Self-incompatibility (S) alleles of the rosaceae encode members of a distinct class of the T2/S ribonuclease superfamily

Stylar riboncleases (RNases) are associated with gametophytic self-incompatibility in two plant families, the Solanaceae and the Rosaceae. The self-incompatibility-associated RNases (S-RNases) of both the Solanaceae and the Rosaceae were recently reported to belong to the T2 RNase gene family, based on the presence of two well-conserved sequence motifs. Here, the cloning and characterization of S-RNase genes from two species of Rosaceae, apple (Malus × domestica) and Japanese pear (Pyrus serotina) is described and these sequences are compared with those of other T2-type RNases. The S-RNases of apple specifically accumulated in styles following maturation of the flower bud. Two cDNA clones for S-RNases from apple, and PCR clones encoding a further two apple S-RNases as well as two Japanese pear S-RNases were isolated and sequenced. The deduced amino acid sequences of the rosaceous S-RNases contained two conserved regions characteristic of the T2/S-type RNases. The sequences showed a high degree of diversity, with similarities ranging from 60.4% to 69.2%. Interestingly, some interspecific sequence similarities were higher than those within a species, possibly indicating that diversification of S-RNase alleles predated speciation in the Rosaceae. A phylogenetic tree of members of the T2/S-RNase superfamily in plants was obtained. The rosaceous S-RNases formed a new lineage in the tree that was distinct from those of the solanaceous S-RNases and the S-like RNases. The findings suggested that self-incompatibility mechanisms in Rosaceae and Solanaceae are similar but arose independently in the course of evolution.     

Self-incompatibility (S) alleles of the rosaceae encode members of a distinct class of the T2/S ribonuclease superfamily

Stylar riboncleases (RNases) are associated with gametophytic self-incompatibility in two plant families, the Solanaceae and the Rosaceae. The self-incompatibility-associated RNases (S-RNases) of both the Solanaceae and the Rosaceae were recently reported to belong to the T2 RNase gene family, based on the presence of two well-conserved sequence motifs. Here, the cloning and characterization of S-RNase genes from two species of Rosaceae, apple (Malus × domestica) and Japanese pear (Pyrus serotina) is described and these sequences are compared with those of other T2-type RNases. The S-RNases of apple specifically accumulated in styles following maturation of the flower bud. Two cDNA clones for S-RNases from apple, and PCR clones encoding a further two apple S-RNases as well as two Japanese pear S-RNases were isolated and sequenced. The deduced amino acid sequences of the rosaceous S-RNases contained two conserved regions characteristic of the T2/S-type RNases. The sequences showed a high degree of diversity, with similarities ranging from 60.4% to 69.2%. Interestingly, some interspecific sequence similarities were higher than those within a species, possibly indicating that diversification of S-RNase alleles predated speciation in the Rosaceae. A phylogenetic tree of members of the T2/S-RNase superfamily in plants was obtained. The rosaceous S-RNases formed a new lineage in the tree that was distinct from those of the solanaceous S-RNases and the S-like RNases. The findings suggested that self-incompatibility mechanisms in Rosaceae and Solanaceae are similar but arose independently in the course of evolution.     

Allelic diversity of S-RNase at the self-incompatibility locus in natural flowering cherry populations (Prunus lannesiana var. speciosa)

In the Rosaceae family, which includes Prunus, gametophytic self-incompatibility (GSI) is controlled by a single multiallelic locus (S-locus), and the S-locus product expressed in the pistils is a glycoprotein with ribonuclease activity (S-RNase). Two populations of flowering cherry (Prunus lannesiana var. speciosa), located on Hachijo Island in Japan's Izu Islands, were sampled, and S-allele diversity was surveyed based on the sequence polymorphism of S-RNase. A total of seven S-alleles were cloned and sequenced. The S-RNases of flowering cherry showed high homology to those of Prunus cultivars (P. avium and P. dulcis). In the phylogenetic tree, the S-RNases of flowering cherry and other Prunus cultivars formed a distinct group, but they did not form species-specific subgroups. The nucleotide substitution pattern in S-RNases of flowering cherry showed no excess of nonsynonymous substitutions relative to synonymous substitutions. However, the S-RNases of flowering cherry had a higher Ka/Ks ratio than those of other Prunus cultivars, and a subtle heterogeneity in the nucleotide substitution rates was observed among the Prunus species. The S-genotype of each individual was determined by Southern blotting of restriction enzyme-digested genomic DNA, using cDNA for S-RNase as a probe. A total of 22 S-alleles were identified. All individuals examined were heterozygous, as expected under GSI. The allele frequencies were, contrary to the expectation under GSI, significantly unequal. The two populations studied showed a high degree of overlap, with 18 shared alleles. However, the allele frequencies differed considerably between the two populations.     

