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Dep. of Agronomy and Plant Genetics and Microbial and Plant Genomics Institute, Univ. of Minnesota, 1991 Upper Buford Cir., St. Paul, MN 55108
* Corresponding author (phill005{at}umn.edu)
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 TOP ABSTRACT FOUNDATIONS OF MODERN DAY...REFERENCES |
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Abbreviations: QTL, quantitative trait loci • PCR, polymerase chain reaction • AFLPs, amplified fragment length polymorphisms • SSRs, simple sequence repeats • MAS, marker-assisted selection
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FOUNDATIONS OF MODERN DAY GENETICS AND GENOMICS |
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TOPABSTRACTFOUNDATIONS OF MODERN DAY...REFERENCES |
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Like the 900-word report by Watson and Crick, CSSA had a rather inauspicious start. In the case of the Nature report, Watson convinced his sister Elizabeth to spend one Saturday afternoon typing a short report (Breo, 1993) that started: "We wish to suggest a structure for the salt of DNA. This structure has novel features which are of considerable biological interest." What an understatement—not only for biology in general, but also because it ushered in an unprecedented era of cell biology and molecular genetics applicable to medicine and agriculture. It has been a long road from the first hybrid plant created by Thomas Fairchild in 1719 [cross of Sweet William and a carnation (Dianthus caryophyllus L.)] (Blair, 1720) to Gregor Mendel's 1866 "Experiments in Plant Hybridization" (Mendel, 1866) to Lewis Stadler's demonstration of the mutagenic effects of X-rays on barley (Hordeum vulgare L.) (Stadler, 1928) to hybrid corn (Zea mays L.) (Shull, 1908) to today. The opportunities to apply science to agriculture are so exciting and extensive that we can hardly imagine the road ahead.
Nobel Prize winners, George Beadle and Barbara McClintock, along with two other pioneers in plant genetics, Charles Burnham and Marcus Rhoades (Fig. 1 ), all worked together to lay much of the groundwork for understanding genetics. In the early years of CSSA, the C-1 Division mainly encompassed breeding and cytology, but soon started to embrace molecular genetics research as well as tissue culture and other important strategies applied to crop improvement. Beadle (a pioneering researcher in maize genetics) and Tatum, working with the bread mold Neurospora, developed the one gene–one enzyme hypothesis (Beadle and Tatum, 1941). The importance of transposable elements (then usually referred to as controlling elements), which could move around on the chromosome or even jump between chromosomes was just being recognized by the genetics community after years of work by McClintock (1951) on color patterns of corn kernels. Her research began to highlight aspects of genome plasticity—a feature of crop genomes still poorly understood but clearly discussed in her Nobel Prize speech (Odelberg, 1984). The discovery of repetitive satellite DNA (Britten and Kohne, 1970) led to the surprising fact that most of the genome is not encoding genes. The function of repetitive DNA is not well defined yet today.
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We also have learned that variation can arise de novo. Variation not present in either parent can appear in progenies, due to a variety of genetic mechanisms, such as gene magnification or reduction, DNA methylation, intragenic recombination, unequal crossing over, transposable elements, and many others. Perhaps some of these mechanisms are operating when breeding programs continue to make excellent progress even when crossing the best by the best. Although such crosses are considered as involving a narrow genetic base, they still can yield excellent results (Rasmusson and Phillips, 1997). The concept that higher yielding types only come from the combination of favorable genes provided by the parents may need to be revisited more seriously in the future.
The discovery of restriction enzymes (Smith and Wilcox, 1970), DNA polymerases (Eom et al.,1996) and ligases (Subramanya et al.,1996) ushered in the era of genetic engineering. The first recombinant DNA techniques were developed by Smith and Nathans (1971) to combine genes from different organisms. This technology together with the ability to regenerate crop plants from cells (e.g., Green and Phillips, 1975; Conger and Carabia, 1978; Lazzeri et al., 1987a, 1987b), since transformation involved shooting DNA into cells (Sanford et al., 1987), opened the door for use of even wider genetic diversity than previously possible. Until that time, we were dependent on existing sexually compatible germplasm as our source of genes, including natural and chemically or physically induced mutations. But for the most part, the new genes for breeding came from germplasm collected from around the world and placed in "gene banks" for everyone's use.
