Genetic relationship of Mediterranean mandarins (Citrus deliciosa Tenore) using RAPD markers

Summary Random amplified polymorphic DNA (RAPD) analysis was carried out to evaluate polymorphism and genetic similarity between 39 Mediterranean mandarin genotypes. One hundred eleven amplification products were identified using 21 random primers. An average of 2.2 RAPD markers was obtained for each primer, corresponding to 42% of the amplification products. Genotype-specific RAPD markers were also found, mainly in known hybrids. UPGMA cluster analysis revealed the low level of genetic variation between accessions of Mediterranean mandarins, whereas their hybrids with other Citrus species showed greater genetic dissimilarity. Twenty accessions yielded very similar patterns, suggesting either that they could be a single clone, or that the technique was not able to detect genomic variation. However, for the other specimens genetic polymorphism can easily be detected by RAPD, although the genetic variation between accessions was quite low. The large number of hybrids and the low polymorphism between accessions support the hypothesis that Mediterranean mandarins are all true hybrid of Common mandarins (Citrus reticulata Blanco).     

Variation in some agronomically important characters in a germplasm collection of beet (Beta vulgaris L.)

Summary A geographically representative selection of germplasm of Beta vulgaris, section Beta has been assessed for characteristics important in sugarbeet breeding, including downy mildew resistance, resistance to aphid colonisation and infection by the beet virus yellow complex. The occurrence of maintainer lines for cytoplasmic male-steriles was also investigated. Desirable qualities were found in some accessions, including nothern European wild vulgaris ssp. maritima and some old multigerm cultivars of fodder beets.     

Assessment of genetic relatedness of the genera Ananas and Pseudananas confirmed by RAPD markers

Genetic relationships among 18 accessions, including 16 of Ananas and two of Pseudananas, were investigated using RAPD molecular markers. The procedure for DNA extraction was adapted from the method of Dellaporta et al. (1983) where an incubation in proteinase K and a purification step were included. From the total of 148 markers scored,132 (89.2%) were polymorphic. The similarity matrix was used for cluster analysis. The phenogram developed from the RAPD bands showed that for most of the cases, the accessions within a species grouped together. Nevertheless, a moderate infraspecific genetic variation was observed. For example, DNA data grouped all A. comosus accessions with a mean similarity coefficient of 0.85. Comparable results were obtained with all other species investigated. The highest genetic divergence was found withinA. lucidus where the mean similarity coefficient among accessions was0.75. A similar level of genetic polymorphism was observed among species,therefore, a definition about which species were involved in the constitution of A. comosus genotypes was not possible. These results agree with the breeders standpoint suggesting that all Ananas species belong to the primary gene pool of pineapple. This revised version was published online in July 2006 with corrections to the Cover Date.     

Partial resistance of sugarbeet to beet cyst eelworm (Heterodera schachtii Schm.)

Summary Within Beta vulgaris and B. maritima origins some partial resistance to beet cyst eelworm was found which could be raised to a very limited extent by selection. However after the second backcross to commercial sugar beet varieties and successive selection of the inbreds this resistance was lost. It was demonstrated that in the rootsystem of resistant plants as much nematodes penetrate and develop as in susceptible ones but the ratio between males and females is different. It was therefore quite probable that this resistance is polyfactorial and merely recessive.     

Molecular marker diversity and bacterial wilt resistance in wild <Emphasis Type="Italic">Solanum commersonii</Emphasis> accessions from Uruguay

Solanum commersonii is a wild tuber-bearing species native to Uruguay with high potential for use in potato breeding programs. Little is known about the genetic diversity within this wild species and the relationship with the resistance to the bacterial pathogen Ralstonia solanacearum. We studied 30 S. commersonii clonal accessions, 20 of which were collected from geographically different areas across the country, while the other ten were grown from seeds from a single plant. Resistance against R. solanacearum was tested and different levels of resistance were found, ranging from delayed wilting to asymptomatic reactions. The genetic variation and the relationships among individuals in this germplasm collection were studied by different molecular markers: Random Amplified Polymorphic DNA (RAPD), Amplified Fragment Length Polymorphism (AFLP) and Microsatellites or Simple Sequence Repeats (SSR). AFLP markers generated the largest number of total and polymorphic fragments per assay unit while SSR revealed the highest frequency of polymorphic bands (100%), followed by AFLP (96.2%) and RAPD (89.4%). In contrast, when comparing the number of different genetic profiles generated, the SSR markers exhibited the lowest discriminatory power. The clustering pattern obtained with the three marker systems showed a similar distribution of the S. commersonii germplasm revealing a high correlation between the three methods employed. All three dendrograms grouped most of the accessions into two main clusters, containing the same accessions regardless of the marker type. Bacterial wilt resistant accessions were present in both clusters. Accessions originated from different seeds of the same plant were grouped within one of the major clusters, and differed in the response to R. solanacearum revealing segregation of resistance. Furthermore, the distribution in two main clusters showed high correspondence with the geographical origin of the accessions, from the north and south of the country, and with the subspecies malmeanum and commersonii morphologically identified.     