Development of microsatellite markers in peach [ Prunus persica (L.) Batsch] and their use in genetic diversity analysis in peach and sweet cherry ( Prunus avium L.)

We report the sequence of 41 primer pairs of microsatellites from a CT-enriched genomic library of the peach cultivar 'Merrill O'Henry'. Ten microsatellite-containing clones had sequences similar to plant coding sequences in databases and could be used as markers for known functions. For microsatellites segregating at least in one of the two Prunus F(2) progenies analyzed, it was possible to demonstrate Mendelian inheritance. Microsatellite polymorphism was evaluated in 27 peach and 21 sweet cherry cultivars. All primer pairs gave PCR-amplification products on peach and 33 on cherry (80.5%). Six PCR-amplifications revealed several loci (14.6%) in peach and eight (19.5%) in sweet cherry. Among the 33 single-locus microsatellites amplified in peach and sweet cherry, 13 revealed polymorphism both in peach and cherry, 19 were polymorphic only on peach and one was polymorphic only on cherry. The number of alleles per locus ranged from 1 to 9 for peach and from 1 to 6 on sweet cherry with an average of 4.2 and 2.8 in peach and sweet cherry, respectively. Cross-species amplification was tested within the Prunus species: Prunus avium L. (sweet cherry and mazzard), Prunus cerasus L. (sour cherry), Prunus domestica L. (European plum), Prunus amygdalus Batsch. (almond), Prunus armeniaca L. (apricot), Prunus cerasifera Ehrh. (Myrobalan plum). Plants from other genera of the Rosaceae were also tested: Malus (apple) and Fragaria (strawberry), as well as species not belonging to the Rosaceae: Castanea (chestnut tree), Juglans (walnut tree) and Vitis (grapevine). Six microsatellites gave amplification on all the tested species. Among them, one had an amplified region homologous to sequences encoding a MADS-box protein in Malus x domestica. Twelve microsatellites (29.3%) were amplified in all the Rosaceae species tested and 31 (75.6%) were amplified in all the six Prunus species tested. Thirty three (80.5%), 18 (43.9%) and 13 (31.7%) gave amplification on chestnut tree, grapevine and walnut tree, respectively.     

基于cDNA芯片的梨品种S基因型鉴定及新S-RNase基因进化分析

梨品种S基因型鉴定对梨栽培中授粉品种选择和遗传育种都具有重要意义。本研究利用梨S-RNase基因荧光标记的特异引物PCR扩增获得梨品种荧光标记的cDNA特异产物;进一步完善梨S-RNase基因cDNA芯片,以被检测梨品种cDNA特异序列与梨S-RNase基因cDNA芯片杂交检测不同梨品种S基因型,并发现新的S-RNase基因。结果表明:利用梨S-RNase基因cDNA芯片鉴定了泸定王皮梨、兴山24号、弥渡百合等35个未知S基因型梨品种,确定了各品种的S基因型。结合PCRRFLP及DNA克隆和测序等技术,发现了7个新的S-RNase基因资源,获得了新S-RNase基因序列。序列分析表明各新S-RNase基因均具有S-RNase基因特异区域序列的典型特征;进化分析显示7个新S-RNase基因主要属于蔷薇科苹果亚科S-RNase类群,且存在种间和属间比种内和属内进化关系更近的现象。7个新的S基因分别命名为:PpS_(53)(Pyrus pyrifolia S53)、PpS_(54)、PpS_(55)、PpS_(56)、PpS_(57)、PpS_(58)和PpS_(59),GenBank登录号分别为:KX581753、KX581754、KX581755、KX581756、KX581757、KX581751和KX581752。     

