Morphological variation within a sour cherry collection

Summary Morphological traits of 28 full-sib sour cherry (Prunus cerasus L.) families developed with pollen from European sour cherry selections were evaluated with principal component (PC) analysis. The traits which loaded on the first PC were size characters such as lateral length, leaf area, fruit weight, and trunk diameter increase. These character loading on the first PC could be interpreted as representing gradations between morphologies characteristic of the 2 presumed progenitor species, sweet cherry (P. avium L.) and ground cherry (P. fruticosa Pall.). Mean family differences in trunk diameter increase, lateral length, leaf area, and fruit weight varied approximately 12, 3.7, 2.5, and 2 fold, respectively. These results suggest that it may be possible to select sour cherry hybrids approaching the tree and fruit size of either progenitor species. The results are discussed in reference to germplasm collection and the potential of certain cultivars and hybrids as parents.     

Selection of a mutant from adventitious shoots formed in X ray treated cherry leaves and differentiation of standard and mutant with RAPDs

Summary An obstacle when using scions orin vitro shoots for mutation induction is the occurrence of chimeras. When adventitious shoots are formed from irradiated material these usually are derived from single cells, this leading to homohistont mutants. SincePrunus avium regenerates adventitious shoots from leaves at a low rate only (Yang & Schmidt, 1992), leaves of the interspecific cherry rootstock ‘209/1’ (P. cerasus ×P. canescens) were irradiated. ‘209/1’ regenerates adventitious shoots readily. Dosages applied were 5, 10, 20 and 40 Gy. Shoot production following 5 Gy irradiation was similar to the control. The application of 40 Gy resulted in strong damage with only few leaves regenerating. Among the adventitious shoots from leaves irradiated with 20 Gy one shoot was evident alreadyin vitro with thicker and smaller leaves having a serrate margin. It was cloned as ‘209/1-20m’. The clone stayed stable since 1990in vitro, in the greenhouse and the field. Compared with standard ‘209/1’ the mutant is very dwarf. Research to differentiate between standard ‘209/1’ and ‘209/1-20m’ was done using RAPDs. Among the decamer primer kits D, J, and T from Operon Technologies, Calif. Only primer OPJ05 (5‘CTCCATGGGG3’) differentiated between ‘209/1’ and ‘209/1-20m’. Rootstock ‘209/1’ showed one band of 2 kb additionally. This band is missing in the mutant.     

Identification of self-incompatibility alleles and pollen incompatibility groups in sweet cherry by PCR based s-allele typing and controlled pollination

A total of 17 pollen incompatibility groups in sweet cherry (Prunusavium L.) were identified among 46 accessions by PCR based S-alleletyping analysis and by controlled test pollinations. Two putativeS-alleles different from S ₁ to S ₆,S _z and S _y were identified. Five S-genotypes, S ₁ S ₅, S ₁ S ₆,S ₂ S ₆, S ₄ S ₆, andS ₅ S ₆, combinations of S ₁ toS ₆ alleles that had not previously been identified from cultivars in NYSAES, were positively confirmed by PCR based S-genotyping analysis. Also, the S-genotypes of cultivars in some pollen incompatibility groups that had previously been incorrectly reported have been clarified. Several popular cultivars, which were previously used as testers for S-allele typing analysis, were found to have been inaccurately genotyped. In addition, the S-genotypes and self-incompatibility groups of some relatively recentlyintroduced cultivars were identified. The molecular typing system ofS-genotypes based on PCR is a useful and rapid method for identifying newS-alleles and incompatibility groups in sweet cherry.     

