Open-pollinated Seeds Research Articles

Using genetic diversity and mating system parameters estimated from genetic markers to determine strategies for the conservation of Araucaria angustifolia (Bert.) O. Kuntze (Araucariaceae)

In order to understand the impacts of forest fragmentation on Araucaria angustifolia populations, we evaluated the genetic diversity and mating system using SSR markers and open-pollinated seeds from four populations of varying sizes and spatial isolation, in and around one of the best-conserved Araucaria Forest remnants in Southern Brazil. The four population types of A. angustifolia include: (1) a continuous forest; (2) a physically isolated cluster located 2 km from the continuous forest; (3) an open population in a field located between the cluster and continuous forest; and (4) a fragment on a private property located 5 km from the cluster. Approximately 28 seeds were collected from ten reproductive trees in each population. We found higher amounts of alleles (113) and exclusive alleles (25) in the continuous forest than in the other populations. The multilocus paternity correlation was significantly higher and effective number of pollen donors was significantly lower in the private population, decreasing the diversity and consequently the variance effective size of families sampled from that population. However, despite its isolation from the other studied fragments, the private population had the second highest number of alleles as well as unique alleles from the other populations. Therefore, strategies for A. angustifolia conservation should focus not only on larger populations, such as those found in protected areas, but also include smaller and isolated fragments on private properties as these populations are able to maintain high levels of genetic diversity and functional connectivity between isolated stands across a landscape.

Seed exchange networks and food system resilience in the United States

Seed exchange is a multidimensional issue with social, political, economic, and agricultural implications. There is a growing concern about the increase of the food system’s vulnerability as a result of loss of agricultural biodiversity. Farmers’ ability to replant, exchange, and distribute saved seed is a way to minimize their dependence on commercial suppliers and thereby maintain control over farming practices. Seed saving is also crucial for conservation because the process of choosing, replanting, and exchanging seeds relies on and increases diversity on the farm and in communities. Seed exchange networks, formal and informal ways that farmers engage alongside institutional plant breeding systems, help to conserve agricultural, social, and cultural diversity and identity as well as enhance resilience against environmental and economic shocks. However, how to build resilient seed systems and move from the innovative but relatively isolated project activities of professionals and farmers to a situation where such approaches are scaled up and networked alongside formal and informal, national and international plant breeding mechanism are a concern. This paper examines grassroots seed exchange through seed libraries, the marketing of new varieties through seed companies, and hybrid civil society-business models to understand their financial and technical abilities as well as challenges they face. Seed exchange networks fulfill an important role in conservation of agricultural biodiversity and building community resilience through their work on breeding, exchange, and propagation of regionally adapted open-pollinated seeds as well as advocacy on seed sovereignty and education on seed saving.

Mendelian inheritance, genetic linkage, and genotypic disequilibrium at microsatellite loci in Genipa americana L. (Rubiaceae).

Genipa americana is a tropical tree species that is widely distributed in the humid tropical and subtropical regions of Central and South America. This study investigated Mendelian inheritance, genetic linkage, and genotypic disequilibrium at six microsatellite loci developed for G. americana. Adult trees (188) and regenerants (163) were sampled and genotyped in a fragmented population of the species. We also genotyped open-pollinated seeds from 12 seed-trees during reproductive events in 2010 and 2011. Significant deviations from the expected 1:1 Mendelian segregation were detected in 29.5% of the tests. Significant genetic linkage between pairwise loci was detected in 54.4% of the tests, but no genotypic disequilibrium was detected between pairwise loci for adult trees and regenerants. Overall, the results indicate that the six loci analyzed may be used in studies of G. americana's genetic diversity and structure, its mating system, and in parentage analyses.

Provenance and seed mass determines the seed germination success of Eucalyptus ovata (Myrtaceae)

Eucalyptus ovata is a medium size tree which grows from near sea level to sub-alpine regions in the south-east of Australia and is increasingly being used for woodland restoration in Tasmania. Given the wide altitudinal range of the species, we investigated whether a wet, cold stratification treatment enhanced seed germination of high altitude provenances. Open-pollinated seeds were harvested from five trees from a high and low altitude provenance within each of the five geographic regions from across E. ovata 's distribution in Tasmania. Chilling imbibed seeds at 5°C for three weeks had no effect on the overall proportion of viable seeds which germinated. There were, however, significant provenance differences in seed weight and the germination traits (i.e. proportion of viable seeds, germination rate and overall proportion germinated) which were variably linked to differences in altitude. Seed viability tended to increase with seed mass and provenance-level correlations amongst germination traits suggested that seeds from different provenances differed in germination vigour. These results indicate that provenance rather than a pre-treatment determines germination success of E. ovata seed, but these provenance differences are not predictable and that selection of heavier seeds may increase germination success in the nursery and in direct seeding applications.