Evolution of CSSA C-7 Division of "Genomics, Molecular Genetics & Biotechnology"
Within this historical framework, the C-7 Division of CSSA was born. There was a great deal of discussion, especially among the C-1 members about the need to stimulate the communication of molecular genetics research within the CSSA membership. There was an awareness of the integral nature of the C-1 Division and a potential C-7 Division, and a concern that the divisions might compete for the same members. Yet there also was the desire to highlight cell biology and molecular genetics research and to attract more plant molecular geneticists to the CSSA. After considerable discussion "on the street," Walt Fehr brought a petition in 1984 to the CSSA Board of Directors, chaired by CSSA President Wayne Keim, to establish a provisional division according to the CSSA constitutional guidelines. The action was: "It was moved to establish Division C-7, Cell Biology and Molecular Genetics, on a 2-yr provisional basis after which time the matter will be presented to the entire voting membership relative to permanent status." The motion was seconded and carried (Anonymous, 1985). Two years later, the approved C-7 provisional division was voted in as a regular division. Although the initial name of C-7 was "Cell Biology & Molecular Genetics," the name "Genomics, Molecular Genetics & Biotechnology" has been used since 2003. The first Chair of the provisional C-7 Division was Glenn Collins, the second Chair was Nick Frey, and the Board Representative was Burle Gengenbach. At the time of the provisional C-7 Division approval vote, I was the C-1 chair, Jerry Nelson was the C-2 chair, Lowell Moser was the C-3 chair, M.B. McDonald was the C-4 chair, D.P. Martin was the C-5 chair, and Dave Sleper was the C-6 chair. On December 1, 1986, the following action was taken by the CSSA Board of Directors: "It was moved to grant permanent status to Division C-7, Cell Biology and Molecular Genetics, subject to membership approval." The motion was seconded and carried (Anonymous, 1987). Today, the C-7 Division is almost 21 yr old.
The members of C-7 have been leaders in much of the application of cell biology and molecular genetics to agriculture. Membership of the C-7 Division of CSSA reads like a "Who's Who" in plant genetics, genomics, and biotechnology with impressive contributions in numerous areas, including male sterility, QTLs (Quantitative Trait Loci), regeneration of crop species, wide hybridization, haploid production, QTL analysis, marker assisted selection, associative mapping, apomixis, bioinformatics, molecular genetic markers, wheat cytogenetics, endosperm mutants, physiology, comparative genetics, disease resistance, transgenic crops, amino acid and protein genetics, centromere structure, anthrax, drought tolerance, polyploidy, genetic control of meiosis, transposable elements, chromosome elimination, DNA technology, and DNA fingerprinting.
Development of Genetic Information and Tools
Over the past 21 yr, many articles in Crop Science have reported research on genetic analysis of specific mutations, QTLs for a wide variety of traits, microsatellite markers, molecular genetic linkage maps, flanking markers for specific genes, genetic diversity studies, transgenics, regulation of genes, doubled haploid lines, gene cloning, gene flow, somaclonal variation, selectable markers, and other transformation techniques.
Interestingly, DNA-based markers have almost replaced the use of traditional genetic markers, such as mutations for dwarfs, chlorophyll deficiencies, anthocyanin color, and a variety of seed mutants, which crop scientists discovered, mapped, biochemically analyzed, and used for a myriad of genetic discoveries. Molecular genetic maps have been developed for a wide variety of crops (Phillips and Vasil, 2001). Genetic markers allow the tracking of minute portions of each chromosome in one experiment as opposed to the two or three-point linkage studies of the past. The PCR (Polymerase Chain Reaction) technology added additional powerful markers by greatly enhancing our ability to amplify DNA (Saiki et al., 1988) from minute quantities, instead of depending on replication of bacteria possessing a segment of the foreign DNA. This rapid and precise technology led to new kinds of molecular genetic markers, such as AFLPs (Amplified Fragment Length Polymorphisms), and SSRs (Simple Sequence Repeats) (Jones et al., 1997).