Genetic linkage maps constructed by using an interspecific cross between Japanese and European pears

Genetic linkage maps of the European pear ( Pyrus communis L.) cultivar 'Bartlett' and the Japanese pear ( Pyrus pyrifolia Nakai) cultivar 'Housui' were constructed based on AFLPs, SSRs from pear, apple and Prunus, isozymes and phenotypic traits by using their F(1) progenies. The map of the female parent Bartlett consisted of 226 loci including 175 AFLPs, 49 SSRs, one isozyme and one S locus on 18 linkage groups over a total length of 949 cM, while that for 'Housui' contained 154 loci including 106 AFLPs, 42 SSRs, two phenotypic traits and the other four markers on 17 linkage groups encompassing a genetic distance of 926 cM. These maps were partially aligned using 20 codominant markers which showed segregating alleles in both parents. Compared with the reports of apple genetic maps, these pear maps were not saturated but were near saturation. Distorted segregation was observed in two and one regions of the genome of Bartlett and Housui, respectively. The position of 14 SSRs originating from apple could be successfully determined in pear maps, which enabled us to compare the two maps. Some SSRs developed from Prunus (peach, cherry) were also mapped. The relationships between pear and the other species belonging to the Rosaceae were discussed based on the position of SSRs.     

Related polymorphic F-box protein genes between haplotypes clustering in the BAC contig sequences around the S-RNase of Japanese pear

Most fruit trees in the Rosaceae exhibit self-incompatibility, which is controlled by the pistil S gene, encoding a ribonuclease (S-RNase), and the pollen S gene at the S-locus. The pollen S in Prunus is an F-box protein gene (SLF/SFB) located near the S-RNase, but it has not been identified in Pyrus and Malus. In the Japanese pear, various F-box protein genes (PpSFBB(-α-γ)) linked to the S-RNase are proposed as the pollen S candidate. Two bacterial artificial chromosome (BAC) contigs around the S-RNase genes of Japanese pear were constructed, and 649?kb around S(4)-RNase and 378?kb around S(2)-RNase were sequenced. Six and 10 pollen-specific F-box protein genes (designated as PpSFBB(4-u1-u4, 4-d1-d2) and PpSFBB(2-u1-u5,) (2-d1-d5), respectively) were found, but PpSFBB(4-α-γ) and PpSFBB(2-γ) were absent. The PpSFBB(4) genes showed 66.2-93.1% amino acid identity with the PpSFBB(2) genes, which indicated clustering of related polymorphic F-box protein genes between haplotypes near the S-RNase of the Japanese pear. Phylogenetic analysis classified 36 F-box protein genes of Pyrus and Malus into two major groups (I and II), and also generated gene pairs of PpSFBB genes and PpSFBB/Malus F-box protein genes. Group I consisted of gene pairs with 76.3-94.9% identity, while group II consisted of gene pairs with higher identities (>92%) than group I. This grouping suggests that less polymorphic PpSFBB genes in group II are non-S pollen genes and that the pollen S candidates are included in the group I PpSFBB genes.     

Characterization of the S-RNase promoters from sweet cherry (Prunus avium L.)

Genomic DNA fragments containing the S(3)-, S(4)-, and S(6)-RNase genes were isolated from the sweet cherry (Prunus avium L.) and sequenced. Comparison of the 5'-flanking sequences of these three S-RNases indicated that a highly conserved region (designated CR) existed just upstream from the putative TATA boxes. We postulate that CR contains cis-regulatory element(s) involved in pistil expression. To examine the activity of the isolated S-RNase promoters of sweet cherry in the pistil, we transiently introduced approximately 650-bp fragments of the S(4)- and S(6)-RNase promoters fused to beta-glucuronidase (GUS) gene into the pistil of the petunia using a particle bombardment technique. Histochemical analysis showed that the 5'-flanking region of each S-RNase was active in the pistil. This suggests that cis-regulatory element(s) for pistil-specific expression may exist(s) within the 650-bp region upstream from the TATA box in the sweet cherry S-RNase promoter.     

