Correlation of isoenzyme polymorphism and Bayoud-disease resistance in date palm cultivars and progeny

Summary Seedlings of date palm (Phoenix dactylifera L.), obtained from seven cultivars crossed with two males, were analyzed by polyacrylamide gel electrophoresis for esterase (EST), glutamate oxaloacetate transaminase (GOT), endopeptidase (ENP) and alcohol dehydrogenase (ADH) polymorphisms. Eleven, eight, five and two phenotypes were revealed for the enzymes tested, respectively. Seedlings of F₁ populations derived from Bayoud (Fusarium)-resistant and low fruit quality cultivars were characterized by a high electrophoretic polymorphism, when compared with progenies of Bayoud-susceptible and high fruit quality cultivars. In almost all cases, the most frequent electrophoretic phenotypes scored for each enzyme in different F₁ populations, were similar to those of the corresponding parent cultivars. Heterozygous phenotypes have been found for, at least, 3 loci; Got-2, Est-1 and Enp, indicating that such loci could be used to screen for hybrid seedlings. The expected Mendelian segregation of allozymes has been observed for these 3 loci, in many F₁ populations. It seems that progenies of Bayoud-resistant cultivars are characterized by a high level of electrophoretic polymorphism. The estimation of this index and the search for genetic linkage with segregating allozymes, may be biochemical criteria useful as an aid in distinguishing date palm seedling populations derived from Bayoud-resistant cultivars and suitable for breeding programs.     

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.     

Genetic variation within and between two cultivars of red clover (Trifolium pratense L.): Comparisons of morphological,isozyme, and RAPD markers

Summary Morphological, isozyme and random amplified polymorphic DNA (RAPD) markers were used to estimate genetic variation within and between cultivars of red clover (Trifolium pratense L.), an important temperate forage legume. Two cultivars of red clover, Essi from Europe and Ottawa from Canada, were evaluated. Six monogenic morphological characters were observed for 80 plants from each of these two cultivars. All six morphological loci were polymorphic in the cultivar Essi whereas only four loci were polymorphic in the cultivar Ottawa. Forty plants from each cultivar were assayed for isozyme markers. A total of 21 enzyme-coding loci with 43 alleles was detected using twelve enzyme systems. Thirteen and nine of these loci were polymorphic in Essi and Ottawa, respectively. The mean number of alleles per locus was 1.81 in Essi and 1.67 in Ottawa. Seventeen random 10-mer primers were screened for RAPD markers. Nine primers which gave clear and consistent amplified products were used to assay 20 individuals from each cultivar. Each primer gave from 7 to 20 amplified bands with an average of 14.8 bands per primer. One hundred and eight of 116 putative loci were polymorphic in Essi and 90 of 98 loci were polymorphic in Ottawa. High within-cultivar variation was observed in both cultivars using both isozyme and RAPD markers. This high polymorphism makes these markers useful for germplasm characterization and genetic studies in red clover.     

Isozyme electrophoregrams of sixteen enzymes in five tissues of cassava (Manihot esculenta Crantz) varieties

Summary Methodology, based on starch and polyacrylamide gel electrophoresis, was developed for determining isozyme electrophoregrams (patterns) of 16 enzymes of cassava (Manihot esculenta Grantz) varieties as potential genotype markers. Extracts of five different tissues (root, stem, leaf, petiole and bud) were examined. In general, the nodal portions of the shoots gave isozyme patterns with the largest number of bands. Petiole extracts gave similar results but bud extracts gave poor patterns. The limited number of varieties that were examined could be distinguished by sequential classification on the basis of the isozyme patterns of acid phosphatase, esterase, glutamate oxaloacetase transaminase and phosphoglucoisomerase.Joint publication of the Centro Internacional de Agricultura Tropical and the Department of Plant Science (No. 716), University of Manitoba. Presented in part at the 2nd International Symposium on Biochemical Approaches to Identification of Cultivars and Evaluation of Their Properties, 5–9 May, 1985, Braunschweig, West Germany.     

Isozyme polymorphism in Festuca rubraL.

Summary Six Festuca rubra populations from Europe and Scandinavia were studied for variation at three isozyme loci; phosphoglucoisomerase (PGI-2), glutamate oxaloacetate transaminase (GOT-3) and superoxide dismutase (SOD-1). Seven alleles were found at the Pgi-2 locus, four at the Got-3 locus and five at the Sod-1 locus. Most plants were heterozygous and up to five alleles were found in the same plant at the Pgi-2 locus. Each population could be distinguished by the presence or absence of certain alleles or by differences in the frequencies of the alleles present. Values for the Shannon diversity index were calculated which showed that there was considerable heterogeneity both within and between loci. In general, 53% of this diversity could be attributed to within population variation.     

