Development of microsatellite markers in potato and their use in phylogenetic and fingerprinting analyses.

Three genomic libraries were constructed using a mixture of DNA from Solanum phureja Juz. & Buk., and S. chacoense Bitt. Two of the libraries were enriched for ATT and GT repeats (a 27-fold enrichment was achieved). In total, 3500 clones of the conventional library, 1,000 of the library enriched for ATT, and 12,000 of the one enriched for GT were screened with five different repeat motifs, and a total of 18 primer pairs was obtained. Another group of 12 primer pairs was obtained from the SSR-containing sequences in the public databases (18 SSR-containing sequences were utilized). From among 30 newly developed primer pairs, 12 previously published ones, and 12 pairs developed for tomato, 7 were used to identify 12 different potato cultivars and introductions, and 12 were used to study phylogenetic distance among seven wild and cultivated potato species. Two SSR markers were sufficient to discriminate the 12 cultivars. The mean number of alleles per polymorphic locus was 5 for the 12 cultivars and 4.5 for the seven species. The results obtained in this study confirm those achieved in similar studies in other plant species regarding the abundance and use of SSR markers in identifying species and cultivars.

Analysis of the genetic composition of anther-derived potato by randomly amplified polymorphic DNA and simple sequence repeats.

Randomly amplified polymorphic DNA (RAPD) and simple sequence repeat (SSR) analyses were used to characterize the genetic composition of anther-derived plants of a diploid potato clone, CP2 (Solanum chacoense 80-1 x S. phureja 1-3). The ploidy of anther-derived plants was first determined by flow cytometry. A total of 44 decamer primers was screened for RAPD polymorphism. The loci that segregated were selected and scored. The monoploids had less than half as many loci carrying RAPD markers compared with the anther donor. Among 14 anther-derived diploids, 5 were identified as homozygous by marker frequency similar to monoploids and 9 as heterozygous. Five of seven SSRs obtained from published potato sequences were polymorphic in CP2. CP2 was found to be heterozygous with two alleles at four SSR loci (TC/TA, AAG, AGA, CTT) and three alleles at a ACTC locus. Primer pairs flanking each of the five polymorphic SSRs revealed that monoploids had only the allele contributed by S. chacoense 80-1. Homozygous diploids had only one band per SSR locus, whereas heterozygous diploids displayed more than one allele for at least one SSR locus. Results of the SSR analysis supported the findings based on RAPD markers; the same five diploid clones were characterized as homozygous by both SSR and RAPD markers.

Variation for 2n pollen production in clones of Solanum phureja Juz. and Buk.

Frequency of unreduced pollen grains was estimated for five genotypes of Solanum phureja (2n=24) growing in three environments; (E1) cool (7.2-13.3°C) and (E2) warm (12.2-17.2°C) growth chambers and (E3) field conditions. Highly variable frequencies were found, with genotype, environment, and genotype x environment interaction as significant components of variance. The frequency of unreduced gametes for two additional genotypes was studied over time in two growth chamber environments (cool and warm). One genotype, characterized by mostly fused spindles at the second meiotic division, expressed a high frequency of big pollen (BP) in both environments, whereas the second, characterized by fused, parallel and tripolar second division spindless was found to increase in BP frequency over time in the cool chamber, but remained consistently low in the warm chamber. The identification of specific environmental components with general effect on the expression of un-reduced gametes is not possible because of the large genotype x environment interaction component of variance. A genetic hypothesis based on incomplete penetrance and variable expressivity of a dominant gene is presented as an alternative to the currently accepted theory of control of parallel spindles by a single recessive gene.