Host preference of the plum curculio

We assessed host preference of adult plum curculio, Conotrachelus nenuphar (Herbst) (Coleoptera: Curculionidae), based on the total number of mark‐released and wild adults recovered and the total distance moved by mark‐released adults in an orchard whose layout was designed to specifically allow foraging plum curculios to choose among host tree species. Host trees included apple, Malus domestica Borkh.; pear, Pyrus communis (L.); peach, Prunus persica (L.) Batsch; apricot, Prunus armeniaca L.; tart cherry, Prunus cerasus L.; sweet cherry, Prunus avium (L.); European plum, Prunus domestica L.; and Japanese plum, Prunus salicina Lindl. (all Rosaceae). We released 2900 marked adults and recovered 17.7%. We used screen traps to provide a measure of the number of adults that arrived at and climbed up particular host trees and found that significantly greater numbers of marked adults and the greatest number of wild adults were recovered from screen traps attached to Japanese plum. We sampled host tree canopies by tapping limbs to provide a measure of the number of adults within a tree canopy at a particular moment. Again, significantly greater numbers of marked and wild adults were recovered from plum species, with no difference between Japanese and European plum cultivars for marked individuals, but with significantly greater numbers of wild individuals recovered from Japanese plum. The preference index (PI) for Japanese plum based on total distances moved by all marked adults recovered on Japanese plum divided by the total distance moved by marked adults recovered on other host trees indicated that Japanese plum was the most highly preferred host, followed by European plum, peach, sweet cherry, tart cherry, apricot, apple, and pear, respectively.     

Effect of several different pollens on the bio-ecological parameters of the predatory mite Typhlodromus athenas Swirski and Ragusa (Acari: Phytoseiidae)

The development, survivorship, and reproduction of the predacious mite Typhlodromus athenas Swirski and Ragusa were studied in the laboratory by rearing the predator on nine different plant pollens [almond(Prunus amygdalis Batsch), apple (Malus domestica Borkh.), apricot (Prunus armeniaca L.), cherry (Prunus avium L.), pear (Pyrus communis L.), plum (Prunus domestica L.), walnut (Juglans regia L.), olive (Olea europaea L.), Typha sp.], and pollen collected from bee hives. All experiments were conducted in environmental chambers at 20 ± 1°C, 65% RH, and a photoperiod of 16:8 (L:D) h. Survival during immature development ranged from 81.1 to 96.0%. The shortest mean developmental time from egg to adult with respect to the range of pollen species was recorded for females and males fed on almond pollen (10.76 ± 0.18 and 10.45 ± 0.21 d, respectively), while the longest was on beehive pollen (26.97 ± 0.23 and 24.00 ± 0.25 d for females and males, respectively). Female longevity varied from 51.63 ± 5.52 d (olive pollen) to 102.81 ± 6.60 d (pear pollen), while fecundity ranged from 5.33 ± 2.35 eggs per female (beehive pollen) to 26.43 ± 1.73 eggs per female (almond pollen). The diet consisting of almond pollen resulted in the highest intrinsic rate of natural increase (r(m)) (1.00 d(-1)) and pollen collected from bee hives resulted in the lowest (0.013 d(-1)). These results showed that various pollen could favor the development of T. athenas, and also support the view that alternative food resources may play an important role in the field for sustaining and increasing the predator's population.     

Loss of pollen-S function in two self-compatible selections of Prunus avium is associated with deletion/mutation of an S haplotype-specific F-box gene

Recently, an S haplotype-specific F-box (SFB) gene has been proposed as a candidate for the pollen-S specificity gene of RNase-mediated gametophytic self-incompatibility in Prunus (Rosaceae). We have examined two pollen-part mutant haplotypes of sweet cherry (Prunus avium). Both were found to retain the S-RNase, which determines stylar specificity, but one (S3' in JI 2434) has a deletion including the haplotype-specific SFB gene, and the other (S4' in JI 2420) has a frame-shift mutation of the haplotype-specific SFB gene, causing amino acid substitutions and premature termination of the protein. The loss or significant alteration of this highly polymorphic gene and the concomitant loss of pollen self-incompatibility function provides compelling evidence that the SFB gene encodes the pollen specificity component of self-incompatibility in Prunus. These loss-of-function mutations are inconsistent with SFB being the inactivator of non-self S-RNases and indicate the presence of a general inactivation mechanism, with SFB conferring specificity by protecting self S-RNases from inactivation.     

