By David N. Leff

BOSTON — Seven healthy Holstein bull calves took a bow at the Massachusetts Biotechnology Council's seventh annual Biotechnology and Trade Exposition here.

The animals' co-creator, developmental biologist Steve Stice, introduced the micro-herd to attendees at a session this week called "Whole organism: Knock in/knock out/transfer technology." Stice is chief scientific officer of Advanced Cell Technology Inc. (ACT), in Shrewsbury, Mass., and co-author of a paper in today's Science, dated May 22, 1998, titled: "Cloned transgenic calves produced from nonquiescent fetal fibroblasts." (See related story, " Transgenic Replication Gets Boost From Fibroblast Cells," in today's issue.)

The article's senior author, James Robl, professor of reproductive and developmental biology at the University of Massachusetts, in Amherst, chaired the session at which Stice reported on "Cloning cattle and pigs for use in biomedicine."

Stice presented the genetically identical calves, George, Charlie, Albert et al., as "the first cloned transgenic cattle produced in the world."

He explained that "what we are trying to replace is the traditional method of making transgenic animals. That is, taking an embryo, putting it under a micromanipulation plate and injecting it with DNA.

"This is a very inefficient procedure," he observed. "You have to produce and inject a lot of embryos, then transfer them into a number of animals, in hopes of getting a couple of transgenic, or genetically modified, animals in the end. It's feasible and workable in mice, but almost unworkable in cattle, because the efficiency of the procedure is so low."

In contrast, Stice summed up for his symposium audience, the procedures that brought George, Charlie, Albert and their bovine ilk into the world earlier this year.

"What we can do with cloning and transgenics," he began, "is start out with some cell line, genetically modify it, select for the cell line that has that genetic modification, and go through the cloning process, using in our case fetal fibroblast cells.

"Whereas with microinjection and making transgenic cattle," Stice continued, "it takes about nine months, if you are successful — and that's a big if — to produce one transgenic calf. If it's a male — hopefully — you can, after two years of artifical insemination, go and produce a large number of transgenic animals.

"With cloning," he pointed out, "we can do all that in about nine months. It's a faster procedure than microinjection, and you can also select the sex. We can start out with a female cell line, say, if we want to produce dairy herds or pharmaceuticals in milk. You can collect only female cell lines, put the gene of interest in them, and make the cloned offspring."

He noted, apropos such DNA insertion, that "the whole area of gene targeting has opened up with cloning technology."

Stice also allowed that "our procedure is still rather inefficient, and we're looking at ways to improve it. We lose about 90 percent of our embryos during the first seven days of development. But once these embryos are developed to blastocyst stage — about the 100-to-200-cell stage — we can get them into cows and produce pretty high pregnany rates. For cattle, our rate of 30 percent isn't too bad."

ACT's chief scientific officer followed up by observing that "the procedure right now is efficient enough in our hands that we think we can use this technology to make transgenic animals for various applications.

"One of the orders that we've taken recently, in collaboration with Genzyme Transgenics Corp. [of Framingham, Mass.], uses ACT's technology along with the proprietary technology of Genzyme Transgenics to produce human serum albumen in the milk of cows.

"It's a perfect product to produce in cows," Stice went on, "because you need a lot of it. About 440 metric tons of plasma derived albumen is used annually worldwide, with sales close to $1 billion. We believe we can produce about 80 kilograms a year from each cow."

Stice concluded his symposium presentation by saying: "There are a number of commercial applications for cloning the transgenics. None of them that I've talked about today are advocating the cloning of humans." *

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