APPLICATIONS OF RNA TECHNOLOGY IN TRANSGENICS
As discussed earlier, transgenic technology at the DNA level allows us to add genes and to inactivate gene expression by insertion. It is also possible to manipulate genes at the RNA level by the use of antisense, ribozymes, or RNA interference. We have already discussed these topics; therefore, this section simply summarizes their application in transgenics.
It is possible to insert an antisense transgene when constructing a transgenic organism. The simplest way to construct an antisense gene is to invert the DNA sequence of the original gene. In higher organisms, the cDNA version of the gene is used as the starting point to avoid complications due to intervening sequences. Such an antisense gene will be transcribed to give antisense RNA, which will then bind to the mRNA of the corresponding sense gene. This results in inhibition of gene expression at the level of translation.
Insertion of antisense genes into mice has resulted in up to 95% inhibition of gene expression, although the effectiveness varies greatly. The antisense RNA may be full length or just a short segment corresponding to part of the sense gene. Short segments complementary to the 5′-untranslated region of the sense mRNA are often very effective at blocking translation. Obviously, antisense constructs can be placed under the control of inducible promoters.
This allows antisense silencing of gene expression to be regulated by the experimenter.
It is also possible to insert transgenic constructs that express ribozymes. In such cases, the ribozyme is usually designed to cleave a specific mRNA. When the ribozyme is expressed it cleaves the mRNA of the target gene and so reduces gene expression. Occasional cases are known in both Drosophila and mice where transgenic expression of ribozymes has been successful in decreasing gene expression.
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