Gene flow in cherry orchards

A survey of Australian cherry orchards identified the cherry cultivars Sunburst, Summit, Merchant, Sam, Sylvia, Tieton, Kordia, Regina, Empress, Nordwunder and Ulster as having low fruit-set associated with poor pollination. Unique orchard sites across Australia where low fruit-set was not a problem for these cultivars were located, and pollen gene-flow-analysis conducted using 6-PGD, GOT, G6PD, GPI, IDH, FDP and SKDH isozyme markers. Pollenisers for the above-mentioned cultivars were determined and Stella was a polleniser for eight of them. Stella's predominance was linked to it reaching full bloom before the other cultivars; anther dehiscence occurs sometime after flowers open, as such newly opened flowers; the most-fertile stage was mainly exposed to Stella pollen. Sunburst a self-compatible cultivar showed no evidence of self-fertilisation. The majority of pollenisers were found to be within 20 m of the tree under examination. Isozyme profiles for 22 cultivars not previously analysed are also presented.Communicated by H.F. Linskens     

Wild and Rare Self-Incompatibility Allele S17 Found in 24 Sweet Cherry (Prunus avium L.) Cultivars

The pollination of self-incompatible diploid sweet cherry is determined by the S-locus alleles. We resolved the S-alleles of 50 sweet cherry cultivars grown in Estonia and determined their incompatibility groups, which were previously unknown for most of the tested cultivars. We used consensus primers SI-19/20, SI-31/32, PaConsI, and PaConsII followed by allele-specific primers and sequencing to identify sweet cherry S-genotypes. Surprisingly, 48% (24/50) of the tested cultivars, including 17 Estonian cultivars, carry the rare S-allele S₁₇, which had initially been described in wild sweet cherries in Belgium and Germany. The S₁₇-allele in Estonian cultivars could originate from ‘Leningradskaya tchernaya’ (S₆|S₁₇), which has been extensively used in Estonian sweet cherry breeding. Four studied cultivars carrying S₁₇ are partly self-compatible, whereas the other 20 cultivars with S₁₇ have not been reported to be self-compatible. The recommended pollinator of seven self-incompatible sweet cherries is of the same S-genotype, including four with S₁₇-allele, suggesting heritable reduced effectiveness of self-infertility. We classified the newly genotyped sweet cherry cultivars into 15 known incompatibility groups, and we proposed four new incompatibility groups, 64–67, for S-locus genotypes S₃|S₁₇, S₄|S₁₇, S₅|S₁₇, and S₆|S₁₇, respectively, which makes them excellent pollinators all across Europe. Alternatively, the frequency of S₁₇ might be underestimated in Eastern European populations and some currently unidentified sweet cherry S-alleles might potentially be S₁₇.

     

Inheritance and linkage of isozymes in sweet cherry (Prunus avium L.)

Eight polymorphic isozyme loci, 6PGD, G6PD, MDH, PGM, SKDH, FDP, GOT and IDH, in sweet cherry where found to be in one linkage group, with a ninth isozyme locus, GPI, being in another linkage group on a different chromosome. Isozymes were also linked to the incompatibility S locus and this explained the disturbed segregation ratios observed in the first generation from controlled hybridisations between different sweet cherry cultivars. Analysis revealed close linkage between the isozyme and S loci. The results supported a pre-existing theory that the S gene in cherry consists of three linked segments each coding for a different function. Progeny derived from selfing of Stella, the self-fertile cherry cultivar, also showed disturbed segregation ratios and an absence of homozygotes for the isozyme loci assayed. This demonstrated that codominant inheritance of the S alleles had not been effected by the self-fertile mutation.     

Molecular characterisation of sweet cherry (Prunus avium L.) genotypes using peach [Prunus persica (L.) Batsch] SSR sequences

A total of 76 sweet cherry genotypes were screened with 34 microsatellite primer pairs previously developed in peach. Amplification of SSR loci was obtained for 24 of the microsatellite primer pairs, and 14 of them produced polymorphic amplification patterns. On the basis of polymorphism and quality of amplification, a set of nine primer pairs and the resulting 27 informative alleles were used to identify 72 genotype profiles. Of these, 68 correspond to unique cultivar genotypes, and the remaining four correspond to three cultivars that could not be differentiated from the two original genotypes of which they are mutants, and two very closely related cultivars. The mean number of alleles per locus was 3.7 while the mean heterozygosity over the nine polymorphic loci averaged 0.49. The results demonstrate the usefulness of cross-species transferability of microsatellite sequences allowing the discrimination of different genotypes of a fruit tree species with sequences developed in other species of the same genus. UPGMA cluster analysis of the similarity data divided the ancient genotypes studied into two fairly well-defined groups that reflect their geographic origin, one with genotypes originating in southern Europe and the other with the genotypes from northern Europe and North America.     

