Allelic breeding- The Allelic complementation test

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A complementation test, which is also occasionally referred to as a “cis-trans” test is a simple diagnostic test for alleles that are recessive in nature. The test refers to an experiment that was developed by the American geneticist Edward B.Lewis, and answered Lewis’s question which was: "Does a wild-type copy of gene X rescue the function of the mutant allele that is believed to define gene X ?” So if there is a recessive allele with an observable phenotype whose gene function can be provided by a wild type genotype, will the function that was lost due to the recessive allele be provided by another mutant phenotype? If not, the two alleles must be defective in the same gene. The sheer beauty of this test is that it serves as a read out of gene function, even if you have no idea what the gene is doing at the molecular level.

What is complementation?

If we have two separate recessive colour gene strains of gerbils that are true breeding for their particular mutation, and these strains were crossed together, the offspring of these gerbils would normally inherit the wild type versions of each mutant gene from their parents if the recessive colour trait was situated on differing loci, so the wild-type phenotype (Golden Agouti) is recovered in the offspring. These strains are said to show “genetic complementation” because the mutations are recessive and as such there is a recovery of function in the pigment pathway which enables the recovery of the wild coat colour. So the test itself is used to find out if two true breeding recessive mutations are caused by mutations in the same gene or in two different genes. If both of the parent strains coat colours are caused by mutations in the same gene, no normal or wild-type versions of the gene can be inherited by the offspring; they still express a mutant phenotype and complementation has failed to occur. There are three possible reasons for this occurring:

  • The mutations occur in the same gene
  • One mutation effects the expression of the other
  • One mutation may result in an inhibitory product.

An example of a complementation test

Let’s take a hypothetical example and say we have three types of Blue gerbils (Strains a, b, c.). All three strains are true breeding for the blue colour, and all the strains display a similar mutation that causes a loss of function in the pigment pathway which gives rise to a blue coat. Firstly we need to cross to wild-type colour (Golden Agouti) to show that each mutant gene is recessive in nature. The test to show whether a particular mutation is due to a recessive gene is crucial, and allows you to move on to a complementation test. Dominant mutations cannot be used in complementation tests.

As you can see from the above demo, when each mutation is crossed to the wild-type colour, the gene behaves in a typical Mendelian recessive manner in both the F1 and F2 generations. In the F1 generation the wild-type gene from the Golden Agouti parent allows for the function to continue in the pigment pathway resulting in all the offspring having the wild-type coat colour. In each case of the above blue strains, the results show that the coat colour mutation is determined by the recessive allele of a single gene. However, are the mutations three alleles of the same locus, or are they situated on two or three differing loci? This question can be answered by asking if the mutations complement each other. So let’s quickly intercross our three Blue strains, a, b, and c.

From the above set of results we can conclude that blue strain ‘a’ and ‘b’ must be causes by the alleles of one gene because they do not complement, but blue strain ‘c’ must be caused by a recessive mutation of another gene.

Practical Examples

If we crossed a pure bred Black gerbil ‘aaCCDDEEGGPP’ with a pure bred Grey Agouti ‘AACCDDEEggPP’, we would expect all the offspring to be Golden Agouti or wild coloured. This is because the dominant wild genes have restored pigment function in the coat at the respective loci where each mutation has occurred. In this case the ‘A’ and ‘G’ locus. Because both of the mutations are recessive and are also situated on different loci, there is a recovery of function in the pigment pathway, so it is possible for the offspring to recover the wild coat colour. Complementation has occurred and the natural wild coat colour has been restored.

However, if we did a cross with a pure bred Dark-Eyed Honey ‘AACCDDeeGGPP’ and a pure bred Schimmel ‘AACCDDe(f)e(f)GGPP’ we wouldn't see any wild type or Golden Agouti offspring, and we still get the mutation appearing (In this case all the offspring are ee(f) ). So we can say that both mutations, ‘e’ and ‘e(f)’ occur on the same locus. Complementation has failed to occur and both mutations represent a loss of gene function in alleles that are situated on the same locus.