Characteristics and transferability of new apple EST-derived SSRs to other Rosaceae species

Genic microsatellites or simple sequence repeat markers derived from expressed sequence tags (ESTs), referred to as EST–SSRs, are inexpensive to develop, represent transcribed genes, and often have assigned putative function. The large apple (Malus × domestica) EST database (over 300,000 sequences) provides a valuable resource for developing well-characterized DNA molecular markers. In this study, we have investigated the level of transferability of 68 apple EST–SSRs in 50 individual members of the Rosaceae family, representing three genera and 14 species. These representatives included pear (Pyrus communis), apricot (Prunus armeniaca), European plum (P. domestica), Japanese plum (P. salicina), almond (P. dulcis), peach (P. persica), sour cherry (P. cerasus), sweet cherry (P. avium), strawberry (Fragaria vesca, F. moschata, F. virginiana, F. nipponica, and F. pentaphylla), and rose (Rosa hybrida). All 68 primer pairs gave an amplification product when tested on eight apple cultivars, and for most, the genomic DNA-derived amplification product matched the expected size based on EST (in silico) data. When tested across members of the Rosaceae, 75% of these primer pairs produced amplification products. Transferability of apple EST–SSRs across the Rosaceae ranged from 25% in apricot to 59% in the closely related pear. Besides pear, the highest transferability of these apple EST–SSRs, at the genus level, was observed for strawberry and peach/almond, 49 and 38%, respectively. Three markers amplified in at least one genotype within all tested species, while eight additional markers amplified in all species, except for cherry. These 11 markers are deemed good candidates for a widely transferable Rosaceae marker set provided their level of polymorphism is adequate. Overall, these findings suggest that transferability of apple EST–SSRs across Rosaceae is varied, yet valuable, thereby providing additional markers for comparative mapping and for carrying out evolutionary studies.     

Microsatellites isolated in almond from an AC‐repeat enriched library

We have isolated 44 SSRs from an AC‐enriched genomic library from almond (Prunus amygdalus Batsch.). Twenty SSRs were screened for their polymorphism in 16 cultivars and for their transportability in seven different Prunus species (peach, nectarine, apricot, European plum, Japanese plum, sweet cherry, sour cherry) and in apple. The expected heterozygosity ranged from 0.62 to 0.89. About 30% of primers gave successful amplification in seven different Prunus species; in two cases amplifications were obtained also in apple.     

Cloning,expression, and characterization of sorbitol transporters from developing sour cherry fruit and leaf sink tissues

The acyclic polyol sorbitol is a primary photosynthetic product and the principal photosynthetic transport substance in many economically important members of the family Rosaceace (e.g. almond [Prunus dulcis (P. Mill.) D.A. Webber], apple [Malus pumila P. Mill.], cherry [Prunus spp.], peach [Prunus persica L. Batsch], and pear [Pyrus communis]). To understand key steps in long-distance transport and particularly partitioning and accumulation of sorbitol in sink tissues, we have cloned two sorbitol transporter genes (PcSOT1 and PcSOT2) from sour cherry (Prunus cerasus) fruit tissues that accumulate large quantities of sorbitol. Sorbitol uptake activities and other characteristics were measured by heterologous expression of PcSOT1 and PcSOT2 in yeast (Saccharomyces cerevisiae). Both genes encode proton-dependent, sorbitol-specific transporters with similar affinities (K(m) sorbitol of 0.81 mM for PcSOT1 and 0.64 mM for PcSOT2). Analyses of gene expression of these transporters, however, suggest different roles during leaf and fruit development. PcSOT1 is expressed throughout fruit development, but especially when growth and sorbitol accumulation rates are highest. In leaves, PcSOT1 expression is highest in young, expanding tissues, but substantially less in mature leaves. In contrast, PcSOT2 is mainly expressed only early in fruit development and not in leaves. Compositional analyses suggest that transport mediated by PcSOT1 and PcSOT2 plays a major role in sorbitol and dry matter accumulation in sour cherry fruits. Presence of these transporters and the high fruit sorbitol concentrations suggest that there is an apoplastic step during phloem unloading and accumulation in these sink tissues. Expression of PcSOT1 in young leaves before completion of the transition from sink to source is further evidence for a role in determining sink activity.     

