4. What Is MOLECULAR GENETICS?
Molecular genetics is the study of the agents
that pass information from generation to
generation. These molecules, our genes, are
long polymers of deoxyribonucleic acid, or
DNA. Just four chemical building blocks
guanine (G), adenine (A), thymine (T), and
cytosine (C)—are placed in a unique order to
code for all of the genes in all living
organisms.
7. Genes determine hereditary traits, such as the color of our hair
or our eyes. They do this by providing instructions for how every
activity in every cell of our body should be carried out. For example,
a gene may tell a liver cell to remove excess cholesterol from our
bloodstream. It will instruct the cell to make a particular protein. It
is this protein that then carries out the actual work.
Many diseases are caused by mutations, or changes in the DNA
sequence of a gene. When the information coded for by a gene
changes, the resulting protein may not function properly or may
not even be made at all. In either case, the cells containing that
genetic change may no longer perform as expected.
8. Isolating DNA from just a single cell provides a complete
set of all a person's genes, that is, two copies of each gene.
However, many laboratory techniques require that a
researcher have access to hundreds of thousands of copies
of a particular gene. One way to obtain this many copies is
to isolate DNA from millions of cells grown artificially in
the laboratory.
Another method, called cloning, uses DNA manipulation
procedures to produce multiple copies of a single gene or
segment of DNA. The polymerase chain reaction (PCR)
is a third method whereby a specific sequence within a
double-stranded DNA is copied, or amplified. PCR
amplification has become an indispensable tool in a great
variety of applications.
9. Cell culture involves growing cells under artificial
conditions, either attached to some type of artificial
surface or suspended in a special solution. In both
cases, the cells are bathed in fluids containing nutrients
that are either synthetically produced or extracted from
related organisms. Certain cell types are more amenable
to being grown in culture than others. Conditions that
serve to sustain one cell type may not apply to other cell
types, or even the same cell type from another species.
Cell culture is a useful technique because it provides a
renewable source of cells for isolating DNA.
10. DNA isolation refers to the process of extracting DNA from a cell in a
relatively pure form. It involves separating DNA from other cellular
components, such as proteins, RNA, and lipids. Whatever the source,
the DNA is isolated by placing the cells in a tube containing a special
solution, called a "cocktail", and mechanically or chemically breaking
them open. This causes the cell to release its contents into the cocktail
containing enzymes, chemicals, and salts. Enzymes are used to chew up
the proteins; chemicals to destroy any RNA present; and salts to help
pull the DNA out of solution. At this point, the DNA will exist in long
strands that form a mucous-like glob within the solution. The DNA is
then harvested by spinning the tube in a machine called a centrifuge.
During spinning, the DNA collects in the bottom of the tube. The
solution is then poured off, and the DNA is dissolved in a second
solution that will make it easy to work with in subsequent procedures.
The result is a concentrated DNA sample containing many thousands of
copies of each gene.
11. In the first step of mRNA isolation, a cell is ruptured, and the
cellular contents are exposed to synthetic beads coated with
strings of thymine nucleotides.
Because adenine and thymine readily bind to each other,
poly(A) mRNA is selectively retained on the beads while the
other cellular components are washed away. Once isolated,
purified mRNA is converted to single-stranded DNA using the
enzyme reverse transcriptase and is then made into a stable
double-stranded DNA using the enzyme DNA polymerase.
DNA produced in this way is called complementary
DNA (cDNA) because its sequence, at least the first strand, is
complementary to that of the mRNA from which it was made.
12. mRNA Isolation
This drawing demonstrates how poly(A) RNA can
be isolated from other RNAs by separation on a
special solid support material. In this example, the
material is made up of glass beads to which
thymine molecules are attached. Because adenine
and thymine molecules readily bind to each other,
mRNAs with poly(A) tails will be selectively
retained on the beads. As seen on the left-hand
side of the diagram, a solution containing various
RNA populations, including mRNAs with poly(A)
tails (red) as well as other RNAs and cellular
material (purple), is applied to the separation
column. Only the poly(A) RNA is retained, because
it is immobilized on the solid support material.
The other RNAs and cellular material pass through
the column. On the right, the bound poly(A)
mRNA is retrieved by treating the column with a
special buffer solution that breaks the thymine
nucleotide–AAA bond. The mRNA can be collected
in a tube for further experimentation.
13. The porous and thin nature of a gel is ideal for separating
DNA fragments using electrophoresis, but as we
mentioned earlier, these gels are delicate and rarely usable
for other techniques. For this reason, DNA that has been
separated by electrophoresis is transferred from a gel to an
easy-to-handle inert membrane, a process called blotting.
Term "blotting" describes the overlaying of the membrane
on the gel and the application of a pad to ensure even
contact, without disturbing the positions of the DNA
fragments. This procedure reproduces the exact pattern of
DNA captured in the gel on the membrane. The membrane
can then be probed with a DNA marker to verify the
presence of a target sequence.
14. The process of determining the order of the nucleotide
bases along a DNA strand is called sequencing. In
1977, 24 years after the discovery of the structure of
DNA, two separate methods for sequencing DNA were
developed: the chain termination method and
the chemical degradation method. Both methods
were equally popular to begin with, but, for many
reasons, the chain termination method is the method
more commonly used today. This method is based on
the principle that single-stranded DNA molecules that
differ in length by just a single nucleotide can be
separated from one another using polyacrylamide gel
electrophoresis, described earlier.
15.
16. Answer the
following
What is What questions.
Molecular happens
Genetics? during RNA When you
Isolation? already saw
the picture,
give their
What is Give the
four essence to
Cell building Human
Culture? blocks.
Life.
What Is CLONNING?
17. Stick It On! Translate the Question using the STICKS Graph.
Then, well of course, ANSWER IT.
J K L S W
A B C
T U X Y
D E F M N O
V Z
G H I P Q R
QUESTION:
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