Correlation of stylar ribonuclease zymograms with incompatibility alleles in sweet cherry

Summary Protein stylar extracts of 16 cultivars of sweet cherry (Prunus avium), from the 10 different incompatibility groups to which incompatibility alleles have been assigned, were separated on acrylamide gels using isoelectric focusing (IEF) and were stained for ribonuclease activity. When two cultivars from the same incompatibility group were analyzed they gave identical zymograms and the cultivars of the 10 different incompatibility groups gave in all eight distinct zymograms. The ribonuclease polymorphism could be correlated with the reported S allele constitutions of the cultivars. Three ribonuclease bands were identified that each consistently corresponded to one of the six known incompatibility alleles (S ₁, S₂ and S ₆), a fourth band apparently corresponded to S ₃ and to the combination of S ₄ and S ₅, and a fifth band to S ₄ and S ₅ in other combinations. Thus, it seems that S alleles of cherry have ribonuclease activity and that IEF is useful for distinguishing S allele constitutions. The ribonuclease pattern of Summit, a cultivar of unknown incompatibility group, indicated its incompatibility genotype to be S ₁S₂, and this was confirmed by controlled pollination. The same band corresponded to S ₄ and S ₄', the mutant allele in self-compatible cultivars. IEF and ribonuclease staining promise to be useful tools for exploring the incompatibility relationships of cherry cultivars and perhaps of other self-incompatible Prunus crops.     

Identification of cultivars of the forage legume Pueraria phaseoloides by electrophoretic patterns of storage proteins

Summary A polyacrylamide gel electrophoresis procedure was developed for discriminating cultivars of the forage legume Pueraria phaseoloides on the basis of the patterns of cotyledon proteins. The genotypic marker proteins were extracted with 5M acetic acid and electrophoresed at pH 3.1 in aluminum lactate buffer. The procedure gave highly reproducible discrimination of ten selected cultivars.     

Study of the parentage of grape cultivars by genetic interpretation of GPI-2 and PGM-2 isozymes

Summary Two highly variable enzyme systems of glucosephosphate isomerase (GPI) and phosphoglucomutase (PGM) were used to investigate the parentages of grape cultivars. Of 35 parent/offspring combinations that we investigated, 30 combinations gave alleles in the offspring which were presented in the reported parents, whereas 5 combinations gave alleles in the offspring which were not extractable from the reported parents. The Gpi-2 genotype of Hiro Hamburg and the Pgm-2 genotype of Pione indicated that Koshu Sanjaku and Cannon Hall Muscat may not have been the paternal parent respectively. The Gpi-2 genotype of New Niagara and the Gpi-2 and Pgm-2 genotypes of Beniyamabiko indicated that Niagara and a hybrid from DxK151 x Delaware may not have been the maternal parent respectively. The Gpi-2 genotype of Cannon Hall Muscat grown in Japan indicated that this cultivar may not have originated as a tetraploid sport of Muscat of Alexandria.     

Identification of tomato cultivars (Lycopersicon esculentum) by polyacrylamide isoelectric focusing

Summary Isozyme analyses have been used for the definitive identification of many plant cultivars, but not for cultivated tomatoes. Six isozyme systems, namely alcohol dehydrogenase, acid phosphatase, phosphoglucomutase, esterase, phosphoglucoisomerase and 6-phosphogluconate dehydrogenase of tomato seed extracts were resolved by isoelectric focusing on polyacrylamide gels with a narrow pH gradient. Nine alcohol dehydrogenase phenotypes were distinguished which, with three acid phosphatase phenotypes, identified twelve of the seventeen cultivars. Fewer differences were found for the other isozymes. Since this method could differentiate between breeding parents and their progeny it is concluded that further investigations are warranted.Abbreviations APS acid phosphatase - ADH alcohol dehydrogenase - EST esterase - IEF isoelectric focusing - PGI phosphoglucoisomerase - PGM phosphoglucomutase - 6-PGDH 6-phosphogluconate dehydrogenase - RFLP restriction fragment length polymorphism - VOPRI Vegetable and Ornamental Plant Research Institute     

Polymorphism and DNA markers for asparagus cultivars identified by random amplified polymorphic DNA