Allozyme differentiation of European populations and cultivars of Lolium perenne L., and the relation to ecogeographical factors

Summary Sixty Lolium perenne populations were screened for allozyme diversity at five loci. Objective was to determine whether allozyme diversity could be used as selection criterion for genebank accessions of L. perenne. Subsampling of locations was tested with allozyme analysis to determine whether genetically different populations could be collected at one location. Correlations between allelic frequencies and environmental factors and morphological data were established, to find ecogeographical patterns in the observed variation. Results indicated that with few examples each allelic variant could be observed in each population screened; very few unique alleles were found. Differences between populations were largely due to differences in allelic frequencies. Few correlations were found with environmental factors and morphological data. For some allele frequencies a north-south cline was observed. Generally, allozymic data of the five screened loci did not appear to be useful for the selection of accessions for genebank storage. Significant genotypic differences between populations collected at one location could be established. In general these results agreed with earlier results concerning phenotypic variation in the same populations.     

Recurrent phenotypic selection for forage yield in rye

Summary Cultivars of Cucurbita pepo and other Cucurbita species were characterized by RFLP analysis using different fragments of the ribosomal intergenic spacer (IGS) of Cucurbita pepo as hybridization probes. Several cultivars could be distinguished by a specific rDNA restriction pattern, whereas some cultivars showed an identical RFLP pattern suggesting a closer relationship. Other species of the genus Cucurbita exhibited strong cross-reaction with the C. pepo spacer probes, in contrast to DNA of Cucumis species which did not cross-hybridize.Abbreviations IGS intergenic spacer - ITS internal transcribed spacer - kbp kilo base pairs - rDNA ribosomal DNA - RFLP restriction fragment length polymorphism - rRNA ribosomal RNA     

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     

Identification of some Populus deltoides Marsh. x P. nigra L. clones,developed in North America,with the aid of allozymes

Summary Horizontal starch gel electrophoresis was done to assay 10 enzyme systems in root tips of 14 Populus deltoides x P. nigra clones developed by controlled hybridization and selected in North America, and some clones sharing one or both parents. The genotypes of the clones were determined for 31 loci coding for 10 enzyme systems. The interclonal allozyme variability was controlled by 12 loci. Each of the 14 clones had unique 12-locus genotypes, thus could be distinguished from each other. The clones differed from each other on an average of 4.2 loci. The first two Principal Components from Principal Component Analysis of the clonal allozyme genotype data accounted for 48% of the total variation in the 12 variant loci. 6-Pgd-3, Per-2, 6-Pgd-1, Aco-1 and Per-1 were found to be the most discriminating loci for the clones. The ordination of the clones on Principal Component axes 1 and 2 was in general agreement with the origin of the clones.     

Identification of triploid Citrus by isozyme analysis

Summary Seedlessness is a desirable horticultural attribute in Citrus and is positively associated with triploidy. The conventional cytological method for triploid identification is a laborious technique involving the preparation of root tips for chromosomal analysis. Digital densitometry of isozymes, however, offers the possibility of distinguishing triploid Citrus from large populations of seedlings both quickly and cheaply. Where there are no gene dosage regulation effects, greater band density should be evident in the allozyme contributed by the diploid gamete for a heterozygous locus. The isozymes of 4 enzymes; malate dehydrogenase, 6-phosphogluconate dehydrogenase, shikimate dehydrogenase, and phosphoglucose isomerase, were investigated with polyacrylamide gel electrophoresis. Band densities of these isozymes for triploid Citrus, their diploid siblings and diploid progenitors were measured using a digital densitometer. Of the 4 enzymes investigated only allozymes for shikimate dehydrogenase exhibited consistent differences over a wide range of Citrus cultivars. Greater band density was evident in the allozyme contributed by the diploid gamete. The band density ratio between allozymes for triploid Citrus was close to 0.5, while for diploid Citrus band density ratios were close to 1.0. This effect is due to the extra protein coded by the additional gene dose and was not observed in diploids. Shikimate dehydrogenase proved to be an accurate molecular marker for distinguishing between diploid and triploid Citrus for heterozygous progeny.Abbreviations PAGE Polyacrylamide gel electrophoresis - MDH malate dehydrogenase - 6PGD 6-phosphogluconate dehydrogenase - SkDH shikimate dehydrogenase - PGI phosphoglucose isomerase     

Diversity in the wild potato species Solanum chacoense Bitt.

