI or EcoR1 - only cuts unmethylated DNA?
EagI
On a southern blot of an X chromosome from a normal male: if digested with EagI-EcoR1, what size kb fragment, and how many fragments, will there be? Is this fragment methylated?
1x 2.8kb fragment
Not methylated - active X chromosome
On a southern blot of the X chromosomes from a normal female: if digested with EagI-EcoR1, what size kb fragment, and how many fragments, will there be? Will they be methylated?
1x 2.8kb fragment - unmethylated (active xs)
1x 5.2kb fragment - methylated (inactive xs)
In a full fragile X mutation, how many repeats are present, and of of what sequence?
CGG>200
How many bands will be seen on a Southern blot of a premutation female for fragile X after EagI-EcoR1 digest?
4 bands
2 normal, 2 premutation
both methylated and unmethylated
What two cellular conditions result from CGG>200 repeats?
abnormal methylation
mitotically unstable
What are hemoglobinopathies?
mutations in the alpha or beta globin chain genes
What do alpha and beta globin chains do?
Complex together with heme to carry O2 in the blood
What are thalassemias?
Mutations in globin chain genes that cause an imbalance of QUANTITY of the alpha and beta chains
What is the result of a sickle cell mutation?
Structural changes (QUALITY) in the beta globin gene
Which restriction sites does the sickle cell mutation affect?
Mnl I
Dde I
What kind of gene mutations occur in beta thalassemia?
point mutations in beta globin gene
Why isn’t beta thalassemia detected before birth?
have gamma globin before birth
beta globin comes in after birth
what is the normal conformation of the beta globin gene?
cluster of 6 beta globin genes; 5 active, 1 inactive (=pseudogene)
What happens with the alpha globin protein in beta thalassemia?
all the alpha can’t complex with beta
too much alpha
forms Heinz bodies (alpha4 homotetramers)
How do Heinz bodies form, and what do they do?
excess alpha globin can’t bind beta globin
precipitate –> destruction of RBC
What causes alpha thalassemia?
misalignment, unequal crossing over
results in deletion –> chromosomes have one alpha gene instead of 2
What is alpha thalassemia 1?
2 alpha globin genes deleted from same chromosome
what is alpha thalassemia 2?
1 alpha globin gene deleted from each of 2 chromosomes
What structures coordinate the appropriate developmental expression of the alpha and beta globin genes within clusters?
locus control regions
What is the alpha Hb Constant Spring Mutation?
mutation of stop codon at codon 142 –> longer, unstable alpha chain
What forms in an individual with the alpha Hb Constant Spring mutation and 2 deleted alpha globin genes?
beta globin tetramer (Hb H) formation
What is the most common mutation in cystic fibrosis?
F508
For what disease have 1900 disease-causing mutations been detected?
Cystic Fibrosis
What enzyme does Third Wave technology use? What is it used for?
Cleavase
used to detect most common Cystic Fibrosis mutations
Where does cleavase cut the invading probe?
ONLY where/if it matches target nucleotide
What happens in Third Wave technology after cleavase cuts?
flap is produced
flap is recognized by second cassette
fluorescent signal is produced
–> nucleotide being tested is present
What can Next Gen Sequencing lead to?
individualized treatment
How many functional beta globin genes does a normal person have?
2 (1 on each xs 11)
