Text Assignment: Chapter 5, Pages 115-122.
Major concepts
Altered F2 phenotypic ratios: We have become quite familiar with the 9:3:3:1 phenotypic ratio that occurs in the F2 generation of a dihybrid cross in which there is independent assortment (absence of any detectable linkage) of alleles at the two genetic loci. The textbook gives substantial attention to the altered phenotypic ratio that occurs with linked loci. This is easiest to see with two recessive mutations that are from separate parental strains, such that each mutant allele is linked to a wild type allele at the other locus.
Nomenclature used: In the text, alleles at linked loci are shown with a line under them to show the linkage. For purposes of posting these notes in html, the alleles from two linked loci are written without a space between them. Thus, ab+ describes a recessive allele at locus a that is linked to a wild type allele at locus b. For two loci on the same autosome, a parental cross of ab+/ab+ x a+b/a+b will yield an F1 hybrid of the composition ab+/a+b, which will be a wild-type double heterozygote in a trans-configuration, that is with each of the recessive alleles a and b carried on a separate chromosome.
Linkage ratio: The F1 hybrid will produce two types of gametes ab+ and a+b. If there is complete linkage, the alleles at the two loci will stay strictly together, and the genotypes of F2 progeny will be 1 ab+/ab+ (mutant a phenotype), 2 ab+/a+b (wild-type) and 1 a+b/a+b (mutant b phenotype). Thus,the F2 will exhibit a 1:2:1 phenotypic ratio similar to that for a monohybrid cross with codominance. Although it is seldom encountered in real life anywhere other than in the progeny of male Drosophila, as described below, the textbook makes a point of referring to this ratio as a linkage ratio.
Crossing over: Because of crossing over during meiosis, which leads to genetic recombination, the linkage between non-allelic mutations on the same chromosome is nearly always less than 100%. However, crossing over does not ever occur on any of the chromosomes in male Drosophila. In addition, in many other species where some degree of crossing over does occur in both sexes, the relative frequencies of crossing over in males and females are often quite different (usually smaller in males and larger in females).
Linkage on the X chromosome: Because it became possible to identify genes carried on the X chromosome well before other chromosomal assignments were possible, much of the pioneering work on linkage was done with X-linked genes, as described in the text. Studies in the laboratory of Thomas Hunt Morgan rather quickly showed that these genes did not remain totally linked. Although each X-linked allele was transmitted from a heterozygous female to half of her male progeny, studies involving females that were heterozygous at two different X-linked loci revealed that alleles that had originally been on different X chromosomes could be transmitted to the same male offspring, although with a reduced frequency compared to either of the alleles alone. This led Morgan to propose that a genetic exchange between paired chromosomes was occurring at the chiasmata that could be seen during meiotic prophase. He also proposed that crossover would be relatively rare for genes that were close together and more frequent for genes further apart.
Undergraduate research: The next step was taken by Alfred H. Sturtevant, who at the time was an undergraduate student working in Morgan's laboratory. Sturtevant realized that crossover frequencies, which had been proposed by Morgan to be larger for genes that were further apart, might be used to construct a map of the relative postions of the genes on the chromosome. He found that distances were roughly additive for the first 3 genes he studied, and went on to construct a map with the relative positions of five different loci. This preliminary mapping, described in 1911, was the beginning of detailed chromosomal maps, such as those shown for Drosophila on page 132 of the textbook.
Mapping with test crosses: Although the textbook gives a lot of attention to altered F2 ratios resulting from linkage, experiments designed to detect linkage are usually done as test crosses. In these experiments, an organism that is heterozygous for the genetic loci whose linkage is being analyzed is crossed with an individual that is homozygous recessive for those genes. In most cases, the heterozygous individual is female because of higher crossover rates. (It is important to use the same sex consistently if results from various experiments are to be compared and used to construct maps). If there is no crossing over, all of the progeny of the test cross will have the same phenotypes as the true-breeding parents of the heterozygote. The amount of crossing over can be evaluated in terms of the fraction of progeny that exhibit recombinant phenotypes (which in a test cross accurately reflects the fraction of gametes with recombinant genotypes).
