Molecular Biology of the Cell by Bruce Alberts, Alexander Johnson, Julian Lewis, David Morgan, Martin Raff, Keith Roberts, Peter Walter by by Bruce Alberts, Alexander Johnson, Julian Lewis, David Morg

STUDYING GENE EXPRESSION AND FUNCTION
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The nucleotide sequence of any genome can be determined rapidly and simply
by using highly automated techniques based on several different strategies. Comparison
of the genome sequences of different organisms allows us to trace the evolutionary
relationships among genes and organisms, and it has proved valuable for
discovering new genes and predicting their functions.
Taken together, these techniques for analyzing and manipulating DNA have
made it possible to sequence, identify, and isolate genes from any organism of interest.
Related technologies allow scientists to produce the protein products of these
genes in the large quantities needed for detailed analyses of their structure and
function, as well as for medical purposes.
STUDYING GENE EXPRESSION AND FUNCTION
Ultimately, one wishes to determine how genes—and the proteins they encode—
function in the intact organism. Although it may seem counterintuitive, one of the
most direct ways to find out what a gene does is to see what happens to the organism
when that gene is missing. Studying mutant organisms that have acquired
changes or deletions in their nucleotide sequences is a time-honored practice
in biology and forms the basis of the important field of genetics. Because mutations
can disrupt cell processes, mutants often hold the key to understanding
gene function. In the classical genetic approach, one begins by isolating mutants
that have an interesting or unusual appearance: fruit flies with white eyes or curly
wings, for example. Working backward from the phenotype—the appearance or
behavior of the individual—one then determines the organism’s genotype, the
form of the gene responsible for that characteristic (Panel 8–2).
Today, with numerous genome sequences available, the exploration of gene
function often begins with a DNA sequence. Here, the challenge is to translate
sequence into function. One approach, discussed earlier in the chapter, is to
search databases for well-characterized proteins that have similar amino acid
sequences to the protein encoded by a new gene. From there, the protein (or for
noncoding genes, the RNA molecule) can be overexpressed and purified and the
methods described in the first part of this chapter can be employed to study its
three-dimensional structure and its biochemical properties. But to determine
directly a gene’s function in a cell or organism, the most effective approach
involves studying mutants that either lack the gene or express an altered version
of it. Determining which cell processes have been disrupted or compromised in
such mutants will usually shed light on a gene’s biological role.
In this section, we describe several approaches to determining a gene’s function,
starting either from an individual with an interesting phenotype or from a
DNA sequence. We begin with the classical genetic approach, which starts with
a genetic screen for isolating mutants of interest and then proceeds toward identification
of the gene or genes responsible for the observed phenotype. We then
describe the set of techniques that are collectively called reverse genetics, in which
one begins with a gene or gene sequence and attempts to determine its function.
This approach often involves some intelligent guesswork—searching for
similar sequences in other organisms or determining when and where a gene is
expressed—as well as generating mutant organisms and characterizing their phenotype.
Classical Genetics Begins by Disrupting a Cell Process by
Random Mutagenesis
Before the advent of gene cloning technology, most genes were identified by the
abnormalities produced when the gene was mutated. Indeed, the very concept of
the gene was deduced from the heritability of such abnormalities. This classical
genetic approach—identifying the genes responsible for mutant phenotypes—is
most easily performed in organisms that reproduce rapidly and are amenable to
genetic manipulation, such as bacteria, yeasts, nematode worms, and fruit flies.
Although spontaneous mutants can sometimes be found by examining extremely