Wide genome physical mapping is becoming a main goal of current genomic research on many plant and animal species. It provides essential basis for large-scale genome sequencing, but requires well-saturated large-insert DNA genomic libraries. This review presents a progress in the construction and application of genomic libraries. New trends towards the improvement of cloning vectors and adjustment to new tasks of molecular genetics are also presented.
Wild pear (Pyrus pyraster, syn. P. communis var. pyraster) is thought to be one of the species that gave rise to all other members of the genus Pyrus, although intraspecific hybridizations with cultivated varieties could cause the disappearance of original species characteristics. S-RNase alleles from 7 different wild pear individuals, collected from various regions of Poland, were cloned on the basis of the PCR method and nucleotide sequence analyses. The hypervariable (HV) region is responsible for allele-specific S-RNase activity in the self-incompatibility mechanism. The high level of polymorphism of its sequences may constitute a source of valuable phylogenetic information. From all individuals, 14 sequences were obtained successfully, and 9 of them were novel alleles. Phylogenetic analysis of these alleles was based on the amino acid sequence interpretation of coding regions and intron nucleotide sequences. The research conducted on a limited pool of available P. pyraster alleles gives only an initial insight into possible S-RNase allele polymorphisms in wild populations. At this stage, the results do not confirm a strong influence of cultivated pear species on the wild pear.
The research was aimed to elaborate a method for selection of male plants (XY, YY) and female ones (XX) as well as for identification of supermale genotypes (YY) among male phenotypes. The population obtained by self-pollination of andromonoecious plants was analysed. In order to identify the bands differentiating the male from the female genotypes, Bulk Segregant Analysis (BSA) was carried out. Primers identified by BSA analysis were used for RAPD amplification on the template of the male and female individuals. Among the products obtained by the use of primer OPB-20, some bands were linked with sex. A band of about 700 bp was found in all female plants, and in 4 phenotypically male specimens. In the male plants, the band showed a much lower intensity, compared with the female specimens. It seems that this fragment can be linked to the X chromosome in the investigated specimens. In the female specimens with XX karyotype, template duplication occurs and hence the band intensity is twice as high as in the XY karyotype. Three male plants did not include the OPB-20-700 fragment so they could potentially have the supermale (YY) karyotype. If the obtained marker proved its usefulness for identification of supermale plants, it could become a valuable tool facilitating breeding work.
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