A future for transgenic livestock

Abstract

The techniques that are used to generate transgenic livestock are inefficient and expensive. This, coupled with the fact that most agriculturally relevant traits are complex and controlled by more than one gene, has restricted the use of transgenic technology. New methods for modifying the genome will underpin a resurgence of research using transgenic livestock. This will not only increase our understanding of basic biology in commercial species, but might also lead to the generation of animals that are more resistant to infectious disease.

Figure 1. Different routes for germline modification.

a | Pro-nuclear injection involves the introduction of the DNA construct into the fertilized egg, which is then transferred to a recipient female^,,,. Only a small proportion of the injected eggs will yield a transgenic founder animal, which is usually identified by Southern blotting after birth. b | Embryonic stem (ES) cells are only available in mice and, so far, this technology is limited to this species. DNA manipulation occurs in the ES cells before embryo manipulation and might involve random gene addition or gene targeting. The modified ES cells, which are identified by Southern blotting, are injected into a host blastocyst that will develop to form a chimaera that consists of both host and ES cells. Only chimeric mice in which the germline has arisen from the modified ES cells can become the founder of a transgenic line. c | Nuclear transfer^, from cultured cells has been achieved in several livestock species including sheep, pigs, cattle, goats, mules and horses, as well as mice and rabbits. The genetic modifications are carried out in the cultured cells before nuclear transfer. Nuclei from the modified cells are transferred to an enucleated oocyte by cell fusion before their development in recipient animals. The process can yield several identical transgenic clones. This technology has been used to add new DNA sequences and for gene-targeting strategies^,,,,,. Modified with permission from Ref. © (2000) Kluwer Academic.

Timeline | Landmark events in transgenic livestock research

Figure 2. Targeted gene deletion in sheep.

Nuclear transfer from a primary sheep fibroblast, in which one copy of the prion protein gene (PrP) had been disrupted through gene targeting by homologous recombination, generated the lamb shown.

Figure 3. Generation of transgenic sheep using lentivectors.

Recombinant lentivector components (a), including a small interfering (si)RNA construct, are transfected into a packaging cell line and the packaged viral particles (b) are purified by centrifuging the cell supernatant. The efficiency of interference with viral replication/transcription can be tested in cell culture (c) before injecting the particles into the perivitelline space of a fertilized egg (d). After in vitro culture the infected early stage embryos are transferred into recipients. After gestation, founder transgenic animals are born (e) and they can be tested for resistance.