Genetics and Health Health Article

MAPPING GENES ON CHROMOSOMES

Very effective methods have been developed to identify which genes for specific traits or diseases are located on which chromosomes, and to pinpoint the location on the relevant chromosome. The smallest genes consist of only a few hundred bases of DNA; the largest known human gene, which is mutated in Duchenne muscular dystrophy, is 2 million base pairs in length. A surprising feature of all nonbacterial genes is their organization into "introns" and "exons." Introns are noncoding stretches of DNA within the gene which are transcribed into RNA but then spliced out before the RNA is translated into protein. There are also untranslated noncoding regions at each end of the gene. There are lots of signals built into the sequence of the DNA—for initiation, stopping, splicing, and other functions crucial to defining the genes in the lengthy DNA molecules; for regulating their expression as mRNA and proteins; and for coordinating regulation of related genes. Some genes have only a single exon; others have up to one hundred interruptions with introns. The reasons are still quite obscure.

Gene mapping before 1950 was limited to the X chromosome, deduced by mother-to-son transmission in pedigrees (diagrams of family histories) for X-linked diseases (e.g., hemophilia) or traits (e.g., color blindness). A few genes were located on abnormal chromosomes by careful clinical correlations. Improved chromosomal analyses permitted formation of double-stranded DNA between fluorescent-labeled probe DNA and the DNA in a particular chromosome. Another method used mouse/human hybrid cells with one each of the different human chromosomes—if a human gene could be detected in the presence of the mouse genes, that gene must be coded for on the single human chromosome present. Once one gene is located, another gene which is linked in transmission from generation to generation can be deduced to be on the same chromosome. For example, the gene for cystic fibrosis was placed near linked markers on chromosome 7. This is truly a needle-in-the-haystack approach, since there are some 30,000 to 40,000 genes, distributed on the 23 pairs of chromosomes.

Everything changed with the new methods of recombinant DNA and the polymerase chain reaction—a way to produce millions of copies of a particular DNA molecule isolated or synthesized in tiny amounts. Increasingly, genes are being identified without the benefit of an initial chromosomal localization. A scan of the entire genome (across all chromosomes) is performed in a search for linkage to fairly common variants of genes that serve as well-spaced markers, even without knowing their function. Many steps in this approach are now automated, thousands of samples can be processed, and powerful computer programs sift through hundreds or thousands of markers to find clues for localization of the presumed gene or genes for a disease. Segments of DNA from the suspected chromosomal region can be cloned into specialized vectors. Linking together all such fragments permits scientists to assemble the genome sequence of humans or of many other organisms (e.g., yeast, fruitfly, bacteria, earthworm, mouse). A much more complicated mapping process is helpful in locating multiple genes for complex diseases like diabetes, high blood pressure, or depression.

THE HUMAN GENOME PROJECT

According to Francis Collins, director of the National Human Genome Research Institute, "mapping the human genetic terrain may rank with the great expeditions of Lewis and Clark, Sir Edmund Hillary, and the Apollo Program." In the early 1970s, obtaining sequence information on DNA or RNA was arduous, typically requiring an entire year to deduce about fifteen nucleotides. Advances in laboratory methods triggered hope in the mid-1980s that a massive scale-up could eventually sequence the entire 3 billion base pairs of human DNA. The project was officially launched in October 1990 in the United States and soon became international, with major efforts in Britain, France, Scandinavia, and Japan. Specific goals, by chromosome, were set to achieve the mapping of genes to chromosomes, the physical map of DNA fragments, and then the DNA sequence. By 1994 there were 5,000 highly useful markers for the genetic map, and overlapping cloned fragments of DNA to create physical maps, using various techniques. Many clever schemes have been put to use to assure sufficient overlaps to orient the location and direction of DNA sequence fragments. Powerful sequencing methods accelerated the target completion date from 2005 to 2003, and then to 2001. Work to "clean-up" the sequence is ongoing.

Nevertheless, having the entire sequence has been likened to having the complete works of William Shakespeare as a sequence of the twenty-six English letters with no punctuation of any kind. Figuring out where the genes are and how they are turned on and off, or up and down, during life's events is a huge remaining task. In reality, the work of understanding the genome has only just begun. Computer algorithms, including one called "GRAIL," have been designed to find and use characteristics that may distinguish coding regions from the other 95 percent of the DNA sequence. Working backward from the mRNA molecules by forming double-stranded complementary DNA with the enzyme reverse transcriptase, and then sequencing the cDNA or even partial cDNA as expressed sequence tags, has accelerated this work.

Another powerful approach has utilized the theme of "unity in diversity" that characterizes all living things. There are amazing homologies between genes in humans and genes for similar functions in mice, earthworms, fruitflies, and even yeast cells, all of which have smaller genomes than humans. Computer databases available to scientists throughout the world permit "virtual experiments" using knowledge of a disease-related gene in the mouse, for example, to deduce what gene might account for a similar disease in humans.

Regulation of gene expression is a crucial feature of differentiated cells in complex organisms and of development from the single fertilized egg cell. Except for red blood cells, which have no nucleus, all other cells in any individual have a nucleus, chromosomes, and DNA—the same DNA. So the information content is essentially the same in all cells, yet quite different sets of genes are active in the blood, liver, kidney, brain, heart, and other organs and in cancer cells. Much is being learned about the ways in which genes are regulated in health and disease.