Download DNA marker analysis - Central Magnet School

DNA marker analysis
Mrs. Stewart
Medical Interventions
Central Magnet School
Standard:
Marker analysis is a technique used to
determine the presence of genetic
mutations associated with cancer.
Objective:
Investigate the marker analysis technique
and analyze results to determine the
presence of a BRCA 2 mutation
BRCA 1 and BRCA 2
Tumor suppressor genes
Repair DNA damage and control cell
growth
Proto-oncogenes
BRCA 2
80,000 nucleotides (larger than average
gene)
600+ mutations identified and linked to
increased risk of cancer
Most mutations are insertion or deletion
mutations (frameshift) of more than one
base
Results in a protein that is unable to help
repair damaged DNA or fix mutations
Marker Analysis Technique
A technique where a gene mutation is
analyzed by using a genetic marker
instead of directly analyzing the gene
itself.
Genetic marker: Alteration in DNA that
may indicate an increased risk of
developing a specific disease or disorder
STRs
Genetic markers used in marker analysis
are short DNA sequences called Short
Tandem Repeats (STRs)
STRs – region of DNA composed of a
short sequence of nucleotides repeated
many times.
# of repeated sequences vary
Alternate forms of STRs correspond
with different alleles.
STRs
Most STRs occur in gene introns (non-coding regions
of DNA)
Does not usually affect gene function
Can use as “markers” to differentiate between different
alleles for certain genes
(because genes located next to each other are inherited
together.)
PCR
The region of the DNA that is the known STR marker
is amplified (and the BRCA unknown gene version
with it)
The amplified DNA is then run on a gel.
Gel Electrophoresis
Because different alleles have a different number
of repeats present in the STR, gel electrophoresis
will separate different alleles based on the number
of repeats present.
The more repeats present = the longer the DNA
fragment
Shorter DNA fragments migrate farther down the
gel
So, the fragments that migrate the farthest, have
the fewest STRs
Smith Family Analysis
We will determine different alleles for
each family member tested by
determining the band lengths for each
family member.
Who was tested?
Diana –Judy’s sister
Jennifer – Judy’s sister
Laura – Judy’s mom
Judy – Sue, Mike and Tucker’s mom
Who has the BRCA 2 mutation?
Each person has 2 chromosomes #13, so each
person will have 2 alleles for the BRCA 2
gene.
You will have to identify which allele is linked
to the “mutant” gene by determining which
alleles Jennifer and Laura have in common
Since both of them are known to carry that
allele.
Analyze Judy and Diana’s results to determine
if they also carry the mutated gene allele.
Analysis
DNA size markers are loaded in the first well.
Use the size markers to determine the sizes of
your unknown fragments.
The known molecular size markers (weights
in base pairs) are written beside each band.
Measure the distance each unknown band
migrated
Table One:
Data
DNA Size
Fragment
Markers
Length in
Distance Migrated
Base Pairs
Distance
Distance to
Rf
Migrated
Reference
(mm)
- Measure
the distance in mm
Point (mm)
A÷B
well to (mm)
each
fragment
in
the
standard
A
B
from the sample
lane. Record in Table One
Fragment 1
Distance
Fragment
2
1353
to 1078
Reference Point - Measure the distance
from the sample well to the end of the gel. Record in
Fragment 3
872
Table One.
Fragment 4
603
This number
will be the same for each size marker
Fragment
5
310
fragment.
Fragment 6
281
Calculate the Rf of each fragment and record in Table
Fragment
234
One. 7
Fragment
8
194 to the nearest one hundredth.
Round
values