Techniques : Roslin

The Details:Techniques:
The Roslin Technique

Timeline: 1995
Timeline: 1996
Timeline: 1997
Honolulu Technique
Nuclear Transfers

Scientist Profiles
Ian Wilmut
Keith Campbell

Web Link:
The Roslin Institute Online
Text of Wilmut's Research Paper in Nature

The cloning of Dolly has been the most important event in cloning history. Not only did it spark public interest in the subject, but it also proved that the cloning of adult animals could be accomplished. Previously, it was not known if an adult nucleus was still able to produce a completely new animal. Genetic damage and the simple deactivation of genes in cells were both considered possibly irreversible.

The realization that this was not the case came after the discovery by Ian Wilmut and Keith Cambell of a method with which to synchronize the cell cycles of the donor cell and the egg cell. Without synchronized cell cycles, the nucleus would not be in the correct state for the embryo to accept it. Somehow the donor cell had to be forced into the Gap Zero, or G0 cell stage, or the dormant cell stage.

First, a cell (the donor cell) was selected from the udder cells of a Finn Dorset sheep to provide the genetic information for the clone. For this experiment, the researchers allowed the cell to divide and form a culture in vitro, or outside of an animal. This produced multiple copies of the same nucleus. This step only becomes useful when the DNA is altered, such as in the case of Polly, because then the changes can be studied to make sure that they have taken effect.

A donor cell was taken from the culture and then starved in a mixture which had only enough nutrients to keep the cell alive. This caused the cell to begin shutting down all active genes and enter the G0 stage. The egg cell of a Blackface ewe was then enucleated and placed next to the donor cell. One to eight hours after the removal of the egg cell, an electric pulse was used to fuse the two cells together and, at the same time, activate the development of an embryo. This technique for mimicking the activation provided by sperm is not completely correct, since only a few electrically activated cells survive long enough to produce an embryo.

If the embryo survives, it is allowed to grow for about six days, incubating in a sheep's oviduct. It has been found that cells placed in oviducts early in their development are much more likely to survive than those incubated in the lab. Finally, the embryo is placed into the uterus of a surrogate mother ewe. That ewe then carries the clone until it is ready to give birth. Assuming nothing goes wrong, an exact copy of the donor animal is born.

This newborn sheep has all of the same characteristics of a normal newborn sheep. It has yet to be seen if any adverse effects, such as a higher risk of cancer or other genetic diseases that occur with the gradual damage to DNA over time, are present in Dolly or other animals cloned with this method.

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