Human Genome Project

In 1990, the U.S. Human Genome Project begun formally. Department of Energy and the National Institutes of Health. The project originally was planned to last 15 years, but rapid technological advances accelerated the completion date to 2003. Project goals were to identify all the approximately 20,000-25,000 genes in human DNA, determine the sequences of the 3 billion chemical base pairs that make up human DNA, store this information in databases, improve tools for data analysis, transfer related technologies to the private sector, and address the ethical, legal, and social issues (ELSI) that may arise from the project.

To help achieve these goals, researchers also studied the genetic makeup of several nonhuman organisms. These include the common human gut bacterium Escherichia coli, the fruit fly, and the laboratory mouse.

A unique aspect of the U.S. Human Genome Project is that it was the first large scientific undertaking to address potential ELSI implications arising from project data.

Another important feature of the project was the federal government's long-standing dedication to the transfer of technology to the private sector. By licensing technologies to private companies and awarding grants for innovative research, the project catalyzed the multibillion-dollar U.S. biotechnology industry and fostered the development of new medical applications.

Landmark papers detailing sequence and analysis of the human genome were published in February 2001 and April 2003 issues of Nature and Science. See an index of these papers and learn more about the insights gained from them.

Discovery of Transposons

The chromosomal basis of heredity was already well established by the time McClintock began her graduate training in the Botany Department at Cornell University. Her experiments laid the groundwork for a serie of cytogenetic discoveries by the Cornell maize genetics group between 1929 and 1935. McClintock developed a method for using broken chromosomes to generate new mutations. Among the progeny of plants that had received a broken chromosome from each parent, she observed unstable mutations at an unexpectedly high frequency, as well as a unique mutation that defined a regular site of chromosome breakage. These observations so intrigued her that she began an intensive investigation of the chromosome-breaking locus. Within several years she had learned enough to reach the conclusion, published in 1948, that the chromosome-breaking locus did something unknown for any genetic locus: it moved from one chromosomal location to another, a phenomenon she called transposition. 

 

The study of transposable genetic elements and transposition became the central theme of her genetic experiments from the mid 1940s until the end of her active research career. This was incredulous at the time, DNA was believed to be stable and invariable. These jumping elements were isolated from the bacterium Escherichia coli in the late 1960's and were further defined as specific, small fragments of DNA which were given the name transposons. The scientific interest in transposons increased during the 1970's, when it appeared that they assisted in the transfer of bacterial resistance to antibiotics. Furthermore, it soon became evident that they caused most of the spontaneous mutations occurring in laboratory populations of more sophisticated organisms, such as yeast and the fruit fly. We now know that transposons are ubiquitous and may comprise up to 20% of an organism's genome.

Retrotransposons

Retrotransposons move by a "copy and paste" mechanism but in contrast to the transposons described above, the copy is made of RNA, not DNA.

The RNA copies are then transcribed back into DNA — using a reverse transcriptase — and these are inserted into new locations in the genome.

Many retrotransposons have long terminal repeats (LTRs) at their ends that may contain over 1000 base pairs in each.

Like DNA transposons, retrotransposons generate direct repeats at their new sites of insertion. In fact, it is the presence of these direct repeats that often is the clue that the intervening stretch of DNA arrived there by retrotransposition.

42% of the entire human genome consists of retrotransposons.

Transposons (Jumping Genes)

Grains of Indian corn come in different colors, such as purple, yellow and white. Sometimes the individual grains are purple with white streaks or mottling. This mottling effect defies Mendel's basic principles of genetics because individual grains may be multicolored rather than a single color. The movement of transposons on chromosomes may result in colored, non-colored and variegated grains that do not fit traditional Mendelian ratios based solely on chromosome assortment during meiosis and random combination of gametes. The explanation for this phenomenon involves "jumping genes" or transposons, and earned Dr. Barbara McClintock the prestigious Nobel Prize in Medicine in 1983 for her life-long research on corn genetics.

Transposons are genes that move from one location to another on a chromosome. In the pigmented aleurone layer of corn grains, the position of transposons may inhibit or block pigment production in some cells. For example, if the transposon moves to a position adjacent to a pigment-producing gene, the cells are unable to produce the purple pigment. This results in white streaks or mottling rather than a solid purple grain. The duration of a transposon in this "turned off" position affects the degree of mottling. If the pigmentation gene is turned off long enough by a transposon, the grain will be completely unpigmented. The reddish-purple patterns caused by transposons may be blotches, dots, irregular lines and streaks.

The following illustration shows how grain color in Indian corn may be affected by transposons. The different cards represent a linear sequence of genes on a chromosome. The ace of spades represents a transposon that moves to different positions on the chromosome. The jack of diamonds represents the gene for purple pigmentation in the corn grain. When the transposon (ace of spades) moves to a position adjacent to the gene for pigmentation (jack of diamonds), the pigmentation gene is blocked and no purple is synthesized (white area):

When the transposon (ace of spades) moves away from the gene for pigment production (jack of diamonds), the production of purple pigment is resumed (continuous purple area). In this example the gene for pigment production (jack of diamonds) is not adjacent to a transposon (ace of spades):

When a transposon moves to different positions within cells of the corn kernel, the coloration gene is "turned on" or "turned off" depending on whether it lands in a position adjacent to the pigmentation gene. Transposons may also have a profound effect on embryonic development and tumor formation in animal cells. Oncogenes (genes that cause tumors) may be activated by the random reshuffling of transposons to a position adjacent to the oncogene. Transposons may also be useful in genetic engineering with eukaryotic cells, by splicing in transposons to activate certain genes. The implications from Barbara McClintock's discovery of transposons may be far-reaching and as significant as Watson and Crick's discovery of the structure of DNA.

