Gene Cloning

The Production and Uses of Transgenic Organisms 375
gene is lowered, this function will no longer be effectively carried out, and the
effect on the phenotype of the organism can be observed). From an applied
perspective, this approach may be used to turn off the expression of a gene
which produces an undesirable trait in the organism.
What kind of DNA sequence could affect the expression of an endogenous
gene in a given plant or animal? One possible strategy to downregulate
an endogenous gene would be to produce, in all cells that express
the gene, an mRNA copy of the gene in the antisense orientation: in other
words, exactly complementary to the mRNA for the gene. This approach
has been tried in numerous different organisms, and although it is not
always successful, it does represent an important application of transgenic
organisms.
Although this technique is widely used, the way in which antisense
approaches work is not fully understood. Originally, it was thought that
mRNA complementary to a given message would bind to that message and
this would prevent ribosomes from binding to the message and translating
it into protein. Subsequent work has shown this is an over-simplified view
of what actually occurs, and in some cases at least antisense is effective in
lowering gene expression because cells contain specific RNA-degrading
enzymes that recognize and break down double-stranded (but not singlestranded)
RNA (Figure 12.7). Various modifications of the original principle
have been used, including the use of catalytic antisense RNAs (called
ribozymes) which not only bind to specific mRNAs but also cut them up,
and of course the RNAi approach discussed in Section 10.2.
Antisense technology has been widely used, and is particularly celebrated
because it was used in developing the first ever GM food to be
Endogenous gene produces sense
mRNA encoding protein
GENE
Pairing of sense and antisense
mRNAs generates a ds RNA
which is not translated and
is rapidly degraded
ENEG
Transgene produces antisense
mRNA which is complementary
to that of endogenous gene
Figure 12.7 Antisense RNA as a method to reduce gene expression.
Pairing of sense and antisense mRNA leads to a double-stranded RNA which is
degraded. This reduces or prevents translation of the sense mRNA.