cis- and trans- configurations: In the simplest experiments, two genes that are known to be linked are examined to determine the amount of crossover that has occurred. (If needed, verification of linkage in Drosophila can be obtained by showing that the loci suspected of linkage stay strictly in the same relationship to each other in heterozygous males). The parental mutations that are being tested can be in the cis-configuration (both of the recessive mutations on the same chromosome, with both of the wild-type alleles on the homologous chromosome, or in the trans-configuration, with one recessive mutation on one chromosome (derived from one of the true-breeding parents) and the other on the homologous chromosome (devived from the other true-breeding parent).
Map distances: The probability of crossing over increases with physical distance between genes on the same chromosome (although not always strictly linearly). If recombination occurs, some of the progeny of the test cross will no longer exhibit the parental pattern. The percentage of recombinants detected in the progeny of the test cross increases with the distance between the genes, and is referred to as map distance. As an example, consider a test cross of a female of genotype AaBb that yields the following phenotypes: 495 AB, 495 ab, 5 Ab, and 5 aB. In a total of 1000 progeny, there are 5 Ab and 5aB phenotypes, which when added together represent 10 recombination events in 1000 gametes. This corresponds to a recombination frequency of 0.01, more commonly expressed as 1% crossing over.
Map units (centimorgans): A recombination frequency of 1% has historically been called one map unit since the time of the pioneering experiments of Sturtevant and Morgan. Although the term "map unit" is still widely used, one map unit is now often referred to as one centimorgan, honoring Thomas Hunt Morgan. One morgan corresponds to 100 map units, but this unit is too large for practical use, as discussed below.
Double crossovers: As the distance between two genes increases, the probability that two crossovers may occur between them begins to increase. If two crossovers occur, the parental genotype is restored, and the test is scored as if there had been no crossover. Thus, map distances measured in two-point tests become smaller than expected for genetic loci that are relatively distant from each other. In addition, because unlinked genes exhibit 50% recombination due to independent assortment, it becomes impossible to do a direct measurement of map distances for genes that are at large distances from one another.
Linkage to intermediate loci: In Drosophila, linkage can always be domonstrated in males, no matter how far apart the loci are, because of the total absence of crossing over. However, in species where crossing over occurs in both sexes, linkage of genetic loci that are quite far from each other on the same chromosome can only be demonstated by showing that both loci exhibit linkage to other loci located at intermediate positions along the chromosome. Total map distances on some large chromosomes can be well over 100 map units (Fig. 5.14, page 132).
Linkage groups: The term linkage group refers to all of the genes that are located on the same chromosome. Those that are close enough together will exhibit linkage in genetic crosses. However, the probability of crossover increases with the physical distance between genes on a chromosome, and genes that are located quite far from each other within a linkage group may not exhibit any detectable linkage in direct genetic tests. The term "linkage group" is also used to refer to genes that exhibit linked behavior in genetic systems where chromosomal assignments have not yet been made.
Additive distances: One way to determine map distance for genes that are located far from each other is to add the map distances between each of a series of pairs of loci positioned at reasonable distances along the chromosome. If three genes are oriented ABC on a chromosome, measured distance AB plus measured distance BC will normally be larger than directly measured distance AC. This is caused by double crossovers, as discussed above. The most accurate chromosomal maps are constructed by adding a series of relatively short map distances between more closely spaced alleles.
Three point crosses: In many cases, even after a gene
has been shown to be linked to a particular chromosome, its exact
placement on the chromosome will not be known. In the next lecture,
we will examine the three point cross, which has proven to be
a particularly useful tool, both for determining the order
of placement of genes on a chromosome, and for calculating
corrected map distances. The special value of the three point cross
lies in the use of relatively less common double crossover events to
identify the middle locus in any group of three genetic loci, and to
refine map distances obained by single crossover measurements.
Unfortunately, the three point cross is also one of the most difficult
concepts in genetics for a beginner to understand fully. You are strongly
advised to read the lecture 13 notes before coming to that lecture. .