WordNet

(genetics) a segment of DNA that is involved in producing a polypeptide chain; it can include regions preceding and following the coding DNA as well as introns between the exons; it is considered a unit of heredity; "genes were formerly called factors" (同)cistron, factor
a blow that renders the opponent unconscious (同)KO, kayo
informal term for information; "give me the gen on your new line of computers"

PrepTutorEJDIC

遺伝子
(ボクシングの)ノックアウト(《略》『KO,K.O.,k.o.』) / 《話》すばらしい人(物) / ノックアウトする

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出典(authority):フリー百科事典『ウィキペディア（Wikipedia）』「2014/03/15 00:43:42」(JST)

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A gene knockout (abbreviation: KO) is a genetic technique in which one of an organism's genes are made inoperative ("knocked out" of the organism). Also known as knockout organisms or simply knockouts, they are used in learning about a gene that has been sequenced, but which has an unknown or incompletely known function. Researchers draw inferences from the difference between the knockout organism and normal individuals.

The term also refers to the process of creating such an organism, as in "knocking out" a gene. The technique is essentially the opposite of a gene knockin. Knocking out two genes simultaneously in an organism is known as a double knockout (DKO). Similarly the terms triple knockout (TKO) and quadruple knockouts (QKO) are used to describe three or four knocked out genes, respectively.

Method[edit]

A laboratory mouse in which a gene affecting hair growth has been knocked out (left), is shown next to a normal lab mouse.

Knockout is accomplished through a combination of techniques, beginning in the test tube with a plasmid, a bacterial artificial chromosome or other DNA construct, and proceeding to cell culture. Individual cells are genetically transfected with the DNA construct. Often the goal is to create a transgenic animal that has the altered gene. If so, embryonic stem cells are genetically transformed and inserted into early embryos. Resulting animals with the genetic change in their germline cells can then often pass the gene knockout to future generations.

To create knockout moss, transfection of protoplasts is the preferred method. Such transformed Physcomitrella-protoplasts directly regenerate into fertile moss plants. Already eight weeks after transfection the plants can be screened for gene targeting via PCR.^[1]

Wild-type Physcomitrella and knockout mosses: Deviating phenotypes induced in gene-disruption library transformants. Physcomitrella wild-type and transformed plants were grown on minimal Knop medium to induce differentiation and development of gametophores. For each plant, an overview (upper row; scale bar corresponds to 1 mm) and a close-up (bottom row; scale bar equals 0.5 mm) are shown. A: Haploid wild-type moss plant completely covered with leafy gametophores and close-up of wild-type leaf. B–D: Different mutants.^[2]

The construct is engineered to recombine with the target gene, which is accomplished by incorporating sequences from the gene itself into the construct. Recombination then occurs in the region of that sequence within the gene, resulting in the insertion of a foreign sequence to disrupt the gene. With its sequence interrupted, the altered gene in most cases will be translated into a nonfunctional protein, if it is translated at all.

A knockout mouse (left) that is a model of obesity, compared with a normal mouse.

A conditional knockout allows gene deletion in a tissue or time specific manner. This is done by introducing short sequences called loxP sites around the gene. These sequences will be introduced into the germ-line via the same mechanism as a knock-out. This germ-line can then be crossed to another germline containing Cre-recombinase which is a viral enzyme that can recognize these sequences, recombines them and deletes the gene flanked by these sites.

Because the desired type of DNA recombination is a rare event in the case of most cells and most constructs, the foreign sequence chosen for insertion usually includes a reporter. This enables easy selection of cells or individuals in which knockout was successful. Sometimes the DNA construct inserts into a chromosome without the desired homologous recombination with the target gene. To eliminate such cells, the DNA construct often contains a second region of DNA that allows such cells to be identified and discarded.

In diploid organisms, which contain two alleles for most genes, and may as well contain several related genes that collaborate in the same role, additional rounds of transformation and selection are performed until every targeted gene is knocked out. Selective breeding may be required to produce homozygous knockout animals.

Knock-in is similar to knock-out, but instead it replaces a gene with another instead of deleting it.

Use[edit]

Knockouts are primarily used to understand the role of a specific gene or DNA region by comparing the knockout organism to a wildtype with a similar genetic background.