Determination of self-incompatibility groups of sweet cherry genotypes from Turkey

Determination of S-allele combinations of sweet cherry genotypes and cultivars has importance for both growers and breeders. We determined S-allele combinations of 40 local Turkish sweet cherry genotypes using a PCR-based method. Ten different S-alleles were detected. Although the most common S-allele was S3, as also found in Western genotypes and cultivars, there were some differences in the frequencies of some S-alleles between Turkish and Western sweet cherry genotypes. According to their S-allele compositions, 30 local Turkish sweet cherry genotypes were assigned to 10 previously identified incompatibility groups. For the remaining genotypes, whose S-allele combinations did not fit to any previous incompatibility groups, three more incompatibility groups, XLII, XLIII and XLIV, were proposed. Results obtained from this study will help both sweet cherry growers and breeders to better manage these local Turkish sweet cherry genotypes in their orchards.     

Pollen performance as affected by the pistilar genotype in sweet cherry (Prunus avium L.)

Summary Differences in pollen performance in higher plants can result in significant selective advantages for some particular genotypes leading to both gametophytic and sexual selection. However, the possibility of selection among male gametophytes has been questioned since natural selection could lead to the fixation of alleles for the best competing male genotypes. These two apparently conflicting hypotheses could be reconciled if pollen performance, rather than operating in absolute terms, could be modulated by the pistilar genotype. Thus, pollen performance in vivo and in vitro has been compared in four sweet cherry (Primus avium L.) cultivars. Differences among the cultivars studied have been recorded in the speed and final pollen germination percentages both in vivo and in vitro. The results obtained show that the female genotype also modulates the final result of pollen performance. These two factors are not merely additive but, on the contrary, the interaction between them affects pollen behavior in vivo. This fact has clear implications for gametophytic and sexual selection since the best male-female combinations can be favored and this could explain the variability observed for pollen performance in nature.     

24份甜樱桃材料自交不亲和S基因型鉴定

甜樱桃品种绝大部分自交不亲和,限制了甜樱桃的正确评价和合理利用,因此自交不亲和基因型的鉴定对于生产具有重要意义。以24个甜樱桃主栽品种为材料,用5对蔷薇科李属引物组合对24个甜樱桃品种进行了S等位基因的PCR扩增,克隆S基因的扩增片段,用核酸序列在Gen Bank上搜索,确定了5种S基因的核酸序列和大小。结果表明:Pru C2+Pru C4R引物组合扩增效果最好;在琼脂糖凝胶上位置相同的扩增带其核酸序列相同,是同一种S基因;5种S基因扩增片段的大小分别是S1为800 bp,S3为762 bp,S4为962bp,S5为300 bp,S6为456 bp,S9为650 bp;24个甜樱桃S基因型是红手球、早红宝石为S1S3,拉宾斯S1S4',红宝石S1S6,布鲁克斯S1S9,那翁S3S4,秦林、泰安大紫、先锋、早大果、丽珠、美早、5-106、左滕锦、桑提娜为S3S6,黑珍珠、红灯、萨米脱、秦樱为S3S9,胜利为S5S9,明珠、红蜜、雷尼、滨库为S6S9。     

Genetic and molecular analysis in Cristobalina sweet cherry,a spontaneous self-compatible mutant

Self-compatibility in a naturally self-incompatible species like sweet cherry is a highly interesting trait for breeding purposes and a powerful tool with which to investigate the basis of the self-incompatible reaction in gametophytic systems. However, natural self-compatibility in sweet cherry is a very rare phenomenon. Cristobalina is a local Spanish sweet cherry cultivar that has proven to be spontaneously self-compatible. In this work, the nature of the self-compatibility in Cristobalina has been studied using genetic and molecular approaches. Pollination studies and microscopic observations of pollen tube growth were carried out to confirm the self-compatible character and the results obtained indicate that self-compatibility is caused by a failure of the pollen and not the style factor. Polymerase chain reaction (PCR) analysis of progenies derived from Cristobalina revealed that self-compatibility in this genotype is not related uniquely to one of the two pollen S alleles, but that pollen grains carrying either of the two haplotypes can overcome the incompatibility barrier. Moreover, PCR analysis and microscopic observation of pollen tube growth in progeny derived from Cristobalina also confirmed that the self-compatible descendants can carry either of the two S haplotypes of their progenitor. Isolation and sequencing of the style S-RNases and pollen SFBs revealed that the DNA sequences of these factors are the same as those described in other self-incompatible sweet cherry cultivars with the same S alleles. Possible mechanisms to explain self-compatibility in Cristobalina are discussed.     

