Analysis of anther-derived plants of Solanum phureja: variation in ploidy, photosynthetic efficiency and structure of the nuclear genome
The ultimate goal of the· breeding scheme, of which the present study is a part, is to introduce exotic germplasm into existing cultivars of Solanum tuberosum through· 4x X 2x crosses using the South American diploid potato species Solanum phureja as the pollen parent. The first phase of this program includes the 'reconstruction' of a highly heterozygous diploid, pollen parent by .first : reducing the chromosome number of the S. phureja clones to the monoploid level and subsequently fusing genomes of two unrelated monoploid plants either by somatic hybridization or by cross-pollination between fertile doubled monoploids.
Within this framework, the objectives of this research were to analyze variation among anther-derived plants of Solanum phureja regarding their: 1) ploidy level and morphology, 2) net photosynthesis and its biochemical components, and 3) nuclear genomic structure, particularly with regard ·to the amplification of rRNA genes as influenced by the anther-culture process.
Based on the analysis of several morphological characters of the anther-derived plants by canonical discriminant function, four characters (anther length, number of chloroplasts/pair of guard cells, leaf width, corolla width) were selected for most effective assignment of plants to their ploidy groups by clustering procedures. Clustering of the anther-derived plants proved to be an efficient means of separating monoploids from higher ploidy levels.
To assess the impact of the process of anther-culture on the physiology of the resulting plants and to evaluate the possibility of selecting anther-derived genotypes for further breeding efforts, monoploid, diploid and tetraploid anther-derived plants were studied regarding their net photosynthesis and its component characteristics. Leaf area, net photosynthesis and chlorophyll content increased with increasing ploidy' Among .the. monoploids I. Rubisco activity and concentration displayed a. significant genotypic effect; whereas in the diploid group variation among genotypes was significant for total protein content and maximum specific activity of ribulose bisphosphate carboxylase. Among the tetraploid genotypes, significant differenc.es were found with respect to net photosynthesis and specific leaf weight. Two exceptional genotypes were identified: a monoploid with an increase of 28% fcfr maximum activity of ribulose bisphosphate carboxylase and a tetraploid with an increase of 30% for net photosynthesis over the anther-donor plant.
To evaluate DNA variation among the anther-derived plants, the nuclear genomes of anther-derived monoploid and diploid plant were studied by DNA reassociation kinetics. It was found that the nuclear genome of the monoploid has undergone differential replication resulting in an increase of sequences consisting of highly· repetitive DNA. Free solution RNA-DNA hybridization showed that the monoploid DNA contained 30% more rDNA sequences than the diploid. Southern blot analysis using rRNA as the probe revealed variation for copy number of certain restriction fragments and for restriction enzyme cleavage sites.