Isozyme variation in Helianthus petiolaris and sunflower,H. annuus

Summary Helianthus petiolaris is a wild species used in genetic improvement of sunflower, as a donor of cytoplasmic male sterility and of genetic resistance to diseases. Isozyme variation for ADH, ACP, EST, GDH, LAP, PGI, PGD and SKDH in this species was studied using starch gel electrophoresis. The patterns thus obtained were compared with zymograms of inbred lines, hybrids and open pollinated varieties of H. annuus. The same alleles for EST and SKDH isozymes were found in both species, while ACP showed an allele that has not been found in sunflower. The rest of the isozyme systems showed both common alleles and characteristic ones for each species. ACP, GDH and PGD were monomorphic in H. petiolaris, while ADH and LAP were monomorphic in H. annuus. The isozyme markers obtained here could be useful in breeding programs involving interspecific crosses, and studies on introgression and on genetic variation in other populations of this wild species.     

Crosses between Beta vulgaris L. and Beta lomatogona F. et M.

Summary A population of 2x^ms sugar beets was crossed with 4x Beta lomatogona F. et M. The 3x F₁-plants were male sterile and were backcrossed with 2x and 4x sugar beets and multiplied without pollination as well. After the 1st backcross mainly 3x apomict types arose again and, among others, a small number of successful 4x backcrosses. After pollination by 4x sugar beets this 4x F₁ B₁ produced. besides predominatly apomictically multiplied 4x plants, also about 7% haploid 2x hybrids. The latter probably possess 1 genome from B. vulgaris and 1 genome from B. lomatogona. In the meiosis of the PMC's a certain amount of homeology between a number of chromosomes of both species could be established. The amphihaploid hybrids can be used as breeding parents for the creation of types in which introgession can occur. During hybridization in addition to 2x and 4x B. vulgaris types a number of 2x-, 3x-, 5x- and 6x-hybrids arose. This is presumably caused by the presence of gametes with the somatic number of chromosomes and the occurrence sometimes of haploid apomictic multiplication.The presence of large numbers of bolters in the F₁ and F₁ B₁ suggests that the bolting tendency of both species is based on different genes.     

Expression in vitro of resistance to Heterodera schachtii in hairy roots of an alien monotelosomic addition plant of Beta vulgaris,transformed by Agrobacterium rhizogenes

Summary Hairy roots, induced by Agrobacterium rhizogenes, were obtained of nematode susceptible beet plants (Beta vulgaris) and of the nematode resistant alien monotelosomic addition AN5, carrying a telosome from B. patellaris. The additional telosome was found to be stably present in vitro in the roots of AN5. The hairy root cultures were inoculated with larvae of the beet cyst nematode Heterodera schachtii. On the root culture of AN5 significantly less cysts developed than on the other root cultures. These results indicate that the resistance to the beet cyst nematode is expressed in the roots after transformation and can be monitored under in vitro conditions.     

Genetic and environmental effects on chromosome doubling of sugarbeet (Beta vulgaris L.) haploids

Summary Chromosome doubling has been a limiting factor for production of doubled-haploids, a means of obtaining fruly homozygous individuals, and a time-saving alternative to repeated selfing for the creation of inbred lines. The existence of genetic, environmental and genotype × environment interaction effects on chromosome doubling ability of sugarbeet (Beta vulgaris L.) haploids was investigated. Haploids were derived from four distinct, highly heterozygous diploid populations through in vitro culture of unpollinated ovules. Ovule-derived plants were treated with colchicine to double their chromosome complement. Environmental effects were determined from replication of the experiment at seven dates. Both genetic and environmental factors were found to affect chromosome doubling ability of sugarbeet haploids. No significant interaction between genotype and environment was identified. The presence of genetic effects on chromosome doubling ability is discussed with regard to its implications on breeding programs.     

