The development of DNA fingerprinting in the 1980s swiftly led to applications in paternity testing and forensic science, however it was soon making inroads in the plant sciences too. As part of a special series of articles on DNA fingerprinting brought together by Investigative Genetics, Hilde Nybom from the Swedish University for Agricultural Sciences, Sweden, Kurt Weising from the University of Kassel, Germany, and Björn Rotter from GenXPro GmbH, Germany, explored how this technique has been applied to botany from its beginnings 30 years ago through to today’s latest advances, as published in their recent review. Here Nybom, Weising and Rotter discuss the impact DNA fingerprinting has had on plant genetics and forensic botany, and what the future holds as this technique continues to develop.
What led to your interest in plant genetics?
BR: I was originally driven by an interest in plant improvement for better yields in arid regions. My first research project was to identify markers for genes involved in drought stress response. After that I developed an interest in molecular genetic techniques in general.
HN: I started as a plant taxonomist but soon realised that I was more interested in the genes responsible for the observed variability than in the classification of the plants. Being able to pursue several DNA fingerprinting projects during my post doc introduced me to molecular techniques.
KW: I started as a molecular plant geneticist in the 1980s, when I wrote my PhD on the topic of chromatin structure of transferred genes in transgenic plants. I then switched to the field of molecular markers in the early 1990s when I did some pioneering work on the establishment and use of DNA fingerprinting technology in plants, especially using methods that are based on microsatellite markers.
What questions does you current research hope to answer?
BR: We now work mostly in classic breeding approaches with the aim of ‘gene identification’, for example to identify beneficial genes from wild-type plants that could be introduced into the gene pool of a high yield crop. Such genes are for example resistance genes to a disease or genes that enable plants to better cope with abiotic stress.
For plants with well described genes, we are trying to identify and understand the biochemical pathways encoded by the identified genes that plants use to cope with stress. Besides that, we are often involved in de novo sequencing projects of as yet unknown plant genomes and functional annotation of the identified genes to related genes from other species.
HN: In my role as an apple breeder, I am especially interested in genes involved in resistance towards various fungal diseases and in various aspects of fruit quality. Unfortunately, very few of these traits are governed by major genes, and the major interest is now directed towards identification of the most important quantitative trait loci (QTL), and how these can be identified and selected for in breeding programs.
KW: In 2000, I became the head of the Plant Molecular Systematics Group in the Department of Biology at the University of Kassel, Germany, now focusing on molecular systematic as well as population genetic analyses (still using DNA fingerprinting like microsatellites and AFLP) in a variety of non-model plants like bromeliads.
With regards to advances in DNA fingerprinting, do you think applications for plants come with a different set of challenges than for animals?
BR, HN, KW: It depends very much on the plant in question. Certain plants with small diploid genomes, like rice, allow researchers to perform whole genome sequencing and consequently gain a direct view of the entire genome with the possibility of observing the breeding process at the highest possible resolution. In contrast, other plants like the octoploid sugarcane, still present a big challenge for breeders. New techniques have however already facilitated breeding tremendously and provide new, high-resolution insights into genomes, even of highly complex genomes.
How has the breakthrough in high-throughput genomic sequencing impacted the field of plant genetics?
BR, HN, KW: These sequencing possibilities clearly represent a new era. We do not know if such an immense technological breakthrough in the life sciences has ever happened in such a short time period before, perhaps when microscopy was invented.
Besides the increasing availability of entire genomes of more and more plants, new techniques allow for comparisons of thousands of plant genomes at a very high resolution, up to a level where entire genomes are compared. This permits association analyses at the highest resolution for the identification of important genes and very accurate phylogenetic analyses.
For breeders working with species of a more restricted economic value or for scientists working with wild plants, genomic sequencing has not had much impact as yet. Thanks to the rapidly decreasing costs we can, however, expect that large steps will be taken in the near future.
How have DNA markers specifically fuelled progress in the field of forensic botany?
BR, HN, KW: Experts in the field told us that currently, simple sequence repeats (SSRs) are typically used to assign plant parts to individual plants (e.g. a leaf found in a suspect´s clothes or car to a plant located at a crime scene) or to find identical genotypes in populations which may consist of vegetatively propagated plants (e.g. Cannabis plantations). DNA markers may also support species identification or chemotype determination for material seized by the police. New sequencing technology will further enlarge the spectrum of possibilities here.
What kind of impact do you see plant DNA fingerprinting having in the future?
BR, HN, KW: Plant DNA fingerprinting was and still is the basis of genetic research for all plants; from the establishment of genetic maps and the identification of genes of interest, to the currently used sorting of DNA contigs in their correct order into genomes using high-throughput sequencing-based fingerprinting techniques. We believe that fingerprinting, especially in the field of plant science, still has a long way to go.
In particular, we believe that a very big impact in breeding will be achieved by the use of a much larger gene pool from natural resources. Using modern high-throughput fingerprinting techniques, beneficial and detrimental alleles can be identified very efficiently and marker assisted breeding will finally leave the ‘academic playground’ and be efficiently applied in breeding programs, even those on a smaller scale. This will hopefully also sustain the notion that natural variation and biodiversity needs to be maintained.
What questions would you like to see answered in the future by molecular genetic techniques that would advance the field of botany?
BR, HN, KW: Once a good knowledge base is established of the diverse repertoire of plant genes – for which decades of research is still required – the next big challenge is to achieve a better understanding of gene regulation, epigenetics and non-coding RNA. Much of the information can and will be achieved by the new sequencing techniques, which will soon be joined by high-throughput proteomics and metabolomics analyses. Finally, the biggest challenge will be to make sense of all of this data.