Various cytogenetic discoveries also have led to efficient mapping systems. One of these approaches is through the use of gametocidal chromosomes that produce a wide variety of chromosome deletions in wheat (Endo and Gill, 1996). The use of these physical deletions allowed researchers to show that wheat genes are largely located in the distal 20% of the chromosomes. This information is helping scientists to identify parts of the huge wheat genome that should receive first priority for mapping (Jiang and Gill, 1994). Another high-throughput approach is the production of oat-maize addition lines and radiation hybrids (Ananiev et al., 1997; Kynast et al., 2000; Kynast et al., 2001; Okagaki et al., 2001; Kynast et al., 2002). Crosses between oat (
) and maize (
), followed by embryo rescue, yields plants that are either oat haploids (2/3 time) or oat-maize addition lines with one or more maize chromosomes present in an oat background (1/3 time).
Figure 2 illustrates how easy it is to identify the maize chromosomes in an oat genomic background by the use of GISH. These plants are then irradiated to break up the maize chromosome, resulting in plants (Radiation Hybrids) with just a piece of a specific maize chromosome (deletion of a portion of the maize chromosome, or translocation of a piece of the maize chromosome to an oat chromosome). Stocks are now available with all 10 maize chromosomes individually in an oat background, as well as several hundred radiation hybrids. These genetic materials greatly simplify the study of the maize genome: the oat-maize addition lines reflect a 10-fold reduction in maize genome complexity, and the radiation hybrids, yet a further reduction.
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A major discovery came from the mapping of molecular genetic markers across many species of grasses. Devos et al. (1993) and others realized that members of the same family had similar gene contents and even the same order of genes on chromosomes. This initiated the discipline of comparative genomics; this similarity of members of a taxonomic family at the DNA level was found to be generally true in plants, animals, and microorganisms. The similarity is certainly not perfect, but generally sufficiently frequent that information in one species significantly advances our understanding of related species. The mapping and sequence information has led to increases in evolutionary understanding of crop species (Moore et al., 1995; Devos, 2005), genetic measurements of diversity (Melchinger et al., 1990), molecular identification of disease resistance genes (Hartl et al., 1995; Warren et al., 1999; Gebhardt and Valkonen, 2001), and genetic control of flowering (Salvi et al., 2002), and many other traits.
As molecular genetic markers became available in crop species, an intense interest developed in locating genomic regions that significantly control agronomic traits. These traits are generally controlled by many genes and influenced by the environment. Quantitative genetics theory had indicated that these traits showed polygenic inheritance. Generally, the theory did not lead investigators to prominently consider the presence of major genes, that is, ones controlling a major portion of the phenotypic variation. The QTL analyses designed to find a significant association of a specific genomic region with the trait of interest often identified genomic regions with a major effect. This information led to the development of "marker-assisted selection" (MAS) protocols (Young and Tanksley, 1989; Edwards et al., 1987; Edwards et al., 1992). Tanksley and his students and colleagues pioneered this area and proposed methods to maximize the utility in breeding programs (Frary et al., 2004). Although MAS often has been effective in transferring more simply inherited traits, the method awaits improvements to be universally valuable. In the meantime, however, considerable information has been acquired on the inheritance of a multitude of phenotypic and biochemical traits (Kianian et al., 2000; Groh et al., 2001; Phillips et al., 1993). We can dream of the day when we know the genomic regions controlling important traits up and down every chromosome, much like the human genome (www.ornl.gov/hgmis/posters/chromosome; verified 19 June 2006).