Summary DNA polymorphism among five Asparagus officinalis L. cultivars-Imperial, Snow, Steline, UC-157 and Larac, as detected by random amplified polymorphic DNA (RAPD), is reported. Thirty one decamer primers were tested. and twenty six of them yielded amplification products. Fourteen primers gave products with at least one polymorphic DNA fragment. Among a total of 119 amplified fragments 33 were polymorphic. These RAPD markers enabled the identification of asparagus cultivars. Unique markers for cultivars were: Snow-bands 475 bp, 772 bp, 412 bp, 935 bp and 820 bp amplified by primers D5, OPA-07, OPA-09, OPA-10 and OPA-18, respectively. Steline-bands 645 bp, 680 bp and 997 bp amplified by primers A32, OPA-03 and OPA-09, respectively. A band 903 bp, amplitied by primer OPA-12, is a marker for Imperial, and a band 420 bp, amplified by primer D52, is a marker for Larac. Cultivar UC-157 could be identified by a combination of shared polymorphic bands. The pairwise marker difference between cultivars ranged from 0.08 to 0.17. A phenogram of the genetic relationship based on RAPD fits with the known origin of the cultivars.     

Genetic variation and cultivar identification of Jamaican yam germplasm by random amplified polymorphic DNA analysis

Summary We have used random amplified polymorphic DNA (RAPD) analysis to characterize eleven cultivars of the five economically most important yam species grown in Jamaica (Dioscorea alata, D. cayenensis, D. rotundata, D. trifida and D. esculenta). Amplification of genomic DNA samples with nine different arbitrary 10mer primers revealed a total of 338 different band positions, ranging in size from 0.3 to 2.5 kb. RAPD patterns proved to be highly reproducible and somatically stable. While no variation was observed among plants belonging to the same cultivar, a large number of intervarietal and interspecific polymorphisms enabled us to reliably discriminate between all Jamaican cultivars investigated.     

Distinguishing among white seeded bean cultivars by means of allozyme genotypes

Summary Allozyme genotypes were determined at 10 loci for 90 cultivars of white seeded snap beans. Within cultivars the loci were homozygous and usually monomorphic, permitting the characterization of most cultivars by a single set of allozymes. A total of 72 allozyme combinations were observed among the cultivars tested, and 52 (58%) of the cultivars could be uniquely distinguished by allozyme genotype alone. The remaining 38 lines could be separated into small groups of 2–5 cultivars each.This paper has been approved by the Director of the New York State Agricultural Experiment Station as Journal Paper No. 3511.     

Identification of cultivars of Stylosanthes capitata Vog. by polyacrylamide gel electrophoresis of seed proteins

Summary A method was developed for identification of cultivars of the pasture legume, Stylosanthes capitata Vog., using electrophoretic patterns of seed proteins in polyacrylamide gels as the genotypic markers. The method can be used for accurate identification of cultivars in germplasm banks, in selecting parents for development of new varieties, and in registering new cultivars for proprietary purposes.     

Genetic variation among cultivars of red clover (Trifolium pratense L.) detected by RAPD markers amplified from bulk genomic DNA

Summary The use of random amplified polymorphic DNA (RAPD) markers obtained from bulked samples was investigated for cultivar identification in red clover. Pooled samples were examined in order to minimize variation within cultivars. To determine the appropriate number of individuals to include in the bulked samples representing each cultivar, DNA samples from two, three, four, five, ten and twenty individuals were pooled. Twenty was found to be an appropriate number of red clover individuals per bulk in order to amplify only the DNA sequences shared among most individuals in each cultivar. Fourteen 10-mer primers were used to amplify genomic DNA from combined leaf samples of 15 red clover cultivars from European, Japanese and North American origins. A total of 79 amplified products, of which 55 were polymorphic, was obtained. Cultivar-specific bands were observed with 13 primers. The amplification patterns obtained from two primers could distinguish all 15 red clover cultivars. Rogers' genetic distances for all 105 pairwise comparisons were calculated to evaluate relationships among these cultivars. Cluster analysis based on these genetic distances separated these 15 cultivars into three groups, with two of the groups consisting of a single Japanese cultivar each, while the third group included cultivars from European, North American, and Japanese origins.     