中国主栽樱花的分子系统发育分析

以88份樱花为试验材料,利用核基因ITS和叶绿体间隔序列trnH-psbA进行测序,经过软件MEGA 6.0处理后,构建了系统发育树。结果表明,山樱群樱花品种系统发育比较混乱,这与其遗传背景比较复杂以及原始亲本系统发育关系较近有关,江户彼岸群和寒绯樱群的大多数品种能够聚类在一起,与山樱群品种能分得开,但涉及到很多杂交品种,ITS和trnH-psbA难以区分,表明DNA条形码技术难以应用到杂交品种的系统发育关系,此外,还可依据某些樱花品种特异的ITS或trnH-psbA序列长度对部分品种进行分子鉴定,也对部分种系不明的品种进行了修订。     

ISSR-PCR fingerprinting of Ukrainian sweet cherry (<Emphasis Type="Italic">Prunus avium</Emphasis> L.) cultivars

ISSR-PCR markers were used to assess genetic diversity and to elucidate relatedness among 21 Ukrainian and three West-European sweet cherry cultivars that are widely cultivated in Ukraine. The discriminatory potential was tested for 11 ISSR-PCR primers, which produced 193 amplicons. UBC 835, 836, 841, and 881 were identified as the best primers suitable for routine application. The studied cultivars appear to be genetically highly heterogenic and can be divided into two main groups. The first one includes closely related cultivars obtained by hybridization of Drogans gelbe Knorpelkirsche, Valerii Chkalov, and some other forms. The second group comprises less similar cultivars derived from several West-European and unknown ancestors. Origin of several Ukrainian cultivars is discussed.     

Characterization of microsatellites in wild and sweet cherry (Prunus avium L.)--markers for individual identification and reproductive processes.

Nuclear microsatellites were characterized in Prunus avium and validated as markers for individual and cultivar identification, as well as for studies of pollen- and seed-mediated gene flow. We used 20 primer pairs from a simple sequence repeat (SSR) library of Prunus persica and identified 7 loci harboring polymorphic microsatellite sequences in P. avium. In a natural population of 75 wild cherry trees, the number of alleles per locus ranged from 4 to 9 and expected heterozygosity from 0.39 to 0.77. The variability of the SSR markers allowed an unambiguous identification of individual trees and potential root suckers. Additionally, we analyzed 13 sweet cherry cultivars and differentiated 12 of them. An exclusion probability of 0.984 was calculated, which indicates that the seven loci are suitable markers for paternity analysis. The woody endocarp was successfully used for resolution of all microsatellite loci and exhibited the same multilocus genotype as the mother tree, as shown in a single seed progeny. Hence, SSR fingerprinting of the purely maternal endocarp was also successful in this Prunus species, allowing the identification of the mother tree of the dispersed seeds. The linkage of microsatellite loci with PCR-amplified alleles of the self-incompatibility locus was tested in two full-sib families of sweet cherry cultivars. From low recombination frequencies, we inferred that two loci are linked with the S locus. The present study provides markers that will significantly facilitate studies of spatial genetic variation and gene flow in wild cherry, as well as breeding programs in sweet cherry.     

Rosaceae conserved orthologous sequences marker polymorphism in sweet cherry germplasm and construction of a SNP-based map

The Rosaceae Conserved Orthologous Set (RosCOS) provides a gene-based genome-wide set of markers that have been used in comparative analyses of peach (Prunus persica), apple (Malus × domestica), and strawberry (Fragaria spp.). In order to extend the use of these RosCOS to sweet cherry (Prunus avium L.), we identified markers that are polymorphic in breeding germplasm. Ninety-five percent (595/627) of previously designed RosCOS primer pairs amplified a product in six sweet cherry cultivars predicted to represent the range of genetic diversity in breeding germplasm. A total of 45% (282/627) RosCOS were polymorphic among the six cultivars, and allele number ranged from 2 to 6, with a genome-wide mean of 2.35. A subset of 92 genome-wide single nucleotide polymorphisms (SNPs) corresponding to 76 RosCOS was analyzed in 36 founder accessions and progeny. The expected and observed heterozygosity suggested that 83% of the RosCOS were in Hardy–Weinberg equilibrium, implying that most RosCOS behave as neutral markers. Principal coordinate analysis (PCO) identified one wild accession and two Spanish landraces that clustered differently from the other accessions. The relatively high number of unique alleles found in the three differentially clustered selections suggested that their use as parents has potential to increase the genetic diversity in future US-bred cultivars. Of the 92 RosCOS SNPs, 81 SNPs that represented 68 genome-wide RosCOS segregated in four mapping populations. These RosCOS were mapped in four F₁ populations, thereby greatly improving the genetic linkage map of sweet cherry.     

