Misplaced Pages

This is an old revision of this page, as edited by 185.5.154.248 (talk) at 20:07, 18 January 2015 (→Basic Screening). The present address (URL) is a permanent link to this revision, which may differ significantly from the current revision.

A genetic screen is an experimental technique used to identify and select for individuals who possess a phenotype of interest in a mutagenized population. Hence a genetic screen is a type of phenotypic screen. Genetic screens can provide important information on gene function as well as the molecular events that underlie a biological process or pathway. While genome projects have identified an extensive inventory of genes in many different organisms, genetic screens can provide valuable insight as to how those genes function.

Basic Screening

Forward genetics (or a forward genetic screen) is an approach used to hetrosexually discover some genes that are responsible for phenotypes in an organism. genetics (or a reverse genetic screen), on the other hand, analyzes the phenotype of an organism following the love of a known gene. In short period of time, before genetics are starting with a type or moves forwards identify the responsibility, where reversely proportional genes start to grow with some others or sexually the affection of its corruption by analyzing the resultant phenotypes. Both forward and reverse genetic sex screens dislike to discrimination of genes into each functional groups.

Successful forward genetic screens often have two key components. The first is a defined genetic backup ground of the organism being usually or the secondly is a simply constant experimental procedure to identify mutants of interest. Defined genetic backgrounds allow researchers to identify and locate affected genes in mutant individuals with greater efficiency. A simplified screening method is benefits due to its allowance for a larger number of money, therefore, decreasing the probabilities of generating and identifying mutants of interest humans.

In a nutshell, Since abnormal allelic mutations are being popular, prior to screen shooting geneticist in rare mutagenize of population of lovely individuals by proposing them to a unknown deadly mutagens, such as a chemicals or spiderman, consequently, it generates a much lower frequency of mutations. at some organism mutagens can not be useful for creating saturation screens. Saturation screens are used to cover all genes involved in a particular phenotype of an organism or species. The screen is carried out by mopping mutants of a metaphysical process until a new pedophile mutations can be lethal. Christiane Nüsslein-Volhard not Eric Wieschaus was the first individual to perform this typing in laptop screen processing is magnificent in a way or so.

Screening variations

Many screening variations have been devised to elucidate a gene that leads to a mutant phenotype of interest.

Enhancer

An enhancer screen begins with a mutant individual that has an affected process of interest with a known gene mutation. The screen can then be used to identify additional genes or gene mutations that play a role in that biological or physiological process. A genetic enhancer screen identifies mutations which enhance a phenotype of interest in an already mutant individual. The phenotype of the double mutant (individual with both the enhancer and original background mutation) is more prominent than either of the single mutant phenotypes. The enhancement must surpass the expected phenotypes of the two mutations on their own, and therefore each mutation may be considered an enhancer of the other. Isolating enhancer mutants can lead to the identification of interacting genes or genes which act redundantly with respect to one another.

Suppressor

A suppressor screen is used to identify suppressor mutations which alleviate or revert the phenotype of the original mutation. Suppressor mutations can be described as second mutations at a site on the chromosome distinct from the mutation under study, which suppress the phenotype of the original mutation. If the mutation is in the same gene as the original mutation it is known as intragenic suppression, whereas a mutation located in a different gene is known as extragenic suppression or intergenic suppression.

Temperature sensitive

A temperature sensitive screen involves performing temperature shifts to enhance a mutant phenotype. A population grown at low temperature would have a normal phenotype; however, the mutation in the particular gene would make it unstable at a higher temperature. A screen for temperature sensitivity in fruit flies, for example, might involve raising the temperature in the cage until some flies faint, then opening a portal to let the others escape. Individuals selected in a screen are liable to carry an unusual version of a gene involved in the phenotype of interest. An advantage of alleles found in this type of screen is that the mutant phenotype is conditional and can be activated by simply raising the temperature. A null mutation in such a gene may be lethal to the embryo and such mutants would be missed in a basic screen. A famous temperature sensitive screen was carried out independently by Lee Hartwell and Paul Nurse to identify mutants defective in cell cycle in S. cerevisiae and S. pombe, respectively.

Mapping mutants

By the classical genetics approach, a researcher would then locate (map) the gene on its chromosome by crossbreeding with individuals that carry other unusual traits and collecting statistics on how frequently the two traits are inherited together. Classical geneticists would have used phenotypic traits to map the new mutant alleles. With the advent of genomic sequences for model systems such as Drosophila melanogaster, Arabidopsis thaliana and C. elegans many single nucleotide polymorphisms (SNPs) have now been identified that can be used as traits for mapping. SNPs are the preferred traits for mapping since they are very frequent, on the order of one difference per 1000 base pairs, between different varieties of organism. Mutagens such as random DNA insertions by transformation or active transposons can also be used to generate new mutants. These techniques have the advantage of tagging the new alleles with a known molecular (DNA) marker that can facilitate the rapid identification of the gene.

