PCR-based markers for assessing genetic diversity in plants

Xiao-Ru Wang and Alfred E. Szmidt

In: Proc. Symposium on Measuring and Monitoring Biodiversity in Tropical and Temperate Forests. August 28 - September 3, 1994, Chiang Mai, Thailand.

For reprint, send e-mail to: Alfred E. Szmidt

Abstract

Accurate understanding of the organization of the existing genetic diversity and the factors shaping it is the prerequisite for devising any reasonable gene conservation programme. Different types of genetic markers have been applied to describe and to measure the existing diversity. However, insights into many aspects of plant genetic system are limited by various shortcomings of the available markers. To date, allozyme electrophoresis has been the most widely used technique to study the genetic diversity of plant populations. The main advantages of allozyme markers are the ease of screening of many individuals at a relatively low cost. However, allozymes reveal only limited portion of the existing genetic variation. DNA based markers appear more appropriate in this respect. In contrast to allozyme markers, any coding or non-coding portion of the genome can be sampled with DNA markers and used for genetic analysis. Among different types of DNA markers, RFLPs have been most widely used, owing to clearly defined variants and genomic origin. The limiting factors for their application in population studies are mainly the cost, time, and the availability of suitable probes. Recently introduced, PCR technique have furnished a wide array of new DNA markers that in several respects are superior to RFLPs. The PCR-based markers require very small amounts of DNA, are technically simpler, cheaper, and less labour intensive than RFLPs. The speed and convenience of PCR reaction permits screening of a larger number of individuals than is possible with other types of DNA markers. Furthermore, unlike allozyme markers for which optimal protocols must be developed separately for each species, standard PCR protocols are readily applicable to virtually any organism. The kind of information revealed by this method chiefly depends on the choice of primer(s). Random priming (RAPD) is a quick and convenient approach for fast screening of nuclear genome variation (see Figure 1). In combination with better separation methods, random priming can also provide the fingerprinting resolution - DNA amplification fingerprinting (DAF). Genetic information contained within the cytoplasmic genomes of higher plants is distinct from that of the nuclear genome. Variation of each of these genomes can be analyzed by employing genome-specific primers. Alignment of known gene sequences from various organisms enables construction of consensus primers that can be applied to other species for which the sequence data is not available. For instance, in combination with restriction fragment analysis, we are able to use this approach for studies chloroplast and mitochondrial gene variation in a wide range of plant species (see Figure 2 and Figure 3 for examples). Fast accumulation of sequence data warrants that soon many additional nuclear and cytoplasmic genes can be studied in a similar way.

by Alfred E. Szmidt