Human Gene Transfer

Human gene transfer is the process of transferring genetic material (DNA or RNA) into a person. DNA may be transferred as "naked" DNA, encapsulated DNA, or DNA within another organism, such as a virus. Use of retroviral vectors in humans also constitutes human gene transfer when the virus contains enzymes that result in a DNA copy of the RNA genome. 

Human gene transfer is experimental and is being studied to see whether it could treat certain health problems by compensating for defective genes, producing a potentially therapeutic substance, or triggering the immune system to fight disease. Human gene transfer may help improve genetic disorders, particularly those conditions that result from inborn errors in a single gene (for example, sickle cell anemia, hemophilia, and cystic fibrosis). It may also hold promise for diseases with more complex origins, like cancer and heart disease. Gene transfer is also being studied as a possible treatment for certain infectious diseases, such as AIDS. This type of experimentation is sometimes called "gene therapy" research. 

Scientists are attempting to determine whether human gene transfer can be safe and effective as a treatment for disease. Some experimental gene transfer procedures involve the introduction of DNA into cells, which then are injected into a person with disease. All such human gene transfer research studies require approval by the UCI Institutional Review Board (IRB), the UCI Institutional Biosafety Committee (IBC), and the NIH Recombinant DNA Advisory Committee (RAC).

Germline Gene Transfer

Gene transfer represents a relatively new possibility for the treatment of rare genetic disorders and common multifactorial diseases by changing the expression of a person's genes. Typically gene transfer involves using a vector such as a virus to deliver a therapeutic gene to the appropriate target cells. The technique, which is still in its infancy and is not yet available outside clinical trials, was originally envisaged as a treatment of monogenic disorders, but the majority of trials now involve the treatment of cancer, infectious diseases and vascular disease. Human gene transfer raises several important ethical issues, in particular the potential use of genetic therapies for genetic enhancement and the potential impact of germline gene transfer on future generations.

 

Scientific Issues

Gene transfer can be targeted to somatic (body) or germ (egg and sperm) cells. In somatic gene transfer the recipient's genome is changed, but the change is not passed on to the next generation. In germline gene transfer, the parents' egg and sperm cells are changed with the goal of passing on the changes to their offspring. Germline gene transfer is not being actively investigated, at least in larger animals and humans, although a great deal of discussion is being conducted about its value and desirability.

Many people falsely assume that germline gene transfer is already routine. For example, news reports of parents selecting a genetically tested egg for implantation or choosing the sex of their unborn child may lead the public to think that gene transfer is occurring, when actually, in these cases, genetic information is being used for selection, with no cells being altered or changed. In addition, in 2001 scientists confirmed the birth of 30 genetically altered children whose mothers had undergone a procedure called ooplasmic transfer. In this process, doctors injected some of the contents of a healthy donor egg into an egg from a woman with infertility problems. The result was an egg with two types of mitochondria, cellular structures that contain a minuscule amount of DNA and that provide energy for the cell. The children born following this procedure thus have three genetic parents, since they carry DNA from the donor as well as the mother and father. Although the researchers announced this as the "first case of human germline genetic modification," the gene transfer was an inadvertent side effect of the infertility procedure.

Gene Therapy

Gene therapy using an Adenovirus vector. A new gene is inserted into an adenovirus vector, which is used to introduce the modified DNA into a human cell. If the treatment is successful, the new gene will make a functional protein.

Gene therapy may be used for treating, or even curing, genetic and acquired diseases like cancer

and AIDS by using normal genes to supplement or replace defective genes or to bolster a normal function such as immunity. It can be used to target somatic (i.e., body) or gametes (i.e., egg and sperm) cells. In somatic gene therapy, the genome of the recipient is changed, but this change is not passed along to the next generation. In contrast, in germline gene therapy, the egg and sperm cells of the parents are changed for the purpose of passing on the changes to their offspring.

There are basically two ways of implementing a gene therapy treatment:

1. Ex vivo, which means “outside the body” – Cells from the patient’s blood or bone marrow are removed and grown in the laboratory. They are then exposed to a virus carrying the desired gene. The virus enters the cells, and the desired gene becomes part of the DNA of the cells. The cells are allowed to grow in the laboratory before being returned to the patient by injection into a vein.

2. In vivo, which means “inside the body” – No cells are removed from the patient’s body. Instead, vectors are used to deliver the desired gene to cells in the patient’s body.

As of June 2001, more than 500 clinical gene-therapy trials involving about 3,500 patients have been identified worldwide. Around 78% of these are in the United States, with Europe having 18%. These trials focus on various types of cancer, although other multigenic diseases are being studied as well. Recently, two children born with severe combined immunodeficiency disorder (“SCID”) were reported to have been cured after being given genetically engineered cells.

Gene therapy faces many obstacles before it can become a practical approach for treating disease.