Heredity and the gene
Knowledge of the principles of heredity is so basic to our fundamental understanding of the biological sciences that it is hard to believe that these principles were discovered only in the 1860s (and their importance was realized less than a century ago). However, a practical knowledge of the hereditary process came long before its mechanism was understood. Archeologists have discovered that as long as 7,000 years ago farmers in Central America were improving crops of corn by planting hybrid corn seeds that had developed preferred characteristics. Over 6,000 years ago, the Chinese learned how to develop superior strains of rice. An ancient Babylonian tablet shows a pedigree of a family of horses through five generations, with detailed information about height, length of the mane, and other traits, revealing that they had some knowledge that these traits were transmitted. Farmers and gardeners have continued to practice this type of selective breeding in both plants and animals. Each time an individual plant or animal appeared with a desired characteristic, it was bred again to produce more with similar traits. For example, at harvest time farmers would select heads of wheat that had the most or largest kernels and save them to use as seed the next year.
Science (or more generally, curiosity) received a large boost during the Renaissance. During the 16th and 17th centuries, the interest in the physical and natural sciences grew. This increased interest laid the foundation for science as we know it now. As gene technology bears on a large number of different disciplines (biochemistry, genetics, cell biology, engineering, etc.), a couple of key findings in different areas will be discussed here. It is interesting to note that some of the great discoveries that were made were not viewed to be very important at the time, just because the proper framework to understand the importance of the findings had not yet been developed. Also, it is important to realize that many of the seminal discoveries were made at the interface between two or more disciplines. The importance of this interdisciplinary insight and research toward developing new concepts and creating new ways of looking at problems persists to this day.
By the end of the 17th century, Hooke in England and van Leeuwenhoek in The Netherlands came up with microscopes. The microscope constructed by van Leeuwenhoek magnified 200 times, enough to see into "a new world" of little beasties that no one had ever seen. Hooke observed how cork was composed of a series of tiny cubicles, much like a honeycomb. He coined the word "cell" for such a cubicle, since it reminded him of the tiny cubicles used by monks in monasteries.
In the beginning of the 19th century, Brown, a Scottish botanist, found "opaque spots" in the cells of orchids. Upon further investigation, he found such spots in cells of all plants. He named the spot in the cell the nucleus, the latin name for "little nut". We will see later that the nucleus is the place where the genetic material is located and its expression is regulated. The current connotation of the word nucleus accurately represents the later realization that the nucleus is the "main office" of the cell, and has replaced the original meaning of the word.
It is striking that the discovery of the principles of heredity was not made by an eminent scientist, but rather by a monk doing experiments in the vegetable garden of a monastery. The monk, Gregor Mendel, was intrigued by the multitude of shapes and colors of all living things, even occurring within a single species growing/living at a single site. He used 34 different varieties of self-fertilizing peas for his experiments (self-fertilizing means that the egg under natural conditions is fertilized by the pollen from the same plant; in peas the flower remains closed during the fertilization period, thus preventing pollen from other plants fertilizing the eggs); he grew these varieties of peas for several pea generations. He crossbred different kinds of peas by opening up a flower, cutting off the stamen, and putting pollen from another flower on the stigma. He tied a tiny bag over the flower to prevent any fertilization by other pollen. He worked on the problem for about eight years, and got results from 10,000 different plants. He selected seven characteristics to study (the color and the shape of the seed, the color of the flowers, the color and form of the pods, the position of the flowers, and the length of the stem), and systematically focused on one or two parameters at a time. In his first experiment, he crossbred plants with round and wrinkled seeds. The progeny only had round seeds, which was in contradiction with the theory current at that time, that the progeny contained a blend of the characteristics of the two parents! He grew plants from the progeny seed, wondering what the seeds of these plants would look like. This time, there were both wrinkled and round seeds, the round seeds outnumbering the wrinkled ones by a factor of three. In the next generation, the plants grown from wrinkled seeds only gave wrinkled seeds, whereas plants from round seeds once again produced a mix of round and wrinkled ones, in a ratio of three to one. He did similar experiments while looking at the other properties he had selected.