Ecological aspects of transgenic sugar beet: transfer and expression of herbicide resistance in hybrids with wild beets

Summary An increasing number of genetically engineered cultivars of several crops is being experimentally released into the environment. In future, crops with new transgenic traits will probably play an important role in agricultural practice. The long-term effect of transgenes on community ecology will depend on the distribution and establishment of transgenic plants in the wild, on the sexual transfer of their new genes to the environment and on the potential ecological impact of the transgenic trait. The starting point was the use of transgenic sugar beet lines, Beta vulgaris subspec. vulgaris var. altissima DÖLL (Helm 1957), with transgenes coding for rhizomania and herbicide (BASTA^®) resistance. The first two questions to answer were: Can the transgenes be transferred via pollen to wild beets, Beta vulgaris subspec. maritima (L.) ARCANG. or cultivated relatives such as red beet or spinach beet and are they expressed in the hybrids? Can transgenes be monitored in young Beta vulgaris-hybrids? The experimental transfer of transgenes was conducted in 1993 at a field location in northern Germany. The beets were hand-pollinated with transgenic pollen. In a non destructive biotest, the hybrid seedlings were tested for herbicide resistance. Transgenic plants showed no noxious phenotypic effects whereas control plants developed leaf necroses. All herbicide resistant hybrids within the biotest were assumed to be transgenic.     

The production of alien monosomic additions in Beta vulgaris as a source for the introgression of resistance to beet root nematode (Heterodera schachtii) from Beta species of the section Patellares

Summary Experiments were carried out for adding the chromosome carrying resistance to beet root nematode (Heterodera schachtii) from the wild Beta species of the section Patellares (B. procumbens, B. webbiana and B. patellaris) to the genome of B. vulgaris. Preliminary experiments indicated that crosses between the wild species and B. vulgaris cultivars of the mangold type yielded on average more viable F₁ hybrids than crosses with sugar and fodderbeet. However, crossability varied strongly between individual parental combinations. It was concluded that most types of B. vulgaris can be hybridized with the wild species of the section Patellares if a sufficient number of pair-crosses is made. Crosses between diploid cultivars or species of the section Vulgares and diploid wild species of the section Patellares yielded many hybrids which, however, were highly sterile. From crosses between tetraploid B. vulgaris and the wild species a great number of viable allotriploid and allotetraploid hybrids was obtained. In the backcross progenies of allotriploid hybrids 26% alien monosomic additions occurred, of which 4.1% carried the resistance bearing chromosome of B. procumbens or B. patellaris. The programme will be continued by sereening progenies of the resistant monosomic addition plants for the occurrence of resistant disomic introgression products.     

Resistance in Lycopersicon hirsutum f. glabratum to the greenhouse whitefly (Trialeurodes vaporariorum) increases with plant age

Summary In this study genetic variation for resistance to the greenhouse whitefly (Trialeurodes vaporariorum) of four genotypes of tomato (L. esculentum) and two subspecies of L. hirsutum was investigated. Resistance was quantified by the whitefly life history components adult survival, oviposition rate, pre-adult survival and developmental period, measured on plants inoculated with whiteflies in clip-on cages.The largest differences between species were found when life history components were measured on adult plants of about four months old. On L. hirsutum f. glabratum whiteflies had the lowest adult survival, oviposition rate and pre-adult survival. On L. hirsutum these components were intermediate whereas on all L. esculentum genotypes they were highest. The variation between plants was low compared to the variation within plants. These results indicate that single plant tests can be used to determine accurately genetic variation between individual plants in a segregating population.     

Species relationships in the subgenus Ceratotropis (genus Vigna) as revealed by RAPD analysis

Summary The genetic variation among 23 accessions of 5 species in the subgenus Ceratotropis, genus Vigna, were investigated by random amplified polymorphic DNA (RAPD) analysis. A total of 404 fragments amplified with 24 primers were scored and analyzed by cluster analysis. The accessions used were separated into two main groups with an average of 70% differences. Within the main groups, five subgroups were recognized, which are in complete agreement with taxonomic species. Wild forms were always grouped with their most closely related cultivated forms and they showed variation in each species. The largest intraspecific variation was found in V. radiata (mungbean), in which wild forms (V. radiata var. sublobata) were highly different from each other and from cultivated forms. V. angularis (adzuki bean) showed the least variation and thus, was probably differentiated in relatively recent times.     

