Page 88 - Genetics_From_Genes_to_Genomes_6th_FULL_Part2

7.5 What Mutations Tell Us About Gene Function 247

Benzer mapped 1612 spontaneous point mutations and Hotspots of mutation
several deletions in the rII locus of bacteriophage T4 Some sites within a gene mutate spontaneously more often
through recombination analysis. He first used recombina- than others and as a result are known as mutation hotspots
tion to determine the relationship between the deletions. (Fig. 7.25c). The existence of hotspots suggests that certain
He then found the approximate location of individual point nucleotides can be altered more readily than others. Treat-
mutations by observing which deletions could recombine ment with mutagens also turns up hotspots, but because
with each point mutant to yield wild-type progeny. mutagens have specificities for particular nucleotides, the
highly mutable sites that turn up with various mutagens are
often at different positions in a gene than the hotspots
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Determining RF between rII mutations resulting from spontaneous mutation.
for precise mapping Nucleotides are the same chemically whether they lie
Benzer next performed recombination tests to measure the within a gene or in the DNA between genes. Furthermore,
genetic distance between pairs of point mutations he had as Benzer’s experiments imply, the molecular machinery
found by deletion mapping to lie in the same small region responsible for mutation and recombination does not dis-
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of the chromosome. The distance between any two rII mu- criminate between those nucleotides that are intragenic
tants could be measured simply by counting the number of (within a gene) and those that are intergenic (between
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rII and total phages in an aliquot of lysate from a phage genes). The main distinction between DNA within and
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cross. The RF (in map units) is simply the number of rII DNA outside a gene is that the array of nucleotides com-
recombinants [plaques on E. coli K(λ)] divided by the total posing a gene has evolved a function that determines phe-
number of phages (plaques on E. coli B), multiplied by 2 to notype. Next, we describe how geneticists discovered what
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account for the rII double mutant reciprocal recombinants that function is.
that must exist but cannot be detected easily:
2[number of plaques on E. coli K(λ)]
RF = essential concepts
(number of plaques on E. coli B)
• A complementation test determines whether two different
Benzer combined the results from deletion mapping
and RF calculations to produce a map of the fine structure recessive mutations occur in the same gene or in different
genes.
of the rII region (Fig. 7.25c). Note that all of the point
mutations in the rIIA complementation group mapped to • At the DNA level, a gene is a linear sequence of
nucleotide pairs in a discrete region of a chromosome
one side of the rII region, and all of the rIIB point muta- that confers a specific unit of function.
tions mapped to the other side.
• Recombination can occur between any two nucleotide
pairs, whether they are within the same gene or not.
How DNA nucleotides are organized into genes
Benzer knew that the genetic distances between all mapped
genes in the T4 genome add up to about 1500 m.u. He also
knew that the T4 chromosome constitutes about 169,000 bp 7.5 What Mutations Tell Us
of DNA, so he could calculate that for bacteriophage T4, About Gene Function
1 m.u. corresponds to 169,000/1500 = 113 bp. The lowest
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RF that he measured between any pair of rII mutants was
0.02 m.u., which would represent about 2 bp. Benzer thus learning objectives
inferred that a mutation can arise from the change of even
a single nucleotide pair, and that recombination can occur 1. Explain how the analysis of arginine auxotrophs implied
between adjacent nucleotide pairs. From the observation that a single gene corresponds to a single enzyme.
that mutations within the rII region form a self-consistent, 2. Describe how missense mutations were used to show
linear recombination map, he concluded that a gene is com- that genes determine the amino acid sequences of
posed of a continuous linear sequence of nucleotide pairs proteins.
within the DNA. And from observations that the positions 3. Differentiate between primary, secondary, tertiary, and
of mutations in the rIIA gene did not overlap those of the quaternary structures of proteins.
rIIB gene, he determined that the nucleotide sequences
composing those two genes are separate and distinct. A
gene is thus a linear set of nucleotide pairs, located within Mendel’s experiments established that an individual gene
a discrete region of a chromosome, that serves as a unit of can control a visible characteristic, but his laws do not ex-
function. plain how genes actually govern the appearance of traits.