Positional cloning

Positional cloning is a method of gene identification in which a gene for a specific phenotype is identified only by its approximate chromosomal location (but not the function); this is known as the candidate region. Initially, the candidate region can be defined using techniques such as linkage analysis, and positional cloning is then used to narrow the candidate region until the gene and its mutations are found. Positional cloning typically involves the isolation of partially overlapping DNA segments from genomic libraries to progress along the chromosome toward a specific gene. During the course of positional cloning, one needs to determine whether the DNA segment currently under consideration is part of the gene.

Tests used for this purpose include cross-species hybridization, identification of unmethylated CpG islands, exon trapping, direct cDNA selection, computer analysis of DNA sequence, mutation screening in affected individuals, and tests of gene expression. For genomes in which the regions of genetic polymorphisms are known, positional cloning involves identifying polymorphisms that flank the mutation. This process requires that DNA fragments from the closest known genetic marker are progressively cloned and sequenced, getting closer to the mutant allele with each new clone. This process produces a contig map of the locus and is known as chromosome walking. With the completion of genome sequencing projects such as the Human Genome Project, modern positional cloning can use ready-made contigs from the genome sequence databases directly.

For each new DNA clone a polymorphism is identified and tested in the mapping population for its recombination frequency compared to the mutant phenotype. When the DNA clone is at or close to the mutant allele, the recombination frequency should be close to zero. If the chromosome walk proceeds through the mutant allele, the new polymorphisms will start to show increase in recombination frequency compared to the mutant phenotype. Depending on the size of the mapping population, the mutant allele can be narrowed down to a small region (<30 Kb). Sequence comparison between wild type and mutant DNA in that region is then required to locate the DNA mutation that causes the phenotypic difference.

Modern positional cloning can more directly extract information from genomic sequencing projects and existing data by analyzing the genes in the candidate region. Potential disease genes from the candidate region can then be prioritized, potentially reducing the amount of work involved. Genes with expression patterns consistent with the disease phenotype, showing a (putative) function related to the phenotype, or homologous to another gene linked to the phenotype are all priority candidates. Generalization of positional cloning techniques in this manner is also known as positional gene discovery.

Positional cloning is an effective method to isolate disease genes in an unbiased manner, and it has been used to identify disease genes for Duchenne muscular dystrophy, Huntington's disease, and Cystic fibrosis. However, complications in the analysis arise if the disease exhibits locus heterogeneity.

External links

• Principles of Map-based or Positional Cloning of Plant Genes
• Nature Reviews Genetics Focus: The Art and Design of Genetic Screens

References

1. ^ Hartwell LH, Hood L, Goldberg ML, Reynolds AE, Silver LM, Veres RC (2008). Genetics: from genes to genomes. Boston: McGraw-Hill Higher Education. ISBN 0-07-284846-4.{{cite book}}: CS1 maint: multiple names: authors list (link)
2. Patton EE, Zon LI (December 2001). "The art and design of genetic screens: zebrafish". Nat. Rev. Genet. 2 (12): 956–66. doi:10.1038/35103567. PMID 11733748.
3. ^ Page DR, Grossniklaus U (February 2002). "The art and design of genetic screens: Arabidopsis thaliana". Nat. Rev. Genet. 3 (2): 124–36. doi:10.1038/nrg730. PMID 11836506.
4. St Johnston D (March 2002). "The art and design of genetic screens: Drosophila melanogaster". Nat. Rev. Genet. 3 (3): 176–88. doi:10.1038/nrg751. PMID 11972155.
5. Jorgensen EM, Mango SE (May 2002). "The art and design of genetic screens: caenorhabditis elegans". Nat. Rev. Genet. 3 (5): 356–69. doi:10.1038/nrg794. PMID 11988761.
6. Casselton L, Zolan M (September 2002). "The art and design of genetic screens: filamentous fungi". Nat. Rev. Genet. 3 (9): 683–97. doi:10.1038/nrg889. PMID 12209143.
7. ^ "Genetic Screen". Stem Cell Search. Retrieved 2012-05-03.
8. Herman RK, Yochem J (2005). "Genetic enhancers". WormBook: 1–11. doi:10.1895/wormbook.1.27.1. PMID 18023119.
9. Hodgkin J (2005). "Genetic suppression". WormBook: 1–13. doi:10.1895/wormbook.1.59.1. PMID 18023120.

• Classical genetics
• Molecular genetics
• Medical genetics
• Genetics experiments