Download Molecular Technologies and Diagnostics

Molecular Technologies and
Diagnostics
Classic DNA Lab Technologies
• Enzyme digestion and electrophoresis
• Sequencing
– Single Nucleotide Polymorphisms (SNPs)
• Southern blot
– Linkage analysis can predict inheritance
• PCR
– A short DNA sequence can be amplified
a billion times
Electrophoresis
• For separation of DNA
fragments that migrate through
a gel
• Shape and size of DNA
determine mobility
• For visualizing genetic
differences
• DNA appears as “bands” in the
gel after staining
Restriction Enzymes
• Active proteins purified from bacteria
• “Molecular scalpels” that cut (digest) double-stranded
DNA
• Recognize specific DNA sequences
• Mutations change DNA sequence, therefore also change
the cutting pattern of restriction enzymes
• Used with PCR, Southern blot, etc.
• Required for cloning techniques
Sequencing
It was discovered that
there are 20 – 25,000
genes in the human
genome, far fewer than
previous estimates.
Sequencing Identifies SNPs
Single Nucleotide Polymorphisms (SNPs)
SNPs are Copying Errors
•Cells sometimes make mistakes during the copying process – (typos). These
typos lead to variations in the DNA sequence at particular locations, called
SNPs
The Consequences of SNPs
•Biological diversity
•Harmful mutations
•Beneficial mutations
•Neutral mutations
SNPs as a Measure of Genetic Similarity
•DNA is passed from parent to child, so you can inherit SNPs
•Number of matching SNPs can determine family relations
Southern Blot
This procedure is tedious, used
less and less, but is an excellent
research and diagnostic tool.
Southern Blot on Family Members Can
Predict Inheritance
Southern Blot
• Laboratory method used to detect specific DNA molecules
from among a many other DNA molecules
• Named after its inventor, Edward Southern
• Can be used to analyze an organism's total DNA (genome)
in order to identify a specific sequence of interest
http://highered.mcgrawhill.com/olcweb/cgi/pluginpop.cgi?it=swf::535::535::/sites/dl/free/0072437316/1200
78/bio_g.swf::Southern%20Blot
Southern Blot for T-cell Lymphoma
M 1 2 3 4 5 6 7 8 9
PCR – Polymerase Chain Reaction
amplification of a specific DNA sequence
Conventional end-point PCR (left)
is rapidly being replaced by real –
time PCR which is faster and uses
fluorescently labeled DNA (below)
New DNA Technologies - Arrays
• Microarrays
– DNA “chips”
– Many sequences analyzed at once
– Subtle cellular changes can be detected
– Gene expression
High Throughput Robotics
New DNA Technologies Genomics
• Pharmacogenomics
– Genetic variety =
metabolic differences
– The correct, safest drug
the first time
– Fewer adverse drug
reactions (ADRs)
– Reduced cost of
healthcare
http://www.pbs.org/wgbh/nova/body/personal-dna-testing.html (Video: 12:35)