Genetic and molecular characterization of three novel S-haplotypes in sour cherry (Prunus cerasus L.)

Tetraploid sour cherry (Prunus cerasus L.) exhibits gametophytic self-incompatibility (GSI) whereby the specificity of self-pollen rejection is controlled by alleles of the stylar and pollen specificity genes, S-RNase and SFB (S haplotype-specific F-box protein gene), respectively. As sour cherry selections can be either self-compatible (SC) or self-incompatible (SI), polyploidy per se does not result in SC. Instead the genotype-dependent loss of SI in sour cherry is due to the accumulation of non-functional S-haplotypes. The presence of two or more non-functional S-haplotypes within sour cherry 2x pollen renders that pollen SC. Two new S-haplotypes from sour cherry, S(33) and S(34), that are presumed to be contributed by the P. fruticosa species parent, the complete S-RNase and SFB sequences of a third S-haplotype, S(35), plus the presence of two previously identified sweet cherry S-haplotypes, S(14) and S(16) are described here. Genetic segregation data demonstrated that the S(16)-, S(33)-, S(34)-, and S(35)-haplotypes present in sour cherry are fully functional. This result is consistent with our previous finding that 'hetero-allelic' pollen is incompatible in sour cherry. Phylogenetic analyses of the SFB and S-RNase sequences from available Prunus species reveal that the relationships among S-haplotypes show no correspondence to known organismal relationships at any taxonomic level within Prunus, indicating that polymorphisms at the S-locus have been maintained throughout the evolution of the genus. Furthermore, the phylogenetic relationships among SFB sequences are generally incongruent with those among S-RNase sequences for the same S-haplotypes. Hypotheses compatible with these results are discussed.     

Primary structural features of rosaceous S-RNases associated with gametophytic self-incompatibility

We isolated cDNA clones encoding five S-RNases (S_1-,_S3- , S_5-, S_6-, S_7-RNases) from pistils of Pyrus pyrifolia (Japanese pear), a member of the Rosaceae. Their amino acid sequences were aligned with those of other rosaceous S-RNases sequenced so far. A total of 76 conserved amino acid residues were stretched throughout the sequence, but were absent from the 51–66 region which was designated the hypervariable (HV) region. The phylogenetic tree of rosaceous S-RNases showed that S-RNase polymorphism predated the divergence of Pyrus and Malus. Pairwise comparison of these S-RNases detected two highly homologous pairs, P. pyrifolia S_1- and S_4-RNases (90.0%) and P. pyrifolia S_3- and S_5-RNases (95.5%). The positions of amino acid substitutions between S_1- and S_4-RNases were spread over the entire region, but in the pair of S_3- and S_5-RNases, amino acid substitutions were found in the 21–90 region including the HV region. The substitutions in this restricted region appear to be sufficient to discriminate between S₃ and S₅ pollen and to trigger the self-incompatible reaction.     

PD1, an S-like RNase gene from a self-incompatible cultivar of almond

 Many flowering plants contain stylar S-RNases that are involved in self-incompatibility and S-like RNases of which the biological function is uncertain. This paper reports the deduced amino acid sequence of an S-like RNase gene (PD1) from the self-incompatible plant Prunus dulcis (almond). The amino acid sequence of PD1, which was derived from cDNA and genomic DNA clones, showed 34–86% identity to acidic plant S-like RNases reported so far, with the highest degree of similarity being to an S-like RNase from Japanese pear (Pyrus pyrifolia). Based on RNA hybridisation experiments it appears that, like for many other S-like RNases, the expression of PD1 is not pistil-specific. Analysis of the genomic structure revealed the presence of three introns, of which one is similar in location to that of the related S-RNase gene from Solanaceae and Rosaceae. At least four bands hybridising to PD1 were found upon Southern hybridisation, suggesting the presence of a multigene family of S-like RNase genes in almond. The putative biological function of PD1 is discussed. Received: 22 November 1999 / Revision received: 18 February 2000 · Accepted: 13 March 2000     

Self-incompatibility alleles in Polish wild pear (Pyrus pyraster (L.) Burgsd.): a preliminary analysis

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 genusPyrus, 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 availableP. 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.