Summary The morphological and isozyme variation was studied in 22 accessions of Solanum chacoense from Paraguay and Argentina. Clear geographic groups were identified through the use of multivariate analyses. S. chacoense from mountain sites in Argentina could be readily distinguished from plains forms from Paraguay, on the basis of several correlated morphological characters. Three isozyme systems, namely phosphoglucose isomerase (PGI), glutamate-oxaloacetate transaminase (GOT) and peroxidase (PRX) were studied using starch gel electrophoresis. The banding patterns indicated that for each isozyme there were several loci, which were polymorphic. A genetic interpretation of one of the PGI loci was made, and indices of genetic diversity and genetic identity calculated. Principal components analysis, cluster analysis and genetic diversity indicated a close relationship between the geographical groups. These results are discussed in the context of in situ genetic conservation.     

Identification of two types of differentiation in cultivated rice (Oryza sativa L.) detected by polymorphism of isozymes and hybrid sterility

Magnitudes of genetic variation within each of major varietal groups of cultivated rice were surveyed in terms of isozyme polymorphism and allelic differentiation of hybrid sterility loci, both of which are considered to have litt le selective value. Allelic differences for 20 isozyme loci were examined in a total of 337 accessions, including Indica and Japonica rices. Aus cultivars from India, Javanica cultivars and both landraces and cultivars from China. Eleven out of the 20 isozyme loci were polymorphic. The Aus cultivars contained more alleles per locus than others. The hybrid sterilities in the crosses among Chinese cultivars, Indica and Japonica cultivars were mainly controlled by locus S-5, whereas the hybrid sterilities of Aus cultivars, when they were crossed to Indica, Japonica or Javanica cultivars, were found to be controlled by allelic interaction at hybrid sterility loci S-5, S-7, S-9 and S-15. Also in terms of the number of alleles at S-5 and S-7, Aus cultivars contained more alleles than others. While the Aus group showed an extreme diversity for both hybrid sterility alleles and isozymes, modern cultivars from the south of YangZi River in China were classified into Indica type and those from the north were into Japonica, which were almost the same as those in Japan. On the basis of the measured polymorphism the Indica-Japonica differentiation was explained by founder effects, i.e., through selection and distribution of two original sources each with a unique set of markers. Contrastingly, the continuous and pronounced diversity in the Aus group was attributed to the absence of such a process. The intermediate groups in Yunnan province and Tai-hu Lake region of China are considered to be isolated from such founder effects, retaining an intermediate diversity in terms of isozymes and hybrid sterility genes. Since hybrid sterility hampers the exchange of genes between cultivars of different groups, the understanding of its genetic basis will be important in rice breeding, particularly in hybrid rice breeding. This revised version was published online in July 2006 with corrections to the Cover Date.     

Genetic analysis and nomenclature for seven isozyme systems in Brassica nigra, B. oleracea and B. campestris

General criteria for the assignment of names to enzyme systems, regions of activity, isozyme loci and allozymes have been lacking in crucifer species. This paper proposes a standard nomenclature for seven isozyme systems in the three diploid species of U's triangle: Brassica nigra, B. oleracea and B. campestris. Gel/electrode buffers, which provided the best resolution for seven isozyme systems, acid phosphatase (APS), aconitase (ACO), leucine aminopeptidase (LAP), 6-phosphogluconate dehydrogenase (6 PGD), phosphoglucoisomerase (PGI), phosphoglucomutase (PGM), and triosephosphate isomerase (TPI), were proposed as standards. Isozyme genetic analysis was determined for B. oleracea and B. campestris from previous studies and by segregation of selfed progenies of heterozygous B. nigra plants. Several populations were studied and 148 allozymes at the 18 loci observed were described for the three species. Their relative mobility was studied using a pure line of oilseed rape as reference. The comparison of the different alleles within and between the species is discussed.     

Spaced-plant traits related to seed yield in plots of perennial ryegrass (Lolium perenne L.)

Summary Seed yield in perennial ryegrass is low and selection criteria for high seed production are lacking. Indirect selection in spaced plants would be efficient, but spaced-plant traits need to be identified that correlate with seed yield in drilled plots. Spaced plants were studied of nine perennial ryegrass cultivars with a different seed production when grown in plots. Genotypic variation within cultivars for seed yield components and other traits was assessed in four environments in replicated clonal experiments. Extensive genotypic variation for most traits was present within each cultivar. Based on cultivar means of 25 or 50 genotypes, significant differences among cultivars were found for most traits. Cultivar-year interactions were found for most traits, but no significant cultivar-location interactions were generally found. Spaced-plant traits in general showed poor correlation to corresponding traits in drilled plots. Cultivars with contrasting seed yield in plots could not be distinguished on the basis of their spaced-plant traits and therefore no criteria for indirect selection for seed production in drilled plots could be identified in spaced plants.     