Emergence of Bioinformatics and Genetic Engineering
In concert with evolving DNA sequence information, database and bioinformatic tool development progressed rapidly. These tools gave us the ability to distinguish potential genes from other sequences, compare sequences across all species with the available data, analyze complex and voluminous datapoints, find gene components, and many other important features (Altschul et al., 1990). Bioinformatics is an essential part of crop genetics today and will become even more useful in the future. With DNA sequence information and our ability to extensively analyze the data, the assignment of function to these gene sequences becomes possible. The National Science Foundation initiated a program called Project 2010 with the goal of deciphering the function of every gene in a plant (Arabidopsis) by the year 2010 (www.nsf.gov/publications/pub_summ.jsp?ods_key=nsf04617; verified 19 June 2006). Most of the Arabidopsis genes also are expected to be in crop species. So, it is not unreasonable to expect that we will know the function of most crop species' genes and their chromosome location within the next decade. Plant breeders will likely track minute segments of each chromosome in segregating generations and select traits based on the molecular genetic genotype, and then check phenotypes in appropriate environments. The correlation of a DNA segment and a phenotype is enhanced by knockout or knockdown mutations. One of the recent approaches involves RNA interference, where small double-stranded RNA molecules destroy the mRNA of a specific gene. The description of cosuppression by Jorgensen (1995) working with petunia was interpreted as due to RNA interference, a technology now employed in maize (Springer et al., 2002) and many other species.
All of these technologies play a role in the new field of genetic engineering, together with recombinant DNA technology (Cohen et al., 1973). The ability to isolate specific genes and incorporate them into the cells of regenerable tissue culture (Vieira and Messing, 1987) has led to the development of plants with genes of interest. Plants with the most useful traits can be advanced for commercial use via a conventional breeding program. Today over 200 million acres are planted to these "biotech varieties" (James, 2004); the principal crops are corn (14% of total acreage) and soybeans (60% of total acreage). Many additional biotech varieties are in the pipeline, including golden rice that is high in pro-Vitamin A (Ye et al., 2000; Paine et al., 2005). The potential of golden rice appears great because of the major deficiency of Vitamin A in the developing world causing 500 000 children to go blind every year and leading to other nutritional maladies. Many issues surrounding the use of this technology have been raised and are being evaluated. Time may show that the biosafety emphasis should be on the product and not the process (National Academy of Sciences, 1987; Scriber, 2001). We also must keep in mind that the technology will continue to advance and many of today's concerns likely will be managed in the future. We should exercise due diligence, and learn from careful experimental testing and experience; we do not want to unnecessarily preclude possible major advances. Full societal integration of high impact technology can take up to 50 yr (Wong, 2005). In the future, for example, rice productivity might be enhanced by converting rice from a C3 species to a C4 species and provide a significant advance in productivity. Heterosis might be explained as we gather more genomic information (Fu and Dooner, 2002), and this knowledge may lead to enhancements in the productivity of hybrids in many species.
The future holds many challenges. Genes do not act alone but interact with other genes, perhaps in the same pathway or in a multitude of other pathways. Multiple transcriptional activator genes exist in species that control the transcription of many other genes. Considering that there are perhaps 25 000 to 50 000 genes in a species, this networking of genes will be difficult to decipher due to the sheer number of combinations. On top of this complexity is the influence of the environment on gene expression. Clearly, there is plenty of work to be done! Making this information useful for the improvement of crops will indeed be a challenge.
Germplasm banks have been developed for major crops and maintained by many members of CSSA. Unlocking the potential harbored in the 100 000's of accessions in germplasm banks around the world is important for the future (Hoisington et al., 1999). Today's crop varieties have been adapted to specific environments and fine-tuned for certain planting and growing conditions. When these modern varieties are hybridized to an unimproved accession held in the germplasm bank, half of the resulting hybrid genome is from the improved variety and half is from the accession. Considering that our crops likely contain 25 000 to 50 000 genes, many genes will be different between the two parents. Sorting out the favorable genes from the modern variety and capturing the few genes desired from the germplasm bank accession is a major task, and one that is infrequently accomplished (Frey, 1998). Because these accessions likely have useful complexes of genes, we must find methods to take greater advantage of the goldmine that these accessions represent. More research is needed to apply the technologies of genomics and molecular biology toward the efficient extraction of these useful genetic regions.