On the basis of the experiments he did, he formulated several rules: (1) heredity is determined by distinct elements contained in the two cells contributed by the parents of the organism, and these elements combine randomly; (2) each characteristic of an organism is determined separately from the others; and (3) when two different characteristics combine, one characteristic will dominate over the other (for his second and third rule he just had luck; it turned out to be not applicable in all cases). He decided to use single letters for each characteristic; a capital letter denoted the dominant factor (the one that determines the phenotype), and the same letter in lower case denoted the recessive one (the one that does not express itself in the phenotype, unless the dominant factor is absent). He realized that progeny combined factors from both parents, and that in the pollen or the egg only half of the information of the parent was present. This insight laid the foundation for modern genetics. He presented his findings and conclusions to a conference of local intellectuals. The response was a polite silence. Nevertheless, they invited him to publish his results in a scientific journal, which he did in 1866. His paper was totally neglected by the scientists of his time. He died 18 years later as a well-respected abbot, but without recognition for his pioneering work.
For more information on Mendelian genetics, go to: http://web.mit.edu/esgbio/www/mg/mgdir.html and click on aspects you want to have more information on.
Mendel's pioneering work chronologically was paralleled by breakthroughs in biochemistry and cell anatomy. In 1869, a Swiss graduate student, Friedrich Miescher, set out to do a chemical analysis of a cell. He was having trouble breaking down all parts of the cell when he got the idea of "digesting" it with stomach enzymes. Cells broke down upon digestion, but they always left a residue from the nucleus. Miescher called the residue "nuclein". He combined pleasure with business angling for salmon in the Rhine river near his home. He had found that salmon sperm had very large nuclei, almost half of the cell itself, and Miescher was able to accumulate a good supply of his newly discovered nuclein. It was fortunate that he did his experiments more than a century ago, because one would be hard-pressed to find any salmon in the polluted Rhine river now. One of Miescher's coworkers observed that nuclein was composed of carbon, oxygen, hydrogen, nitrogen and phosphorus. Jars and jars of the white sticky stuff, the nuclein, stood on Miescher's laboratory shelves for years, but he did not realize that what he had isolated was a crude preparation of DNA, the genetic material that contained the molecular basis for Mendel's results. The chemical characterization of the nuclein was published by Miescher's friends after his death, but at that time it was just a curiosity, a piece of the puzzle that could not be fitted into its proper perspective.
In the middle of the 19th century, a young lad by the name of William Perkin was asked by his employer to find ways to synthetically make quinine, a natural drug used to treat malaria and other maladies. He went ahead, and mixed all kinds of chemicals. One mix, aniline and potassium dichromate resulted in a purple goo. Adding alcohol made a wonderful purple solution that did not cure malaria, but could be used as a dye. A quarter of a century later, a German scientist, Walter Flemming, stained some cells with Perkin's dye, to see how that would look under the microscope. Suddenly, the nucleus Brown had described half a century earlier became distinctly clear, since it absorbed the dye. Flemming called the mass of colored material in the nucleus chromatin. When he stained sections of growing tissue, he saw the cells in the process of division. He watched the chromatin bunch up into short, threadlike bundles, and he called these chromosomes. As he studied the bunching up process, he saw the chromosomes double in number. He watched the chromosomes begin to pull apart. Half of them moved to one end of the cell, the other half went to the other end. Because the chromosomes spread out like threads during this process, Flemming called the process mitosis ("thread" in Greek). Half the chromosomes moved into one of the new daughter cells that formed when the cell divided, and half went into the other new cell. They were exact duplicates. When Flemming wrote about mitosis and chromosomes in 1882, he did not connect it to inheritance. Mendel's laws had not yet been rediscovered.
Five years later, van Beneden, a Belgian scientist, discovered that every cell in a body has the same number of chromosomes, with the exception of the sperm or egg cell, which contains only half the "usual" number of chromosomes. He also found that different species have different numbers of chromosomes: a human has 46, a fruit fly has 8, and a crayfish has some 200. The giant sequoia can live with 22. Note that up to this point all important findings in this area had been made in Europe. Basic sciences in the US had not come of age yet.