Knockouts organisms are also used as screening tools in the development of drugs, to target specific biological processes or deficiencies by using a specific knockout, or to understand the mechanism of action of a drug by using a library of knockout organisms spanning the entire genome, such as in Saccharomyces cerevisiae.^[3]

References[edit]

^ Ralf Reski (1998): Physcomitrella and Arabidopsis: the David and Goliath of reverse genetics. Trends Plant in Science 3, 209-210
^ Egener et al. BMC Plant Biology 2002 2:6 doi:10.1186/1471-2229-2-6
^ http://www-sequence.stanford.edu/group/yeast_deletion_project/deletions3.html

External links[edit]

Diagram of targeted gene replacement
Frontiers in Bioscience Gene Knockout Database (available on archive only)
International Knockout Mouse Consortium
KOMP Repository

UpToDate Contents

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English Journal

Reduction of biogenic amines production by eliminating the PEP4 gene in Saccharomyces cerevisiae during fermentation of Chinese rice wine.

Guo X1, Guan X1, Wang Y1, Li L1, Wu D1, Chen Y1, Pei H1, Xiao D2.
Food chemistry.Food Chem.2015 Jul 1;178:208-11. doi: 10.1016/j.foodchem.2015.01.089. Epub 2015 Jan 24.
Biogenic amines in Chinese rice wine have a potential threat of toxicity to human health. In this study, PEP4 gene in Saccharomyces cerevisiae was knocked out in order to evaluate its effect on biogenic amines production; the enzyme encodes proteinase A (PrA), an enzyme that is responsible for the p
PMID 25704703

Effects of glucose-dependent insulinotropic polypeptide receptor knockout and a high-fat diet on cognitive function and hippocampal gene expression in mice.

Lennox RR1, Moffett C1, Porter DW1, Irwin N1, Gault VA1, Flatt PR1.
Molecular medicine reports.Mol Med Rep.2015 Jul;12(1):1544-8. doi: 10.3892/mmr.2015.3447. Epub 2015 Mar 6.
It has been previously demonstrated that compromise of glucose‑dependent insulinotropic polypeptide receptor (GIPR) action and chronic consumption of a high‑fat diet can independently impair memory and learning ability, however, the underlying pathology remain to be elucidated. The present study
PMID 25760229

HIF1α is necessary for exercise-induced neuroprotection while HIF2α is needed for dopaminergic neuron survival in the substantia nigra pars compacta.

Smeyne M1, Sladen P1, Jiao Y1, Dragatsis I2, Smeyne RJ3.
Neuroscience.Neuroscience.2015 Jun 4;295:23-38. doi: 10.1016/j.neuroscience.2015.03.015. Epub 2015 Mar 19.
Exercise reduces the risk of developing a number of neurological disorders and increases the efficiency of cellular energy production. However, overly strenuous exercise produces oxidative stress. Proper oxygenation is crucial for the health of all tissues, and tight regulation of cellular oxygen is
PMID 25796140

Japanese Journal

Generation of albino Xenopus tropicalis using zinc-finger nucleases

Nakajima Keisuke,Nakajima Taeko,Takase Minoru [他]
Development, growth & differentiation 54(9), 777-784, 2012-12-00
NAID 40019526018

Social memory, amnesia, and autism: Brain oxytocin secretion is regulated by NAD + metabolites and single nucleotide polymorphisms of CD38

Higashida Haruhiro,Yokoyama Shigeru,Huang Jian-Jun,Liu Li,Ma Wen-Jie,Akther Shirin,Higashida Chiharu,Kikuchi Mitsuru,Minabe Yoshio,Munesue Toshio
Neurochemistry International 61(6), 828-838, 2012-11-00
… When CD38 was disrupted, social amnesia was observed in Cd38 knockout mice. … Hence, there is a rationale for investigating single nucleotide polymorphisms (SNPs) in the human CD38 gene in ASD subjects. … We found several SNPs in this gene. … A) was associated with high-functioning autism (HFA) in American samples from the Autism Gene Resource Exchange. …
NAID 120004966608

ビタミンE特異的輸送タンパク質と先天性ビタミンE欠乏症の解明

新井洋由
ビタミン 86(10), 529-538, 2012-10-25
… Moreover, we demonstrated that the α-TTP gene is mutated in the patients with familial vitamin E deficiency. … α-TTP knockout mice were also established and served as a good model for human patients with familial vitamin E deficiency. …
NAID 110009544359