INTRODUCTION
Genetic diversity is the basis of the ability of organisms to adapt to changes in their environment through natural selection. Populations with little genetic variation are more vulnerable to the arrival of new pests or diseases, pollution, changes in climate and habitat destruction due to human activities or other catastrophic events. The inability to adapt to changing conditions greatly increases the risk of extinction. Gene conservation management aimed to save adaptive genetic diversity should be based on the knowledge of the genetic basis of adaptation.
This note is extracted from a new FAO Working Paper. The goal of the Working Paper is to briefly describe how adaptive genetic diversity can be measured using new molecular genetic approaches and achievements in forest genomics. The summary of the conclusions is presented below:
SUMMARY OF FINDINGS & CONCLUSIONS
The study of adaptation is fundamental to forestry and forest genetic conservation. Forest geneticists have long used field experiments (common-garden experiments) and, to a lesser extent, molecular markers to study patterns of adaptation in forest trees. Field experiments are means to estimate genetic parameters of measurable traits, but they can neither provide information on which particular genes and how many of them are involved in adaptation, nor how much of phenotypic variation can be explained by genetic variation in these genes.
Another, and generally complementary, approach for estimating adaptive genetic diversity is to measure genetic variation using molecular genetic markers. There are numerous molecular marker technologies available, but most measure either neutral or highly conservative genetic variation of limited adaptive value. There is a need for developing rapid and informative diagnostic techniques for evaluating large numbers of adaptive genes and genetic variation for in situ conservation. Genomics provides new tools to study adaptation in trees. Forest geneticists can use automated, efficient and fast technologies to identify DNA sequences and to determine the genotype of a large number of individuals. They can ultimately identify genes responsible for forest tree adaptation via quantitative trait loci (QTL) and candidate gene mapping using expressed sequence tag (EST) markers for adaptive genes that are expressed in the genome. Then, using modern genotyping technologies and association studies geneticists can determine allelic diversity for these candidate genes in forest tree populations and directly measure adaptive allelic diversity for thousands of genes simultaneously.
Despite remarkable progress much work remains to be done to understand the nature of genetic variation that underlies adaptive forest tree phenotypes. Comprehensive understanding will first require discovering, annotating and cataloguing all genes in the forest tree genome. A modern approach towards achieving this goal is to determine the DNA sequence of the entire genome and infer the genes from the DNA sequence, although a complete sequence alone is not sufficient to understand the genetic control of adaptive traits. Added to the challenges in forestry are the complexity and large size of tree genomes. The size of the pine genome (20,000-30,000 million nucleotide base pairs (bp)), for example, is 6 to 8 times larger than the human genome (3,400 million bp), and 150 to 200 times larger than the genome of the model plant species, Arabidopsis thaliana (125 million bp). Even the relatively small physical size of the Populus genome (500 million bp), which is 40 times smaller than the best-studied conifer, Pinus taeda, and, therefore, can be a good forest tree model species, is still about 4 times as large as that of Arabidopsis (although similar to rice and 6 times smaller than maize, both of which are almost completely sequenced). Moreover, adaptive traits are usually very complex, have quantitative inheritance and are controlled by many genes each with relatively small effects.
An alternative (or parallel) approach is to determine the DNA sequences for the expressed genes only. This can be accomplished by isolating messenger RNA (mRNA), preparing complementary DNA (cDNA) libraries from this mRNA and sequencing cDNA. These EST sequences are submitted to databases and compared to all other sequences in the databases to see if they match to genes whose function has been determined. EST databases of tens of thousands of ESTs have been already produced and are publicly available for Pinus, Picea, Populus, and Eucalyptus.
The second step towards understanding adaptation involves construction of genome, QTL, comparative and consensus linkage maps for most forest tree species (e.g., Sewell et al. 1999). Genetic maps show the position of genes and are valuable for understanding genome organization and evolution. Maps are extremely useful tools for identifying genes controlling interesting phenotypes. Loci controlling quantitatively inherited traits, QTLs, have been already identified in many forest trees for a variety of growth, wood quality, and other economic and adaptive traits. These data are immediately useful for tree improvement and gene conservation.
Next, DNA microarray analysis can be used to study the expression patterns of genes, and to understand the function of all genes and their interactions. The relationship between the vast amount of allelic diversity in genes and the array of different phenotypes found in forest tree populations can be studied. A catalogue of common coding-sequence variants in forest tree genes can be created and tested for association with a phenotype. Genome-wide high-resolution maps of known polymorphisms can be used to scan the genome for marker-adaptive trait associations.
The analysis needs not be limited to coding sequences. It may be that the majority of relevant mutations reside in regulatory regions. Thus, it is important to identify variants in at least the proximal and distal regulatory sequences as our poor understanding of ‘regulatory’ elements dictates the need for a more global approach. An approach in which marker-trait associations are sought will require the construction of a high-resolution map of genetic variants. Single nucleotide polymorphism (SNPs) are the natural candidates for this map because they are abundant, have a smaller mutation rate than microsatellites and can be genotyped en masse using microarray technology.
A map-based association search for multiple adaptive loci, each contributing to the total phenotype in a small yet measurable way, is feasible via haplotype analysis. The alleles of these loci can be indirectly recognized by their historical associations with other genetic variants (e.g. SNPs) in their neighbourhood. The non-random association of variants with one another (linkage disequilibrium) is a well-known feature of the plant and animal genomes. DNA microarrays will have a major role in genotyping thousands of genes simultaneously, in the creation of fine maps and in mapping the components of complex adaptive phenotypes. Forest genomics has a bright future and awaits exiting applications in forest tree management and gene conservation.
SELECTED REFERENCES:
Boshier, D.H and A.G. Young (2000) Forest conservation genetics: limitations and future directions. In: Forest conservation genetics: Principles and practice (A. Young, D. Boshier and T. Boyle, eds), pp. 289-297. CABI Publishing, United Kingdom.
Mandal, A.K. and G.L. Gibson (eds.) (1998) Forest genetics and tree breeding. CBS Publishers, New Delhi, India.
Sewell, M.M. and D.B. Neale (2000) Mapping quantitative traits in forest trees. In: Molecular biology of woody plants. Forestry Sciences, Volume 64 (S. M. Jain and S. C. Minocha, eds), pp. 407-423. Kluwer Academic Publishers, The Netherlands.
Young, A., D. Boshier and T. Boyle (eds) (2000) Forest Conservation Genetics: Principles and Practice. CABI Publishing, United Kingdom.
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THE PRESENT NOTE IS EXTRACTED FROM THE FOLLOWING DOCUMENT: Krutovskii, K.V. and Neale, D.B. 2001. Forest Genomics for Conserving Adaptive Genetic Diversity. Forest Genetic Resources Working Papers, Working Paper FGR/3, Forest Resources Development Service, Forest Resources Division. FAO, Rome (published in English only, will later be available at FAO Homepage). The working paper is based on a lecture given by Dr. Krutovskii, K.V., entitled: Forest genomics and new molecular genetic approaches to measuring and conserving adaptive genetic diversity in forest trees. Presented at a training workshop organized by the International Plant Genetic Resources Institute (IPGRI), Rome and the Austrian Federal Ministry of Agriculture and Forestry, Environment and Water Management (BMLFUW), in technical collaboration with FAO. The training workshop was held in Gmunden, Austria, 30 April to 11 May 2001. The longer version of the document, available free of charge from FAO, Rome, includes the following main sections: 1 Introduction- Why it is important to measure and save adaptive genetic diversity in forest tree populations2. How Forest Genetic Conservation Can Benefit From New Achievements in Genomics
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