Biomolecules and
DNA Analysis
Forensics and DNA fingerprinting
Detecting Carbohydrates
• To detect monosaccharides and most
disaccharides (reducing sugars) in stomach
contents, a sample would be mixed with
Benedict’s solution.
• A positive result is seen by the formation of a
brownish-red cuprous oxide precipitate.
• You will test dextrose as a known for a positive
result.
Detecting Carbohydrates
• Iodine is typically used to detect starch. It
turns a dark blue/black in the presence of
starch.
• You will test a known starch solution to see
the result before testing the unknown
“stomach contents.”
Detecting proteins
• Biuret solution is used to detect the presence
of proteins.
• To observe the known reaction, you will mix
biuret with albumin. A pinkish-purple color is
observed if proteins are present.
Detecting Lipids
• Sudan III crystals are used to detect the presence
of lipids.
• The procedure is important to note! A nonpolar
solvent, like hexane is mixed with the sample and
then poured off into an evaporating dish.
• The solvent must be completely evaporated
before the crystals are streaked across the dish. A
red streak is a positive test for lipids. No color
change indicates no lipids are present.
Detecting Vitamin C
• The addition of 2,6-dichloroindophenol, which
is a blue solution, turns light amber to
colorless in the presence of ascorbic acid
(vitamin C).
• You will test a solution of ascorbic acid to see
the positive test.
• Then you should perform the test on the
stomach contents to determine if vitamin C is
present.
DNA
• Except for identical twins, no two people on
earth have the same DNA.
• Since the 1980s, DNA has been used to solve
crimes, identify victims, and establish
paternity.
• It is especially useful in large-scale disasters,
like plane crashes, hurricanes and tsunamis to
identify victims.
Review of DNA
• DNA holds all the information needed to make
proteins and to replicate itself.
• The genetic material is stored in
chromosomes. If you dissolved the histone
packaging, you would see the double helix
strand of DNA tightly coiled around protein
molecules and itself.
• If you could unwind the DNA, it would
measure 6 feet in length!
DNA Structure
• DNA is composed of a sugar-phosphate backbone and ladder
of nitrogenous base pairs.
• Adenine pairs with thymine
• Cytosine pairs with guanine
• There are 46 chromosomes in the nucleus of most cells
(except sex cells…they have 23). This is called nuclear DNA.
• One chromosome pair is inherited from the mother and one
from the father, so each person inherits exactly half of their
genetic information from each parent.
• Mitochondrial DNA is in a circular loop and is inherited only
from the mother. It is passed to the offspring in the cytoplasm
of the egg cell.
Genes and Alleles
• Genes are DNA sequences that have instructions that
determine our inherited traits.
• Examples: blood type, hair color, eye color, etc.
• An allele is one of two or more alternative forms of a
gene. One allele comes from the mother and one
from the father.
• Alleles are what is analyzed for dominant and
recessive traits.
• The human genome is the total amount of DNA in the
cell.
Coding and Noncoding
genes
• The chromosomes in the nucleus of the cells
contain 23, 688 encoded genes.
• This accounts for 1.5% of the DNA in the cell.
• The other 98.5% is noncoding and used to be
referred to as junk DNA. However, more
research is proving that even noncoding DNA
is important in gene splicing.
Using DNA for
Identification
• In the noncoding DNA, there is variation
among humans. There are repeated base
sequences that are unique to individuals.
• Certain sequences are repeated many times.
These differences are referred to as
polymorphism.
DNA Fingerprinting
• Dr. Alec Jeffreys determined that these repeating
sequences could be isolated and analyzed. Thus,
the beginning of DNA fingerprinting. (1984)
• Two types of repeating sequences are used: STRs
(short tandem repeats) and VNTR (variable
numbers of tandem repeats).
• These repeating sequences show up as different
bands in electrophoresis separation techniques.
VNTR
• Certain short sequences of noncoding bases
repeat multiple times.
• However, the number of times varies in each
individual. One individual may have the sequence
repeat 3 times while another is 7 times.
• Since the number of repeats varies, it is called
variable number of tandem repeats (VNTR).
• The length of repeating bases in a sequence can
vary from 9-80 bases.
STR
• The DNA sequences with a high degree of
polymorphism are most useful for DNA analysis.
• These sequences are short- only 2-5 bases in
length in the noncoding DNA.
• This method is useful by analyzing how many
times the base sequence repeats in an individual.
• This is becoming the preferred method of analysis
because the long VNTR sequences are harder to
separate clearly.
DNA Profiles
• When several STR or VNTR sequences are
examined, the DNA profile or fingerprint of a
person is developed that is unique to that
individual.
• This information is used in 2 ways:
• 1) Matching a tissue to see if it is from the same
person. All bands match. (victim/perp ID)
• 2) Inheritance markings- since half of genetic
material is from mother and half from father…half
of the bands will align with a parent.
Sources of DNA
• Saliva, blood, seminal fluid, skin and hair may
all contain DNA left at a crime scene.
• Since very small (or trace) amounts of DNA
might be all that is available at the scene, a
technique was needed to multiply the DNA to
have enough to analyze.
• The use of the PCR (polymerase chain
reaction) is employed to replicate the DNA
samples
Collecting DNA
• 1) Wear disposable gloves and change them often.
• 2) Use disposable instruments for handling the
sample (pipets, containers, etc.)
• 3) Avoiding touching, talking, coughing on,
sneezing on areas where DNA is suspected to be.
• 4) Air-dry evidence before packaging in a paper
envelope or bag.
• 5) If you cannot dry wet evidence before
packaging, it can be frozen.
DNA Separation
• When the specific enzymes are used to cut the
DNA in different fragments, the fragments are
different sizes so they are different masses.
• The process of gel electrophoresis will allow
them to be separated based on their mass.
• As more technology is developed, the
different fragments are able to be separated
by mass spectrometers based on their time of
flight between charged plates.
DNA Fingerprinting
• 1) DNA must be extracted from its cells in the
evidence sample.
• 2) DNA is “cut” by using restriction enzymes that
recognize a unique base pattern in the DNA.
• 3) Amplification using the polymerase chain
reaction (PCR).
• 4) Electrophoresis or other methods of separating
fragments by mass. Smallest fragments move
fastest through the gel.
Electrophoresis
• A gel, of aragose, is made which is porous.
• A buffer solution is placed in the gel tray. This aids
the movement of the DNA based on differential
charge.
• A power supply is attached and must be in contact
with the solution. The DNA is placed in wells in the
gel that is towards the negatively charged end of
the gel.
• The DNA moves towards the positive charge. The
smaller fragments move faster, and the larger
fragments move more slowly. These create a band
pattern that is unique for each individual.