Inter and intra-population variability of <Emphasis Type="Italic">Jatropha curcas</Emphasis> (L.) characterized by RAPD and ISSR markers and development of population-specific SCAR markers

Jatropha curcas (Euphorbiaceae) is an oil-bearing species with multiple uses and considerable potential as a bioenergy crop. The present investigation has been undertaken to assess the extent of genetic diversity in a representative set of 42 accessions of J. curcas encompassing different crop growing regions in India along with a non-toxic genotype from Mexico as a prelude for utilization of promising and genetically divergent materials in the breeding programmes. Molecular polymorphism was 42.0% with 400 RAPD primers and 33.5% with 100 ISSR primers between accessions indicating modest levels of genetic variation in the Indian germplasm. The within-population variation based on RAPD polymorphism was 64.0% and was on par with the inter-population variation. Polymorphic ISSR markers have been identified that could differentiate the Indian accessions from the Mexican genotype and two of them were converted to SCAR markers. The SCAR primer pair ISPJ1 amplified a 543 bp fragment in all the Indian populations, while ISPJ2 with a specific amplicon of 1,096 bp was specific to the Mexican genotype. Population-specific bands have been identified for the accession from Kerala (2 RAPD markers), Neemuch-1 from Rajasthan (1 each of RAPD and ISSR markers) and the non-toxic genotype from Mexico (17 RAPD and 4 ISSR markers), which serve as diagnostic markers in genotyping. The study indicates an immediate need for widening the genetic base of J. curcas germplasm through introduction of accessions with broader geographical background.     

Assessment of natural and induced genetic variation in Alstroemeria using random amplified polymorphic DNA (RAPD) markers

Summary We have used random amplified polymorphic DNA (RAPD) markers to study genetic variation in Alstroemeria. The first objective was to examine the discriminatory power of RAPD markers in different genotypes of Alstroemeria obtained by traditional breeding. All genotypes examined, including commercial Alstroemeria varieties, could be distinguished on the basis of their RAPD profiles. Progeny plants could be distinguished from their parents. A second objective of this study was to investigate whether RAPD markers can be used as a routine tool to detect mutant plants, as an alternative to glasshouse testing. To address this objective, we analysed Alstroemeria plants that carried phenotypically visible mutations that either were induced by irradiation using X-rays or were the result of somaclonal variation. In eight out of a total of 13 mutant Alstroemeria plants obtained after irradiation or tissue culture we detected no polymorphisms when compared to control plants that were considered to be non-mutated. Only in five of the mutant plants analysed we detected one to two polymorphisms. These results suggest that frequent genome rearrangements had not occurred in the mutant plants analysed. These results also demonstrate that the RAPD technique is an inappropriate tool for the rapid screening of Alstroemeria for induced variation. It that the RAPD technique is an inappropriate tool for the rapid screening of Alstroemeria for induced variation. It seems probable that this conclusion would be equally applicable in other plant genera in which induced variation has occurred. However, the RAPD technique is a simple and effective tool for genetic fingerprinting of Alstroemeria varieties, provided their differences are due to sexual propagation.     

Evaluating genetic variation and relationships among two bromegrass species and their hybrid using RAPD and AFLP markers

The two most widely grown bromegrass species in North America are smooth bromegrass (Bromus inermis Leyss.), a hay type grass, and meadow bromegrass (Bromus riparius Rehmann), a pasture type grass. Recently a hybrid bromegrass population between the two species has been produced as a dual-purpose hay-pasture grass. Molecular markers have the potential to improve selection procedures to enhance bromegrass breeding. The objective of this study was to use RAPD and AFLP markers to determine genetic relationships and variations among bromegrass populations. Forty-three RAPD markers from 21 primers and 83 AFLP markers from seven primer combinations were used. Both marker types were able to group the individuals into their respective populations. The relationships among the individuals within each of the populations were not similar between the two marker types. Analysis of molecular variance (AMOVA) detected greater within-population variation than among-population variation for both marker types. The highest variation was observed in the hybrid population followed by meadow and then smooth bromegrass. The inter-population distance from both markers indicated that the highest genetic distance was between meadow and smooth bromegrass and lowest between smooth and hybrid bromegrass, which reflect the breeding history of the hybrid population. This study showed that both markers are capable of differentiating bromegrass genotypes into their respective populations, detecting genetic variation and relationships of the populations. Results of this study suggest that these two markers can be used in the future to enhance the current breeding practices in bromegrass, however, AFLP markers would be the marker of choice due to the high number of polymorphic markers. This revised version was published online in July 2006 with corrections to the Cover Date.     

