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What is meant by forward genetics

Forward genetics is the method by which you identify the function of a gene and then identify the gene itself


How are forward genetic screen carried out

An organism is taken and its genome is randomly mutated using a mutagen. Then, interesting phenotypes are looked for in the offspring of the mutagenized animals. Once the interesting phenotypes have been identified the next stage is to identify the actual gene that the mutation is in


What chemical mutagen is commonly used in forward genetic screens



Why are more simple organisms used preferentially over more complex mammalian systems in forward genetic screens

Because the mutagenesis process occurs at random and effects the entire genome the probability if hitting a specific gene is relatively low. This increases the number of mutagenized animals you would need to analyse to find the interesting phenotypes. Thus simpler organisms are used to speed up this process


What examples of simple systems are used in forward genetic screens

Drosophila, C. elegans, Zebrafish and Yeast


Most genetic screens are done for recessive alleles but domain forward screens can also be done, T or F



What technique allows you to identify the specific gene mutated in a particular phenotype

Positional cloning/linkage analysis


Explain how a forward genetic screen is carried out in flies

Male flies are exposed to a mutagen such as EMS which is then integrated into the body and introduces mutations in the sperm. This generation of males, referred to a P0 are then outcrossed with non-mutagenised wild type female flies. The male offspring from this cross are then collected, these are referred to as the F1 generation. Each male in the F1 will have 4 or 5 mutated genes. These males are then crossed with wild type females once again to produce the F2 generation. The F2 generation will contain both males and females with mutant copies of the genes. This family is then incrossed to see what the homozygous mutants look like.


What percentage of the F2 generation of flies produced by a forward genetic screen will show a phenotype if there is one

A quarter


How many generations does it take to make a particular mutation homozygous

3 generations; Po, F1 and F2


To identify genes that affect a specific process, thousands of F2 families need to be screened, T or F



What is meant by complementation testing

Complementation testing is used to determine if different mutations that have the same phenotype are alleles of the same gene or if these mutations in fact lie in separate genes.


Explain the process of complementation testing

Complementation analysis allows the sorting of mutations into distinct groups that correspond to individual genes. Carriers of a mutation that has the same phenotype are crossed and the offspring produced from this cross are analysed. If ¼ of the offspring produced are homozygous mutants then the mutations fail to complement each other and they are mutant alleles of the same gene. If no offspring with the mutant phenotype are produced then mutations are said to complement each other and lie in different genes


What is the name given to the groups of separate genes that when mutated produced the same phenotype

Complementation groups


Explain how linkage analysis allows the identification of a particular gene

Linkage analysis involves analysing recombination between an identified allele and a known marker on the same chromosome. This enables the determination of whether the gene and marker are linked in any way. Carriers of the mutation are crossed with carriers of the marker and the number of offspring where crossing over has occurred in, are counted. These are the number of offspring that exhibit both the mutation and the marker. The greater the distance between the gene and the marker on the chromosome the more frequent that crossing over events will occur and the greater the number of offspring produced with both attributes.


What is indicated by linkage analysis experiments that reveal few offspring that show both the mutation and the marker

It means that the mutation and the marker lie close to each other and hence crossing over is rare


What maker is often used in fruit fly screens



Recall the equation that calculates the genetic distance between mutation and marker in linkage analyses

cM = R/T x 100 (genetic distance = number of recombinant gametes/total number of gametes)


How are genetic distances calculated in humans

A correlation between known SNPs and diseased phenotypes are investigated. SNPs act as the markers and over 1m have been identified in humans. The offspring of a diseased individual are analysed to determine if particularly SNPs are always present in the diseased state. If an SNP is always present in diseased offspring but not healthy ones you can determine that the mutant gene is linked to that SNP. You can then correlate the location of the known SNP with known genes to derived candidate genes.


What are SNPs

Single polynucleotide polymorphisms are single base pair changes that are frequent in the population, varying from individual to individual but without conferring a disease phenotype


What is the result of changes in the regulatory sequences of the DNA

These affect transcription and occur in regulatory sequences such as enhancers


What are the downstream implications of changes in the non-coding sequence of the transcript of a gene

These may effect RNA splicing, stability or folding


Give an example of a change in the non-coding sequence of a gene transcript

Mutations that lead to the deletion of an intron


What are the results of mutations in the coding sequence of a gene

May alter amino acid composition which can impact folding


What are the main two types of mutations in the coding sequence

Missense mutations which are characterised by the substitution of an amino acid. Nonsense mutations create premature stop codons which result in a truncated protein


Below is a table containing a list of features of loss of function mutations, determine if they are characteristic of amorphic, hypomorphic or antimorphic mutations?




Hypomorphic and amorphic









What is meant by a hypermorphic mutation

Mutation that confers a gain of function of the gene product. This is caused by the overexpression of a transcription unit or overactivity of a gene production.


Are hypermorphic mutations recessive or dominant