Detection and analysis of genetic variation can help us to understand the molecular basis of various biological phenomena in plants. Since the entire plant kingdom cannot be covered under sequencing projects, molecular markers and their correlation to phenotypes provide us with requisite landmarks for elucidation of genetic variation. Genetic or DNA based marker techniques such as RFLP (restriction fragment length polymorphism), RAPD (random amplified polymorphic DNA), SSR (simple sequence repeats) and AFLP (amplified fragment length polymorphism) are routinely being used in ecological, evolutionary, taxonomical, phylogenic and genetic studies of plant sciences. These techniques are well established and their advantages as well as limitations have been realized. In recent years, a new class of advanced techniques has emerged, primarily derived from combination of earlier basic techniques. Advanced marker techniques tend to amalgamate advantageous features of several basic techniques. The newer methods also incorporate modifications in the methodology of basic techniques to increase the sensitivity and resolution to detect genetic discontinuity and distinctiveness. The advanced marker techniques also utilize newer class of DNA elements such as retrotransposons, mitochondrial and chloroplast based microsatellites, thereby revealing genetic variation through increased genome coverage. Techniques such as RAPD and AFLP are also being applied to cDNA-based templates to study patterns of gene expression and uncover the genetic basis of biological responses. The review details account of techniques used in identification of markers and their applicability in plant sciences.

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Advances in molecular marker techniques and their applications in plant sciences

Comparative genomics, the study of similarities and differences in structure and function of the hereditary information in different taxa, uses molecular tools to investigate notions that far preceded the discovery that DNA was the hereditary molecule. Vavilov’s (1922) “law of homologous series in variation” was an early suggestion of the possibility of underlying commonality in the genetic blueprints of different (plant) species. In plants, genetic analysis based upon morphological and isozyme markers provided early hints that the arrangements of genes along the chromosomes of various taxa may have retained parallels since their divergence from common ancestors. DNA-level investigations in diverse taxa point to two broad messages: 

(1)

The small but essential portion of most plant genomes encoding genes evolves relatively slowly, with corresponding genes retaining recognizable DNA sequences and similar order along the chromosomes of taxa that have been reproductively-isolated for millions of years.




(2)

A wide range of factors, such as DNA sequence mobility, gene deletion, and localized rearrangements, are superimposed on the relatively slow tempo of chromosomal evolution and cause many deviations from co-linearity.




(3)

Genetic loci that account for common phenotypes in different taxa are often at corresponding genomic locations, and may represent orthologous genes or members of orthologous clusters of genes.

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Comparative Genomics of Plant Chromosomes

Faba bean (Vicia faba L.)

We have evaluated three DNA-based marker types for linkage map construction in Populus: RFLPs detected by Southern blot hybridization, STSs detected by a combination of PCR and RFLP analysis, and RAPDs. The mapping pedigree consists of three generations, with the F1 produced by interspecific hybridization between a P. trichocarpa female and a P. deltoides male. The F2 generation was made by inbreeding to the maximum degree permitted by the dioecious mating system of Populus. The applicability of STSs and RAPDs outside the mapping pedigree has been investigated, showing that these PCR-based marker systems are well-suited to breeding designs involving interspecific hybridization. A Populus genome map (343 markers) has been constructed from a combination of all three types. The length of the Populus genome is estimated to be 2400–2800 cM.

Molecular genetics of growth and development in Populus. III. A genetic linkage map of a hybrid poplar composed of RFLP, STS, and RAPD markers.

Linkage maps for two apple clones, White Angel and Rome Beauty, were constructed using isozyme and DNA polymorphisms segregating in a population produced from a Rome Beauty x White Angel cross. The linkage map for White Angel consists of 253 markers arranged in 24 linkage groups and extends over 950 cM. The Rome Beauty map contains 156 markers on 21 linkage groups. The White Angel map was taken as the standard, and we were able to identify linkage groups in Rome Beauty homologous to 13 White Angel linkage groups. The location of several genes not segregating in the Rome Beauty x White Angel population could be determined on the basis of known linkages with segregating markers. Hence, the standard map for apple now contains about 360 markers, with most linkage groups saturated at 10-15 cM. The double pseudotestcross format of the mapping population permitted the comparison of recombination frequencies in male and female parents in certain regions of the genome where appropriate markers were available. The recombination frequencies observed for the approximately 170 cM that were comparable gave no indication that a sex-related difference in recombination rate was characteristic of apple.

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Molecular Marker Linkage Map for Apple

Segregating allozyme and DNA polymorphisms were used to construct a preliminary linkage map for faba bean. Two F2 populations were analyzed, the most informative of which was segregating for 66 markers. Eleven independently assorting linkage groups were identified in this population. One of the groups contained the 45s ribosomal array and could be assigned to the large metacentric chromosome I on which the nucleolar organizer region is located. This linkage group also contained two isozyme loci, Est and Tpi-p, suggesting that it may share some homology with chromosome 4 of garden pea on which three similar markers are syntenic. Additional aspects of the map and the extent of coverage of the total nuclear genome are discussed.

A. M. Torres

Papers

Linkage among isozyme, RFLP and RAPD markers in Vicia faba