Biologists Develop Efficient Genetic Modification Of Human Embryonic Stem Cells

[Guest guest]

Biologists Develop Efficient Genetic Modification Of Human Embryonic Stem Cells

http://www.medicalnewstoday.com/articles/175490.php

Biologists have developed an efficient way to genetically modify human embryonic

stem cells. Their approach, which uses bacterial artificial chromosomes to swap

in defective copies of genes, will make possible the rapid development of stem

cell lines that can both serve as models for human genetic diseases and as

testbeds on which to screen potential treatments, they say.

" This will help to open up the whole human embryonic stem cell field. Otherwise,

there's really few efficient ways you can study genetics with them, " said Yang

Xu, professor of biology at the University of California, San Diego who directed

the research. Xu and co-authors Hoseok Song and Sun-Ku Chung, both postdoctoral

fellows in Xu's research group, describe their technique in the January 8 issue

of the journal Cell Stem Cell.

Most attempts to alter the genetic makeup of the cells have proved too

inefficient, Xu said. His group used bacterial artificial chromosomes, or BACs,

to improve the yield.

BACs are synthesized circles of human DNA, which bacteria will replicate just

like their own native chromosomes. Commercially available BACs can be modified

within bacterial cells to insert altered copies of specific genes. Once the

modified BACs are introduced into human cells, they will sometimes pair up with

a matching segment of a human chromosome and swap segments of DNA, a process

called homologous recombination.

The advantage of using BACS to alter the genetic code in human cells comes from

the long flanking sequences on either side of the modified gene, which increases

the chance that the BAC with line up with native DNA in position for a swap.

Other genetic approaches have been limited by shorter segments of DNA.

Using BACs, the team was able to substitute modified genes in 20 percent of

treated cells. Standard methods of genetic modification typically achieve

modification in fewer than one percent of cells, Xu said.

His group successfully transferred a defective copy of the gene p53, which

suppresses cancer, into a human embryonic stem cell line. By repeating the

process in a second round, they developed a line of cells in which both copies

of the genes were disrupted.

They also report success with a different gene, ATM, which when mutated in

humans causes Ataxia-telangiectasia, a disease characterized by a host of

systemic defects including increased cancer risk, degeneration of specific types

of brain cells and degraded telomeres, the protective caps at the end of each

chromosome.

Genetically engineered mice with two bad copies of the ATM gene share some of

these traits with human patients, but not all. Neurons don't degenerate in ATM

mice, for example, and the telomeres are long. " If you want to study accelerated

shortening of telomeres, you can't do it in the mouse. You can only do it in

human cells, " Xu said.

Those differences propelled Xu's group to develop human cell lines instead, with

the hope that some of the processes that go wrong in human patients could be

studied in the lab. Already, they have demonstrated that their ATM-deficient

embryonic stem cell line has damaged telomeres. Other characteristics, such as

the degeneration of specific types of neurons, will be the subject of future

experiments, Xu said.

The authors say their approach can easily be adapted to modify other human genes

within other stem cells lines. For their initial work, Xu's group used a cell

line that easily forms new colonies from single cells, but they also repeated

the procedure in a cell line called H9, which has proved difficult to

manipulate.

Because H9 was among the few cells lines approved for use by researchers funded

by the federal government before new lines began to be approved in mid-December

2009, many researchers already have considerable experience with coaxing the

cells into differentiating into specific types of tissues, for example, which

would make the ability to genetically modify them particularly valuable.

The California Institute for Regenerative Medicine, the state's stem cell

research agency, funded this project.