Intellectual Property, Biosafety, and Bioethics
Coupled with the developments in genetics has been an explosion in intellectual property rights (Sullivan, 2004) and regulatory processes related to the biosafety of the new technologies (Ellstrand, 2003). Ethical issues have been raised as well by the new developments (Dhanda, 2002). Although the Plant Patent Act was passed by the U.S. Congress in 1930, patent protection was extended to artificially engineered microorganisms via the Chakrabarty decision in 1980. The Plant Variety Protection Act (1970, amended 1994) and the use of utility patents also gave protection to new varieties of crops. These intellectual property provisions allowed biotechnology to become big business and greatly impact agriculture. The potential risks from genetic engineering were presented to the world via the Asilomar Conference in 1975 when 150 molecular biologists met and decided on a moratorium on recombinant DNA research until it was better explored (Berg et al., 1975). Subsequent discussions led to the U.S. Coordinated Framework for regulation of genetically engineered organisms, as published by the Office of Science and Technology Policy (1986), and to extensive discussions and implementation of biosafety protocols around the world (Gogo, 2001). Such powerful technologies that can modify organisms in many different ways have generated broad discussion about bioethics, which will continue to be an important topic in the foreseeable future as science is increasingly applied to agriculture.
What We Don't Know and Future Opportunities
In 2005, on the 125th anniversary of Science magazine, Editors Donald Kennedy and Colin Norman posed 125 questions under the title, "What don't we know"? Several of these questions are of importance to C7 scientists:
This list strongly illustrates the point that many of the basic questions underlying crop science remain to be answered—and these are important questions! Of course, we can add many more questions specifically for crop scientists, such as:
This sample of questions emphasizes the amount of information yet to be obtained or at least exploited. It also shows a glimpse of the exciting careers that lie ahead for young scientists entering the field of crop science.
In spite of the challenges in applying basic genetic information to the improvement of crops, we do have the historical documentation that genetic changes account for about 50% of the improvements in crop productivity in most crops (Fehr, 1984). The other 50% is due to management and efficient use of natural resources. The genetic component of the increase in productivity may be even greater in the future due to the general interest in reducing the use of chemicals in agriculture. This goal coupled with great leaps in genetic knowledge and the ability to manipulate plants using molecular genetic techniques may well raise the percentage to over 50%.
In 1974–75, Maxam and Gilbert (1980) developed a method for DNA sequencing that used four separate reactions to determine the sequence of the nucleotides making up DNA. At the same time, Sanger et al. (1977) developed the chain termination method for sequencing DNA. These technologies revolutionized the field and allowed millions of fragments of DNA to be sequenced. The new sequencing technologies being developed today, such as the 454 sequencing method (Margulies et al., 2005), hold the promise of being able to sequence a genome the size of the human genome for perhaps $1000 (Pennisi, 2005). If that becomes the norm, imagine the information that will be at our fingertips in the future. The great international achievement of sequencing the 430 million base pairs of the rice genome may seem trivial in the future. Even today, we are expecting to sequence at least the gene space of the sorghum genome in a year (National Sorghum Producers, 2005) and the maize genome in two or more years. Just as the sequenced human genome was published concurrent with the 50th anniversary of the double helix discovery, the completed rice genome was published in the same year as the 50th anniversary of CSSA.
One aspect of today's world is clear: the use of genomics, molecular biology and biotechnology to keep food production at the level required to feed a billion more people added to this world every 14 yr will require the entire complement of crop sciences represented by Divisions C-1 through C-8. The C-7 Division stimulates, communicates, and focuses biological discoveries on agriculture worldwide. The success of agriculture in the future will depend on the appropriate application of science and the general understanding of that science by the public. The CSSA will play a major role in agriculture and in the general welfare of people around the world, recognizing the high correlation of conflict and lack of adequate food (Borlaug, 2000). All of the CSSA divisions will play an increasingly important role. A quote from CSSA member and Nobel Peace Prize Laureate Norman Borlaug reflects the need to work together: "No matter how excellent the research done in one specific discipline, its application in isolation will have little positive effect on crop production. What is needed are venturesome scientists who can work across disciplines to produce appropriate technologies and who have the courage to make this case with political leaders to bring advances to fruition."
Received for publication November 8, 2005.
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TOPABSTRACTFOUNDATIONS OF MODERN DAY...REFERENCES |
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