Cytogenetic analysis of interspecific hybrids in genus Citrullus (Cucurbitaceae)

Investigations on interspecific crossability in two species of the genus Citrullus were studied based on their morphology, chromosome pairing and pollen fertility in their hybrids resulting from 12 combinations(including reciprocals). These have been utilized for tracing the phylogenetic relationship between the taxa and factors responsible for their differentiation. A collective evaluation of data on cross compatibility,morphology, chromosome association and pollen fertility indicated a close phylogenetic relationship between the species C. colocynthis (wild2n = 22) and C. vulgaris (cultivated 2n = 22). Significant differences for crossability and pollen fertility were observed between hybrids of C. colocynthis × C. vulgaris and reciprocals. Meiotic analysis of F₁ hybrids revealed that these two species are freely crossable and provide normal chromosome association, involving an average of 9 to maximum 11 pairs of chromosomes. This indicated the possibility of high recombination and thereby genetic introgression of desirable traits. As both the species are of economic significance, breeding strategies can be developed for the genetic improvement of these species. This revised version was published online in August 2006 with corrections to the Cover Date.     

Genetic diversity of wheat wild relatives in the Near East detected by AFLP

In order to reveal the molecular genetic diversity of wheat wild relatives, an AFLP analysis was conducted with 16 accessions of five Triticum andAegilops species originating from the Near East. Variation within population was studied with at least seven individuals per accession. Four primer combinations were used for selective amplification. Based on the scored bands, we estimated percentage of polymorphic bands, 1 – proportion of shared bands (1-psb) and nucleotide diversity (π). Of the five species used in this study, Ae. speltoides had the highest level of `within population' variation. This species had also the highest value of the variation among populations. As for Triticum species, the level of variation within population was low in diploid species (T. urartu and T. boeoticum),whereas two tetraploid species (T. dicoccoides and T. araraticum) had relatively high levels of variation within population. While the two diploid Triticum indicated a clear interspecific divergence, the two tetraploid wild wheats were not clearly divergent in this study. The variance portioning analysis indicated that the variation detected for diploid Triticum species was mainly composed of `between species' variation, on the other hand that for tetraploid Triticum was mostly composed of `within population' variation. In conclusion, AFLP analysis reveals molecular variation in all accessions used in this study, suggesting a potential genetic diversity of the wheat wild relatives in natural populations. These results have implications for the design of strategies to maintain genetic diversity within genebank collections. This revised version was published online in August 2006 with corrections to the Cover Date.     

Genetic diversity and relationships among Lablab purpureus genotypes evaluated using RAPD as markers

Summary Genetic variation in 40 accessions of Lablab purpureus was evaluated using random amplified polymorphic DNA as markers. A high level of genetic variation in this species was detected but this was mainly restricted to the difference between cultivated and wild forms. Of the cultivated genotypes, genetic variation among Asian collections was significantly higher than that among African collections. The three most divergent cultivated genotypes were all from Asia. Four of the five wild accessions, two from Zimbabwe and the other two from Zambia, were closely related. The other one, CPI 31113 collected from Uganda, was highly divergent. The two commercial forage varieties used in Australia, Rongai from Kenya and Highworth from India, were not very different.     

The taxonomic characterisation of annual Beta germplasm in a genetic resources collection using RAPD markers

Summary Annual beets in the genus Beta section Beta represent an important genetic resource. Representative accessions of annual beets from a beet germplasm collection were analysed using RAPD to assess the patterns of variation and relationships among them. Using arbitrary primers, markers showing variation across accessions were identified. A dendrogram of similarity was produced using these molecular markers. All the accessions analysed were classified into three major groups corresponding to species or subspecies macrocarpa, adanensis and maritima. Macrocarpa was shown to be the most divergent group in this section. Using RAPD molecular markers, it was possible to ascribe an accession to one of three taxonomic groups and overcome much of the confusion encountered when morphological traits are used for identification. The group of maritima was found to be more polymorphic than either the group of macrocarpa or adanensis at both accession and subspecies levels.