Registration of ‘Centennial’ Sand Bluestem

‘Centennial’ (Reg. No. CV-15, PI 670042) sand bluestem (Andropogon hallii Hack.) is a synthetic cultivar selected for greater seed germination at a low water potential and seedling establishment under field conditions. Two cycles of recurrent selection were used to develop Centennial (ABmedium Syn-2) from ‘Chet’ sand bluestem (AB-medium Syn-0). Cycle 1 consisted of germinating 3500 open-pollinated seeds of population AB-medium Syn-0 in a −0.8 MPa D-mannitol solution for 7 d. All germinated seeds (~250) were selected to create population ‘AB-medium Syn-1’. Cycle 2 selection was similar to Cycle 1 except that population AB-medium Syn-1 was used to create population AB-medium Syn-2. Germination of seeds of Centennial was 16.3% higher than for those of Chet in a water potential of −0.8 MPa and 12.8% higher than Chet in deionized water. Seeds from Centennial had significantly greater field emergence (62.7%) than either the ABmedium Syn-1 (59.6%) or Chet sand bluestem (54.6%). Thus, percentage field emergence of sand bluestem was increased by recurrent selection for increased seed germination at a low water potential. Centennial sand bluestem was cooperatively released by the USDA–ARS and USDA–NRCS to provide forage for pasture, hay, or complementary rangeland–forage production systems and to stabilize soils by reclaiming marginal croplands in the central and southern Great Plains of the United States.

Variabilidad en el fruto y semillas de zapallo Cucurbita moschata Duch. y Cucurbita argyrosperma subsp. sororia L.H. Bailey Merrick & D.M. Bates

The cucurbits presents one range of variants of color in the seed: from white to brown, you may have evolutionary significance and geographical. Variability was evaluated in 295 seed Cucurbita moschata Duch introductions and 99 of Cucurbita argyrosperma subsp. sororia, all from open-pollinated seeds. The cluster analysis revealed 11 groups for each of the species, which expressed high variability between and within introductions for the characters under study: percentage of seed ether extract (EE), seed weight per fruit (SWPF) and number of fruits per plant (NFP). In C. moschata, two groups ( 5 and 9) stood out for the highest averages for yield per plant (PPL ), seed weight per fruit ( PSPF ) and ether extract per plant ( EEPP ) , with coefficients of variation ( CV ) between 52% and 93 % . In C. argyrosperma subsp. sororia, same characters stood out to groups 5 and 11, with a CV between 53 and 79 %. High coefficients of variation showed wide variability between and within introductions to the characters of agronomic and agro industrial importance. Given the cluster analysis and the weighted selection index (PSI) six introductions of each species was selected.

An open-pollinated design for mapping imprinting genes in natural populations.

With the increasing recognition of its role in trait and disease development, it is crucial to account for genetic imprinting to illustrate the genetic architecture of complex traits. Genetic mapping can be innovated to test and estimate effects of genetic imprinting in a segregating population derived from experimental crosses. Here, we describe and assess a design for imprinting detection in natural plant populations. This design is to sample maternal plants at random from a natural population and collect open-pollinated (OP) seeds randomly from each maternal plant and germinate them into seedlings. A two-stage hierarchical platform is constructed to jointly analyze maternal and OP progeny markers. Through tracing the segregation and transmission of alleles from the parental to progeny generation, this platform allows parent-of-origin-dependent gene expression to be discerned, providing an avenue to estimate the effect of imprinting genes on a quantitative trait. The design is derived to estimate imprinting effects expressed at the haplotype level. Its usefulness and utilization were validated through computer simulation. This OP-based design provides a tool to detect the genomic distribution and pattern of imprinting genes as an important component of heritable variation that is neglected in traditional genetic studies of complex traits.

Propagation for the Conservation of Pityopsis ruthii, an Endangered Species from the Southeastern United States