Identification of cultivars of pasture legumes (Centrosema macrocarpum,C. pubescens and C. sp.n.) by acid polyacrylamide gel electrophoresis of cotyledons storage proteins

Summary An electrophoretic procedure was developed for storage proteins which can discriminate cultivars of forage legumes, Centrosema macrocarpum, C. pubescens and C. sp.n. Proteins extracted from cotyledons were separated in 10% polyacrylamide gel containing 10% sucrose. Electrophoretic patterns are presented to illustrate the results that can be obtained with the procedure described. Cultivars of all three species were distinguishable based upon the variation in their acid soluble seed proteins.Joint publiccation of the Department of Food Science, University of Manitoba (No 101) and Centro Internacional de Agricultura Tropical-CIAT.     

Polyacrylamide gel electrophoresis of wheat gliadins: the use of a moving boundary for improved resolution

Summary The process of the separation of gliadin proteins of wheat, using polyacrylamide gel electrophoresis, was studied in detail. It was shown that electrophoresis is a dynamic process during which the pH of the gel changes together with the potassium ion concentration. The most positive effect on the separation of the gliadins was generated by a moving front, which is a boundary between regions with a low concentration of K⁺ ions and a low pH, and region with a high concentration of K⁺ ions and a high pH, after optimization of the concentrations of the cations and anions in the electrode solutions. The finding was exploited for the development of an extremely simple electrophoresis system, in which buffers were not needed for obtaining a high resolution. The system was further improved by applying a stacking gel. The advantages of this system are discussed. This new approach may be useful for improving electrophoresis systems for other applications.     

The use of reversed-phase high performance liquid chromatography as an aid to the identification of European barley cultivars

Summary A reversed-phase high performance liquid chromatography (RP-HPLC) system was used to separate the storage proteins (hordeins) extracted from European barley cultivars. From a total of 38 barleys tested, 26 types of hordein patterns could be distinguished after RP-HPLC. This appears to be a marked improvement in resolution over that achieved in a similar survey of European barley cultivars using SDS polyacrylamide electrophoresis (32 hordein patterns resolved by SDS PAGE from a total of 160 spring and winter barleys tested).Different hordein patterns were resolved by RP-HPLC within each of two groups of barley previously classified by SDS PAGE as indistinguishable within groups (three distinct patterns identified in a total of five cultivars tested from group 1A and five patterns observed among eight cultivars from group 3B). Thus RP-HPLC achieves a higher resolution than undirectional electrophoresis and promises to be a valuable aid in the identification of European barley cultivars.     

Utilization of isozyme polymorphism for cultivar identification of 45 commercial peas (Pisum sativum L.)

Forty five Pisum sativum cultivars were analysed by isozyme electrophoresis with the objective to find protein markers for exact and reproducible discrimination of individual genotypes. The combination of six enzyme systems (acid phosphatase, amylase, esterase, leucine aminopeptidase, shikimate dehydrogenase and phosphoglucomutase) with two electrophoretic techniques (NATIVE-PAGE, isoelectric focusing) and use of seed and leaf tissue enabled to identify all 45 studied cultivars. Critical factors which may affect utilization of isozyme electrophoresis for commercial applications in pea breeding and seed production and testing are discussed. This revised version was published online in July 2006 with corrections to the Cover Date.     

DNA fingerprinting and classification of geographically related genotypes of olive-tree (Olea europaea L.)

