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A conditional gene knockout allows gene deletion in a tissue in a tissue specific manner. This is required in place of a gene knockout if the null mutation would lead to embryonic death, [13] or a specific tissue or cell type is of specific interest. This is done by introducing short sequences called loxP sites around the gene.
Gene knockdown is an experimental technique by which the expression of one or more of an organism's genes is reduced. The reduction can occur either through genetic modification or by treatment with a reagent such as a short DNA or RNA oligonucleotide that has a sequence complementary to either gene or an mRNA transcript.
Floxing a gene allows it to be deleted (knocked out), [5] [6] translocated or inserted [7] (through various mechanisms in Cre-Lox recombination). The floxing of genes is essential in the development of scientific model systems as it allows spatial and temporal alteration of gene expression.
Gene silencing is often considered the same as gene knockdown. [3] [4] When genes are silenced, their expression is reduced. [3] [4] In contrast, when genes are knocked out, they are completely erased from the organism's genome and, thus, have no expression.
Using the conditional gene knockout technique eliminates many of the side effects from traditional gene knockout. In traditional gene knockout, embryonic death from a gene mutation can occur, and this prevents scientists from studying the gene in adults. Some tissues cannot be studied properly in isolation, so the gene must be inactive in a ...
Gene knock-in originated as a slight modification of the original knockout technique developed by Martin Evans, Oliver Smithies, and Mario Capecchi.Traditionally, knock-in techniques have relied on homologous recombination to drive targeted gene replacement, although other methods using a transposon-mediated system to insert the target gene have been developed. [3]
Targeted gene knockout using CRISPR/Cas9 requires the use of a delivery system to introduce the sgRNA and Cas9 into the cell. Although a number of different delivery systems are potentially available for CRISPR, [37] [38] genome-wide loss-of-function screens are predominantly carried out using third generation lentiviral vectors.
For example, deletion of a gene by gene targeting (gene knockout) can be done in some organisms, such as yeast, mice and moss. Unique among plants, in Physcomitrella patens , gene knockout via homologous recombination to create knockout moss (see figure) is nearly as efficient as in yeast. [ 4 ]