Pityopsis ruthii is an endangered species endemic to the Hiwassee and Ocoee Rivers in Tennessee. As part of a recovery effort focused on P. ruthii, vegetative propagation and in vitro multiplication and seed germination techniques were developed. Plants were vegetatively propagated using greenhouse stock plants and wild-collected stems. Rooting occurred with and without auxin treatments but was greatest when 0.1% indole-3-butyric acid (IBA) talc was applied to the vegetative cuttings; rooting was lowest when flowering stems were used. Pro-Mix BX substrate provided the most consistent rooting. In vitro multiplication was accomplished by the removal of lateral shoots from in vitro-grown plants that were rooted on Murashige and Skoog (MS0) basal medium with 270 clones produced from a single individual after 4 months. Nineteen clones were transplanted and secured with bonded fiber matrix into their natural habitat and 14 survived for 1 year. To avoid genetic swamping of native populations with the introduction of large numbers of genetically identical individuals through clonal propagation, seed-based propagation efforts were explored. Open-pollinated seeds were collected, disinfested and germinated, and seedlings established on MS medium. Seeds were submersed in 70% ethanol for 1 minute and briefly flamed. Seeds were surface-sterilized in a range [10% to 50% (v/v)] Clorox® bleach solutions with vigorous shaking for 20 minutes, rinsed three times in sterile water, and germinated on MS0. Removal of pappus from seeds was required for successful disinfestations, but the bleach concentration was not critical. Successful propagation is a step toward the conservation and recovery of P. ruthii and should allow future reintroduction projects.

Genetic evidence for paternal inheritance of the chloroplast in four Australian Callitris species (Cupressaceae)

The mode of chloroplast inheritance was investigated in the four Australian conifer species; Callitris columellaris, C. intratropica, C. gracilis and C. verrucosa (Callitroideae, Cupressaceae), as a basis for future use of chloroplast genetic markers for the genus. Open-pollinated seeds and the foliage tissue of the mother trees were collected from one population per species, in which two distinct chloroplast haplotypes at the trnL (UAA) 5′ exon–trnF (GAA) region were co-distributed. In total, the DNA sequences of the chloroplast fragments in 255 samples were determined, and the haplotypes of the germinated seedlings and the mother trees were compared. Among the 16 mother trees examined across all species, 12 trees were shown to have produced at least one offspring with a different haplotype from the mother, indicating the occurrence of paternal inheritance of chloroplast in the fertilization processes. In binomial GLMM analyses, the haplotype of mother trees in C. columellaris, C. gracilis, and C. verrucosa did not significantly affect those of their offspring, and the predicted frequency of seedlings' haplotypes was compatible with the observed frequency in adult canopy trees. These findings provide a genetic support that paternal inheritance of chloroplast DNA via pollen dispersal would be dominant in the Callitris species.

Somatic embryogenesis and plant regeneration from immature cotyledons of European chestnut (Castanea sativa Mill.)

Here, we established a protocol for induction of somatic embryogenesis and plant regeneration from immature cotyledons of open-pollinated seeds of European chestnut (Castanea sativa Mill.) cultivars ‘Osmanoglu’ and ‘Sariaslama’. Basal media, Murashige and Skoog medium (MS), Driver and Kuniyuki Walnut medium (DKW), and Woody Plant Medium (WPM) supplemented with l-glutamine or casein hydrolysate, with or without silver nitrate, agar or gelrite, and various plant growth regulator (PGR) combinations were tested in initial cultures for induction of somatic embryos. The effects of initial cultures on the percentage of somatic embryos and average number of embryos per cotyledon explant, subcultured monthly, were determined at the end of 4 mo. Interactions were observed among the different treatments for ‘Osmanoglu’ cultivar, with the highest rates of somatic embryogenesis (4.7–9.7%) being obtained in MS, DKW, or WPM basal media supplemented with (1) 6-benzyladenine (BA; 1 mg/L) + kinetin (KIN; 2 mg/L) + indole-3-butyric acid (IBA; 0.01 mg/L); (2) BA (1 mg/L) + 1-phenyl-3-(1,2,3-thiadiazol-5-yl; TDZ 0.1 mg/L) + IBA (0.01 mg/L), and (3) KIN (2 mg/L) + TDZ (0.1 mg/L) + IBA (0.01 mg/L) PGR combinations plus l-glutamine or casein hydrolysate, with or without silver nitrate, and with either gelrite or agar. The highest percentages (12.0% and 11.2%) of somatic embryogenesis for ‘Sariaslama’ were obtained in DKW supplemented with PGR combinations of (1) BA (1 mg/L) + KIN (2 mg/L) + IBA (0.01 mg/L), (2) BA (1 mg/L) + TDZ (0.1 mg/L) + IBA (0.01 mg/L), respectively. The average number of somatic embryos ranged between 0 and 0.65 per explant for ‘Osmanoglu’ and between 0 and 0.49 per ‘Sariaslama’ explant. For germination of somatic embryos, root, shoot, and plantlet regeneration, different treatments included desiccation, cold and gibberellic acid (GA3), and BA alone or in combination with auxins (IBA or α-naphthaleneacetic acid, NAA; 0.1 mg/L). The highest rate of somatic embryos regeneration (27.5%) occurred using MS basal media with half-strength microelements containing 0.1 mg/L BA + 0.1 mg/L NAA, after treatments of desiccation, or desiccation plus cold or GA3 (3 mg/L).