Málaga is a province of Spain where olive-trees are cultivated in a large range of environments, climates and soils. We have developed a reliable and reproducible method to detect RAPD and AP-PCR polymorphisms, using DNA from olive-tree (Olea europaea L.) leaves. Starting from their natural orchards, fifty-six olive-tree cultivars throughout Málaga province, including oil and table olive cultivars, were screened and grouped into 22 varieties. A total of 62 informative polymorphic loci that provide 601 conspicuous bands were enough to differentiate the varieties. Clustering analyses managing 3 different pairwise distances, as well as phylogenetic analyses, led to the same result: olive-trees in Málaga can be divided into three main groups. Group I (90% of certainty) contains wild type and two introduced varieties, group II (83% of certainty) covers some native olive-trees, and group III (58% of certainty) is an heterogeneous cluster that includes varieties originating and cultivated in a number of Andalusian locations. Geographic location seems to be the first responsible of this classification, and morphological traits are needed to justify the group III subclustering. These results are consistent with the hypothesis of autochthonic origin of most olive-tree cultivars, and have been used to support a Label of Origin for the olive oil produced by the varieties included in group II. This revised version was published online in July 2006 with corrections to the Cover Date.     

Field bean (Phaseolus vulgaris L.) cultivar identification by electrophoregrams of cotyledon storage proteins

Summary Electrophoretic procedures were developed for seed proteins which can discriminate cultivars of field beans. Proteins were extracted from seven varieties and the extracts were analysed using acid and SDS polyacrylamide gel electrophoresis. Electrophoregrams are presented to illustrate the results that can be obtained with the methods described. Results indicate that sufficient variation is present among the seven cultivars examined to afford unambiguous discrimation and identification of the cultivars. Banding patterns were stable for each genotype.Joint publication of the Department of Plant Science, University of Manitoba (No. 747) and Centro Internacional de Agricultural Tropical-CIAT.     

Identification of self-incompatibility (S) alleles in Latvian and Swedish sweet cherry genetic resources collections by PCR based typing

The Latvian and the Swedish sweet cherry (Prunus avium L.) genetic resources collections comprise valuable material for breeding. The collections represent local Latvian and Scandinavian genetic resources: semi-wild samples, landraces, and cultivars developed in local breeding programmes, as well as diverse germplasm from the northern temperate zone. The objective of this investigation was to determine which S ₁ –S ₆ alleles are most important in the sweet cherry genetic resources collections and to compare the identified allelic and genotypic frequencies in material of different origin. Accessions in the two collections were screened for the presence of the self-incompatibility (S) S ₁ to S ₆ alleles, using PCR based typing. Significant differences (P < 0.05) between screened collections were found in frequencies of S ₄ and S ₅ alleles. Analysis of allele combinations identified the high occurrence of selections with the S-genotype S ₃ S ₆ in both collections. Compared to the S-allele frequencies published for over 250 sweet cherry cultivars from Western and Southern Europe, the Latvian and Swedish germplasm appeared to have a high frequency of the S ₆ allele in both collections, and a relatively high frequency of the S ₅ allele in Latvian germplasm. This study represents the first comprehensive S-allele screening for the sweet cherry genetic resources collections in Latvia and Sweden. Both sweet cherry collections contain high proportion of accessions adapted to north central European growing conditions, not typical for the majority of the documented sweet cherry genetic resources, which explains differences in certain S-allele occurrence.     

Seed protein variation for identification of common buckwheat (Fagopyrum esculentum Moench) cultivars

Summary Total seed proteins of 24 common buckwheat cultivars and cultivated populations within a molecular weight range of 30 to 54 kDa were analysed by SDS-PAGE. Single seed analysis of six cultivars identified a total of 18 alternative protein bands with different mobilities. Differences of individual protein band frequencies extracted from single seeds among six buckwheat cultivars varied distinctively, indicating high intravarietal polymorphism. The relation between frequencies of protein bands revealed by single seed analysis and their appearance on the bulk seed analyses was demonstrated. Regarding to band mobility rate and relative band intensity among 24 bulk samples analysed, 14 had distinctive electrophoregrams while the other 10 were ranged into four distinct groups. Analysis of endosperm and cotyledon proteins showed that proteins stored in these main seed parts are tissue specific. The observed electrophoretic polymorphism related to proteins stored in the cotyledons while there was no apparent variability with endosperm proteins.