The Effect of Isozyme Selection on Yield and Flowering Time in Lolium perenne

A series of lines, differing in isozyme genotype, were constructed by selection from within three cultivars of Lolium perenne. Field assessment of the progeny of these selected line? revealed variation in yield and timing of inflorescence emergence. This variation between lines from within a cultivar could not be accounted for by the differing levels of homozygosity attained. In some instances differences in yield were associated with homozygosity for specific alleles at the PGI/2 locus, background genotype was however, of major importance. The variation identified could be attributable to founder effects or possible linkage of quantitative trail loci to the PGI/2 locus.     

Application of isozyme electrophoresis for purity testing and cultivar identification of F1 hybrids of Brassica oleracea

Summary Six isozyme genes were analyzed in seed samples of 65 commercial F₁ hybrids of four horticultural groups of Brassica oleracea (cabbage, Brussels sprouts, sprouting broccoli and cauliflower). Results obtained from electrophoretic assays led to the following conclusions: 1) the electrophoretic test of F₁ hybrid purity was possible in 59 (91%) of the hybrids analyzed, since their inbred parents were apparently fixed each for a different allele in at least on of the loci studied; 2) forty-eight (74%) of the hybrids were individually distinguished by their isozyme phenotype; 3) high levels of segregation in the inbred parents were inferred from the analysis of a sample of seeds of each hybrid.     

Identification of rape seed cultivars (Brassica napus) by starch gel electrophoresis of enzymes

Summary Isozyme patterns were used to characterize ten commercial rape seed (Brassica napus) cultivars. Extracts of cotyledons were made 4–7 days after germination and separated by electrophoresis on starch gels. A sample of more than 100 plants of each variety was analysed and stained for 4 polymorphic enzyme loci (Lap, Gpi, Acon, and Sdh). Therewith, it was possible to distinguish all ten rape varieties qualitatively by at least one typical enzyme pattern. Further, it could be demonstrated that all pairs of varieties showed clear qualitative differences in isozyme patterns, when only tow loci were screened (Acon, Sdh). Using ² test of homogeneity all pairs of varieties differ significantly in their frequencies of isozyme patterns for Acon and in all but one for Sdh.     

Assessment of quantitative resistance of rice cultivars to Xanthomonas campestris pv. Oryzae: a comparison of inoculation methods

Summary Lesion size and lesion number were measured on cultivars of rice inoculated by clipping or spraying with virulent isolates of Xanthomonas campestris pv. oryzae. Mean percentage diseased leaf area (%DLA) gave a similar ranking for the two inoculation methods but differences in lesion size among cultivars were much more evident after clip than after spray inoculation. Correlation between the methods was high (r=0.82^**), but some cultivars responded differently with the two techniques. Cultivars which had low scores following spray inoculation showed low disease progress during the first nine weeks after transplanting into a screen-house experiment. Assessment after clip inoculation measures resistance due to spread of bacteria within the leaf xylem system, an important component of quantitative resistance. Assessment after spray inoculation measures all resistance, including resistance to entrance of bacteria into the leaf. In order to select rice entries with improved quantitative resistance to X. c. oryzae based on both components, a screening based on lesion length after clip inoculation, followed by a test for lesion number after spray inoculation, is advised.     

Cultivar identification and genetic relationship among selected breeding lines and cultivars in chickpea (Cicer arietinum L.)

Twenty two RAPD and 22 ISSR markers were evaluated for their potential use in determination of genetic relationships in chickpea (Cicer arietinum L.) cultivars and breeding lines. We were able to identify six chickpea cultivars/breeding lines by cultivar-specific markers. All of the cultivars tested displayed a different phenotype generated either by the RAPD or ISSR primers. Though ISSR primers generated less markers than RAPD primers, the ISSR primers produced higher levels of polymorphism (% of polymorphic markers per primer) than RAPD primers. A high level of within cultivar homogeneity was observed in chickpea. Cultivars/breeding lines originating from a common genetic background showed closer genetic relationship. Chickpea lines with similar seed type(kabuli or desi) had a tendency to cluster together. This revised version was published online in August 2006 with corrections to the Cover Date.     

New Isozyme Markers in Vicia faba: Inheritance and Linkage

Results on the inheritance of 6 enzyme systems: LDH, PGM, FDH, SKD, SOD, AAT from seeds of Vicia faba and the linkage relationships among these isozyme loci are presented. The allozymes at each one of these loci behaved in a codominant manner and segregated in the expected Mendelian ratio. Linkage tests between these loci showed that they segregate independently.