Assessment of genetic diversity of Czech sweet cherry cultivars using microsatellite markers

We analyzed 24 sweet and wild cherry genotypes collected in Czech Republic to determine genetic variation, using previously described 16 SSR primers to adapt a fast, reliable method for preliminary screening and comparison of sweet cherry germplasm collections. All SSRs were polymorphic and they were able all together to distinguish unambiguously the genotypes. These SSR primers generated 70 alleles; the number of alleles per primer ranged from 2 to 7, with a mean of 4.4 putative alleles per primer combination. The primer UDP-98-412 gave the highest number of polymorphic bands (totally 7), while Empa2 and Empa3 gave the lowest number (2). The allele frequency varied from 2.1% to 87.5%. We observed 10% of unique alleles at different loci. The observed heterozygosity value ranged from 0.25 to 0.96 with an average of 0.72 while expected heterozygosity value varied from 0.22 to 0.75 with an average of 0.59. The PIC value ranged from 0.21 to 0.71 with a mean value of 0.523. Cluster analysis separated the investigated cultivars in two groups. High level of genetic diversity obtained in the collection and proved to be sufficiently genetically diverse and therefore these genotypes would be useful to breeders for the development of new cherry cultivars.     

Identification and genetic diversity of plum cultivars grown in Belarus

A study of the collection of sour cherry, sweet cherry, common plum, diploid and tetraploid types of plums, and apricots grown in Belarus carried out using 20 SSR markers showed that they are characterized by high genetic diversity. Among 106 genotypes, 524 polymorphic alleles were identified. The average number of alleles was 15.4 in common plum samples, 11.3 in diploid and tetraploid plum, 9.3 in sour cherry, 6.0 in apricot, and 4.9 in sweet cherry. The greatest genetic diversity is characteristic of common plum cultivars (PD = 0.811). The genetic diversity decreases as follows: diploid plum (PD = 0.741), sour cherry (PD = 0.721), apricot (PD = 0.673), and sweet cherry (PD = 0.655). Cluster analysis shows that the degree of intraspecific divergence in sour cherry and sweet cherry cultivars is less than that of common plum, diploid plum, and apricot plum. Although apricots and plums belong to the subgenus Prunophora, according to the results of SSR analysis, apricot cultivars form a cluster that is more distant from both Cerasus and Prunophora. A set of seven SSR markers (EMPA001, EMPA005, EMPA018, EMPA026 and BPPCT025, BPPCT026, BPPCT039) was selected for DNA identification of cultivars of sour cherry, sweet cherry, common plum, diploid plum, and apricot, as well as species and interspecies hybrids.     

Phenotypic and genotypic variation in Iranian sour and duke cherries

Phenotypic and genotypic variation and structure of 29 sour cherry (P. cerasus) and duke cherry (P. x gondouinii) genotypes from different regions of Iran were identified using random amplified polymorphic DNA (RAPD) markers and morphological characters. Furthermore, one Prunus mahaleb genotype was used as an outgroup for molecular analysis. For morphological analysis, 23 variables were recorded to detect similarities between and among studied sour and duke cherries. Most studied characteristics were showing a high degree of variability. Principal component analysis showed that the first three components explained a total of 73.87 % of the whole phenotypic variability. Based on the morphological cluster analysis, studied sour and duke cherry genotypes were placed into three main clusters. The first main cluster included 16 sour cherry genotypes. The second main cluster contained all duke cherry genotypes and eight sour cherry genotypes, while, only one sour cherry genotype was placed in third main cluster. For RAPD analysis, 17 primers generated a total of 233 discernible and reproducible bands across genotypes analyzed, out of which 214 (91.51 %) were polymorphic with varied band size from 300 to 3000 bp. According to the similarity matrix, the lowest similarity was obtained between P. mahaleb, as an outgroup, and sour cherry. Dendrogram based on molecular data separated genotypes according to their species and geographic origin. Low correlation was observed between the similarity matrices obtained based on morphological and RAPD data. The information obtained here could be valuable for devising strategies for conservation of Iranian sour and duke cherries.     