Around the beginning of the 20th century, three scientists (De Vries in The Netherlands, Correns in Germany, and Tschermak in Austria) independently and virtually simultaneously performed plant breeding experiments that greatly resembled the work done by Mendel half a century earlier. All three only discovered Mendel's publication while doing a literature study in preparation of their publication. Around the same time, the name "gene" was coined to describe the hereditary substance, and "genetics" became the name of the rapidly emerging discipline of gene study and characterization.
Advances in the field of genetics were also fueled by the discoveries by Flemming, van Beneden, and others on the cellular/sub-cellular level. In the twenties and thirties of this century, chromosomes became the subject of intense study, and it is mainly the pioneering work of two groups (those of Morgan and Castle) that has contributed greatly to the further development of the concept of genes in relation to chromosomes. Both groups studied the fruitfly (Drosophila melanogaster), whose eight chromosomes are all readily visible under the light microscope. Six chromosomes could be divided up in three pairs of morphologically identical chromosomes, whereas the other two (named X) were identical in females while in males one of the two was replaced by a much smaller chromosome (Y). It was correctly assumed earlier (from work on grasshoppers by Walter Sutton) that these two chromosomes would be the ones that determined the sex of the organism. Morgan found a fruitfly mutant which had white eyes rather than the usual red ones. When this mutant, a male, was mated with a red-eyed female, all the progeny had red eyes, indicating that the red eyes were dominant. Upon self-breeding of the progeny, the red-eyed flies outnumbered the white-eyed ones by a factor of about three, as would be expected. However, it was found that all white-eyed flies were male. Thus, according to Morgan, the gene for eye color must be on the X chromosome. He was right. This was the first time that anyone had placed a specific gene on a specific chromosome.
Of course, Morgan could not do all this work by himself. He had a large group of graduate students (nicknamed the "fly squad"), some of whom later became independent researchers. One of them was Hermann Muller. He was wondering what caused the mutations. He tried outside changes: no luck: if you cut off the tail of a mouse, all progeny had tails. He tried accidents of all kinds, but no mutations were produced. He then began thinking in terms of the "world of the little". Perhaps mutations came from an ultramiscroscopic accident. Maybe the gene could not escape a speeding electron. He then tried radiation (small, energy-rich particles); he reasoned that there is some natural radiation in the atmosphere, causing rare mutations, and that he might be able to get many more mutations by irradiating his flies with X-rays (invented a couple of years before). He put hundreds of flies in gelatin capsules, irradiated them, and then bred them with untreated flies. The progeny was an interesting mix: there were flies with bulging eyes, flat eyes, dented eyes, purple eyes, yellow eyes, broad wings, curly wings, bumpy wings, etc.. Some were hyperactive, others dopy. Some were big, others small. The conclusion: mutations are caused by ultramiscroscopic collisions, making changes in the hereditary information.
The question now could focus on the chemical identity of the carrier of the hereditary information. As indicated earlier, Miescher had isolated nuclein in large amounts from salmon sperm, and unlike protein, nuclein was found to have a high concentration of phosphorus. Therefore, it was unlikely that nuclein was just a type of protein. Eventually, chemical analysis revealed that chromosomes are made of both protein and nuclein, and that nuclein is deoxyribonucleic acid (DNA). With both protein and DNA present in chromosomes, which of these carries hereditary information? Almost everyone thought that proteins must be the hereditary material because chemically it is much more complicated than DNA, and the molecule of heredity had to be able to contain an extraordinary amount of information. DNA seemed too simple..... How wrong this reasoning would turn out to be.
In 1928 Frederick Griffith, a scientist who was interested in pathology rather than in genes, made a serendipitous discovery that led to the identification of the chemical composition of genetic material. He was trying to understand the differences between the strain of Diplococcus pneumoniae that cause pneumonia (virulent strains) and the strains that are harmless (non-virulent). The virulent forms had polysaccharide capsules around them that gave them a smooth appearance. The non-virulent bacteria had no capsules and were rough. Griffith hoped that either heat-killed virulent strains or live non-virulent strains could be used as vaccine. He mixed heat-killed virulent and live non-virulent strains together in hopes of making an effective vaccine. Instead, upon injecting this mixture into mice, the mice died. How could dead bacteria be virulent? He retrieved virulent bacteria from the blood of the dead mice, and he saw that the bacteria were smooth. Something had been transferred from the dead virulent bacteria to the live non-virulent ones. Griffith called this substance the "transforming factor". This experimentation is nicely illustrated at the web site: http://web.mit.edu/esgbio/www/dogma/dogmadir.html. However, it is unfortunate that Griffith is not given any credit at the MIT web site, and only the people who followed up on it (Avery et al.) are mentioned there.
