As we have seen earlier, if two genes are located on different chromosomes, phenotypic traits associated with them assort independently, whereas if they reside on different copies of the same chromosome the coincidence of both traits in the same individual occurs less frequently than expected. In fact, in the absence of recombination, traits specified by two genes on different homologs will never segregate together. The degree to which two traits do co-segregate therefore tells us something about the occurrence of recombination between the respective genes and it is this property that allows us to map genes relative to one another along the chromosome. This is because the frequency of recombination that occurs between them during meiosis, is directly related to the physical distance separating the two genes on the chromosome. The greater the distance between the genes, the greater the probability that a chiasma will form between them. Thus calculating the frequency of recombination between genes allows the genetic map position along the chromosome to be calculated.

Breeding experiments performed to determine the location of genes are known as mapping crosses and usually involve at least two markers on the same chromosome as the gene to be mapped. The example shown in Figure 6-2.1 illustrates the mapping of the Antp mutation relative to Sb and to another dominant marker on the same chromosome, Lyra (Ly; thin wings). First females heterozygous for the mutant-bearing chromosome and the marker-bearing chromosome are generated. Because Antp has been inherited from one parent and Sb and Ly from the other in this female, the Antp mutation is on a different copy of the homologous chromosome that carries Sb and Ly and is said to be in trans or in repulsion to these markers. The Sb and Ly mutations, by contrast, are on the same homolog and are said to be in cis or in coupling. Meiotic crossing over takes place in female Drosophila, so recombination will have occurred within the vicinity of these genes in a proportion of meioses in this fly, giving rise to recombinant chromosomes in which Antp is now linked to one of the markers. We can detect these recombination events by mating the female with a wild-type male and examining the phenotype of the progeny.

The majority of the progeny of the cross have legs instead of antennae with normal wings and bristles or have normal antennae but shortened bristles and thin wings: each of these has inherited one or other of the parental chromosomes and they thus represent the so-called non-recombinant progeny. Flies that have inherited recombined forms of the original chromosomes, by contrast, manifest different combinations of the three mutant traits. Recombination between Ly and Antp gives rise to flies that are phenotypically antennapedia with thin wings and others which have short bristles, whereas recombination between Antp and Sb gives rise to flies with legs instead of antennae and short bristles and others with just thin wings. In addition, a small proportion of the progeny will inherit chromosomes that have undergone recombination between Antp and both markers; these will either be phenotypically wild type or will manifest all three mutant traits. To map the position of Antp, we first count the number of recombinant flies of each type and then add together those generated by recombination between the same pairs of genes. Thus 18/200 flies have either legs in place of antennae and short bristles with normal wings, or wild-type antennae and bristles with thin wings, reflecting recombination between Antp and Sb; while 14/200 have legs in place of antennae and thin wings with normal bristles, or normal wings and antennae with short bristles, reflecting recombination between Antp and Ly. The larger number of recombinants between Antp and Sb than between Antp and Ly reflects the fact that Antp is further from Sb than it is from Ly. To obtain the total number of recombinations between Antp and each of the markers, we must also add the rare double recombinants: there are two of these, giving 18+2 = 20 recombinants between Antp and Sb and 14+2 = 16 recombinants between Antp and Ly. To convert these numbers into standardized map distances, we express them as a percentage of the total numbers of flies in the cross, to generate the distance in map units known as centiMorgans (cMs) after the geneticist Thomas Hunt Morgan who worked out how to map genes. One cM is thus equivalent to the distance between two loci that show 1% recombination. In our experiment we can conclude that Antp is (14+2)/200 = 8 cMs from Ly and (18+2)/200 = 10 cMs from Sb. The order of the genes along the chromosome can be deduced by calculating the map distance between Sb and Ly. In our example, 18+14 flies show non-parental inheritance of these two markers, giving a map distance of 32/200 or 16 cMs between them. Since Antp is 8 cMs from Ly and 10 from Sb, it follows that it must lie between the two markers. Significantly, the distance between Ly and Sb calculated by summing the distance of each from Antp (10+8 = 18) is greater than the calculated value of 16 cMs. This is because the double crossovers - detected when scoring flies for all three markers - are not included when the segregation of only Ly and Sb is considered. This is an important factor to consider when constructing genetic maps: the more markers that are used, the more accurate the map.

In organisms such as Drosophila, with its large number of marker mutations that manifest visible phenotypes, genes can be mapped simply by visual inspection of the progeny of mapping crosses. A more generally applicable method of mapping, essential in organisms that lack this wealth of visible marker mutations, exploits the existence of molecular polymorphisms between strains. Such polymorphisms are randomly distributed throughout genomes and can be used as molecular markers to construct genetic maps.

Even in Drosophila, single nucleotide polymorphisms (SNPs) are now commonly used for mapping. Comparisons of genomic sequences between different strains reveal SNPs at a relatively high frequency (around 1/200 base pairs between different strains of Drosophila). In many cases these abolish recognition sites for restriction enzymes and thus give rise to restriction fragment length polymorphisms (RFLPs) that are readily detected by restriction digestion of the relevant PCR amplified fragment. Alternatively, SNPs can be detected by direct sequencing, mass spectrophotometry or single stranded conformation polymorphism (SSCP) analysis (see Chapter 2). Mutations that result in a particular phenotypic trait - be it antennal transformation in Drosophila or a human disease model in fish or mouse - can be mapped relative to such molecular markers in exactly the same way that a trait is mapped using visible markers, as illustrated in Figure 6-2.2, and explained in the legend.

centiMorgan: the unit of genetic map distance between two loci that show 1% recombination.

frequency of recombination: the number of cross over events observed between two linked loci expressed as a proportion of the total number of meioses sampled.

genetic map position: the location of a gene on the genetic map deduced from recombination frequencies.

mapping crosses: crosses in which recombination between molecular or visible markers and a gene of interest is assayed to calculate recombination frequencies.