Fruit size QTL identification and the prediction of parental QTL genotypes and breeding values in multiple pedigreed populations of sweet cherry

Large fruit size is a critical trait for any new sweet cherry (Prunus avium L.) cultivar, as it is directly related to grower profitability. Therefore, determining the genetic control of fruit size in relevant breeding germplasm is a high priority. The objectives of this study were (1) to determine the number and positions of quantitative trait loci (QTL) for sweet cherry fruit size utilizing data simultaneously from multiple families and their pedigreed ancestors, and (2) to estimate fruit size QTL genotype probabilities and genomic breeding values for the plant materials. The sweet cherry material used was a five-generation pedigree consisting of 23 founders and parents and 424 progeny individuals from four full-sib families, which were phenotyped for fruit size and genotyped with 78 RosCOS single nucleotide polymorphism and 86 simple sequence repeat markers. These data were analyzed by a Bayesian approach implemented in FlexQTL? software. Six QTL were identified: three on linkage group (G) 2 with one each on groups 1, 3, and 6. Of these QTL, the second G2 QTL and the G6 QTL were previously discovered while other QTL were novel. The predicted QTL genotypes show that some QTL were segregating in all families while other QTL were segregating in a subset of the families. The progeny varied for breeding value, with some progeny having higher breeding values than their parents. The results illustrate the use of multiple pedigree-linked families for integrated QTL mapping in an outbred crop to discover novel QTL and predict QTL genotypes and breeding values.     

PCR markers for mutated <Emphasis Type="Italic">S</Emphasis>-haplotypes enable discrimination between self-incompatible and self-compatible sour cherry selections

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, the 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. We previously determined that sour cherry has non-functional S-haplotypes for the S ₁ -, S ₆ - and S ₁₃ -haplotypes that are also present in diploid sweet cherry (P. avium L.). The mutations underlying these non-functional S-haplotypes have been determined to be structural alterations of either the S-RNase or SFB. Based on these structural alterations we designed derived cleaved amplified polymorphic sequence (dCAPS) markers and S-haplotype specific primer pairs that took advantage of either the length polymorphisms between S-haplotypes, differential S-haplotype sequences, or differential restriction enzyme cut sites. These primer pairs can discriminate among the mutant and wild-type S-haplotypes thereby enabling the identification of the S-haplotypes present in a sour cherry individual. This information can be used to determine whether the individual is either SC or SI. In a sour cherry breeding program, the ability to discriminate between SI and SC individuals at the seedling stage so that SI individuals can be discarded prior to field planting, dramatically increases the program’s efficiency and cost-effectiveness.     

Purification and properties of two pectinesterases from tomatoes.

Pectinesterase is present in green tomato fruit and increases several-fold during ripening. Several isoenzymes of pectinesterase are known to exist in tomatoes, but one isoenzyme predominates in the fruit of most cultivars. A few cherry tomato cultivars have been identified that contain low levels of this isoenzyme and much higher levels of another pectinesterase that is unique to those cultivars. The two major pectinesterases were purified to homogeneity and characterized. There were significant differences in the pectinesterases but they cross-reacted with antibodies raised against them and their N-terminal amino acid sequences were similar.     

The mutated S1-haplotype in sour cherry has an altered S-haplotype-specific F-box protein gene

Gametophytic self-incompatibility (GSI) is an outcrossing mechanism in flowering plants that is genetically controlled by 2 separate genes located at the highly polymorphic S-locus, termed S-haplotype. This study characterizes a pollen part mutant of the S(1)-haplotype present in sour cherry (Rosaceae, Prunus cerasus L.) that contributes to the loss of GSI. Inheritance of S-haplotypes from reciprocal interspecific crosses between the self-compatible sour cherry cultivar Ujfehértói Fürt?s carrying the mutated S(1)-haplotype (S(1)'S(4)S(d)S(null)) and the self-incompatible sweet cherry (Prunus avium L.) cultivars carrying the wild-type S(1)-haplotype revealed that the mutated S(1)-haplotype confers unilateral incompatibility with a functional pistil component and a nonfunctional pollen component. The altered sour cherry S(1)-haplotype pollen part mutant, termed S(1)', contains a 615-bp Ds-like element within the S(1)-haplotype-specific F-box protein gene (SFB(1)'). This insertion generates a premature in-frame stop codon that would result in a putative truncated SFB(1) containing only 75 of the 375 amino acids present in the wild-type SFB(1). S(1)' along with 2 other previously characterized Prunus S-haplotype mutants, S(f) and S(6m), illustrate that mobile element insertion is an evolutionary force contributing to the breakdown of GSI.