Griffith did not follow up on identifying the chemical nature of his "transforming factor". This work laid dormant for a while until 1943, when O.T. Avery and colleagues purified the "transforming factor" and found that it was DNA. This finding was viewed with a lot of suspicion by other scientists, as everyone thought that protein (consisting of a long string of subunits, each of which could have 20 different chemical structures) must be the location of hereditary information. DNA (consisting of a much longer string of subunits, each of which could have 4 different chemical structures) was viewed as inadequate to store much information. This way of thinking is typically B.C. (before computers) because we know now that an enormous amount of information can be stored even using a binary (on / off or 0 / 1) mode.
Developments were rapid from then on. In 1952 the DNA-vs-protein debate was settled conclusively by showing using radioactive isotopes that protein is not passed on from a virus to its progeny, but DNA is. The polynucleotide (DNA) molecule has a number of relatively unusual characteristics for a macromolecule, including the repeated negative charge of the phosphate groups, and the innate affinity of pyrimidine and purine bases to each other. Erwin Chargaff discovered in the forties that the base compositions of DNA from different organisms vary over a relatively wide range (for example, adenine makes up between 23 and 32% of the bases), but that the amount of A consistently was very close to the amount of T, and the amount of C essentially identical to that of G. These two equalities were the first indication that stoichiometric complexation occurs between A and T and between G and C. These issues are also illustrated at http://web.mit.edu/esgbio/www/dogma/dogmadir.html
Chargaff shared his findings with James Watson, who with the help of molecular models discovered that H-bonded base-paired structures could be formed between A and T, and between G and C, and that these structures have the same overall dimensions. Two H-bonds can be formed between A and T, and three between G and C. Watson brought this information to the attention of Francis Crick, a crystallographer. Methods had been developed to determine the structure of crystals (repetitively arranged groups of molecules) to atomic resolution, and Crick was an expert in using this technique. This culminated in the elucidation of the DNA structure in 1953. The X-ray diffraction pattern of crystallized DNA could be interpreted in terms of a helix (essentially a winding staircase) composed of two polynucleotide strands with H-bonded pairs formed between the bases of opposing strands (see the cover of Molecular Biotechnology). Molecular-modeling studies showed that the H-bonded base pairs could form only when the directional senses of the two interacting chains were opposite (anti-parallel). Also, the DNA duplex was found to be "folded" in a three-dimensional structure, a double-helix (like two helices folded into each other). Watson and Crick Crick (but not Chargaff . . . life not always is fair) got the Nobel Prize in 1962. The Nobel Committee noted that the discovery had "no immediate practical application, but determining the molecular structure of the substance that is responsible for the forms that life takes is a discovery of tremendous importance". The understatement of the year? Without some of these breakthroughs, molecular biology and biotechnology might still be in their infancy.
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Instructors | Aims
Lecture Part: Schedule | Expected Background & Textbook Info | Historical Perspective
Intro to Biotechnology | DNA, RNA and Protein Synthesis | Chemical Synthesis, Sequencing, and Amplification of DNA |
Directed Mutagenesis and Protein Engineering | Vaccines | Antibiotics & Proteins | Bioremediation |
Microbial Insecticides | Plant Genetic Engineering: Methodology | Plant Genetic Engineering: Applications | Transgenic Animals
Human Molecular Genetics | Regulatory & Ethical Aspects | Biotech Inventions | Additional Materials
Lab Part: Aims and Expectations | Schedule
Center for Bioenergy & Photosynthesis
Arizona State University
Room PSD 209
Tempe, AZ 85287-1604
29 August 2007
phone: (480) 965-1963
fax: (480) 965-2747