Genetic variation of drought-induced cavitation resistance among Pinus hartwegii populations from an altitudinal gradient

Hydraulic failure can cause massive die-back of forest trees during drought. With extreme climatic events set to become more frequent and severe due to climatic change, it is essential to study resistance to water stress-induced cavitation. We investigated the genetic differentiation for cavitation resistance among Pinus hartwegii populations, the pine species growing at the treeline in Mexico. Open-pollinated seeds were collected from seven natural populations along an altitudinal gradient (3,150–3,650 masl) from Pico de Tancitaro, Michoacan, western Mexico. Seedlings were raised in a nursery and then established in a randomized complete block design in a common garden experiment. Resistance to cavitation (P 50, xylem pressure inducing 50 % loss of hydraulic conductance and S, slope of the vulnerability curve) and specific hydraulic conductivity (k s), were evaluated on branches of 5-year-old seedlings using the Cavitron technique. Mean P 50 was −3.42 ± 0.05 MPa, indicating that Pinus hartwegii is one of the more vulnerable pine species to cavitation. No significant genetic differentiation was detected between populations for cavitation resistance traits (P 50 and S), but a significant altitudinal cline was found for S. In contrast, k s exhibited a significant differentiation among populations and a significant decline with increasing altitude. The lack of genetic differentiation among P. hartwegii populations for cavitation resistance is likely to represent a limitation for adapting to the warmer and drier climates that are expected to occur in Mexico under climatic change. Finally, a worldwide comparison within the Pinus genus showed that pines growing at the treeline were on average more vulnerable to cavitation than those from lowland. This might reflect an adaptation to dry environmental conditions at low elevation.

Edge effects enhance selfing and seed harvesting efforts in the insect-pollinated Neotropical tree Copaifera langsdorffii (Fabaceae)

Edge effects may affect the mating system of tropical tree species and reduce the genetic diversity and variance effective size of collected seeds at the boundaries of forest fragments because of a reduction in the density of reproductive trees, neighbour size and changes in the behaviour of pollinators. Here, edge effects on the genetic diversity, mating system and pollen pool of the insect-pollinated Neotropical tree Copaifera langsdorffii were investigated using eight microsatellite loci. Open-pollinated seeds were collected from 17 seed trees within continuous savannah woodland (SW) and were compared with seeds from 11 seed trees at the edge of the savannah remnant. Seeds collected from the SW had significantly higher heterozygosity levels (Ho=0.780; He=0.831) than seeds from the edge (Ho=0.702; He=0.800). The multilocus outcrossing rate was significantly higher in the SW (tm=0.859) than in the edge (tm=0.759). Pollen pool differentiation was significant, however, it did not differ between the SW (=0.105) and the edge (=0.135). The variance effective size within the progenies was significantly higher in the SW (Ne=2.65) than at the edge (Ne=2.30). The number of seed trees to retain the reference variance effective size of 500 was 189 at the SW and 217 at the edge. Therefore, it is preferable that seed harvesting for conservation and environmental restoration strategies be conducted in the SW, where genetic diversity and variance effective size within progenies are higher.

Long-distance dispersal in a fire- and livestock-protected savanna

Savannas are highly diverse and dynamic environments that can shift to forest formations due to protection policies. Long-distance dispersal may shape the genetic structure of these new closed forest formations. We analyzed eight microsatellite loci using a single-time approach to understand contemporary pollen and effective seed dispersal of the tropical tree, Copaifera langsdorffii Desf. (Fabaceae), occurring in a Brazilian fire- and livestock-protected savanna. We sampled all adult trees found within a 10.24 ha permanent plot, young trees within a subplot of 1.44 ha and open-pollinated seeds. We detected a very high level of genetic diversity among the three generations in the studied plot. Parentage analysis revealed high pollen immigration rate (0.64) and a mean contemporary pollen dispersal distance of 74 m. In addition, half-sib production was 1.8 times higher than full-sibs in significant higher distances, indicating foraging activity preference for different trees at long distances. There was a significant and negative correlation between diameter at breast height (DBH) of the pollen donor with the number of seeds (r = −0.640, P-value = 0.032), suggesting that pollen donor trees with a higher DBH produce less seeds. The mean distance of realized seed dispersal (recruitment kernel) was 135 m due to the large home range dispersers (birds and mammals) in the area. The small magnitude of spatial genetic structure found in young trees may be a consequence of overlapping seed shadows and increased tree density. Our results show the positive side of closed canopy expansion, where animal activities regarding pollination and seed dispersal are extremely high.

Mendelian inheritance, genetic linkage, and genotypic disequilibrium at nine microsatellite loci of Cariniana legalis (Mart.) O. Kuntze

Cariniana legalis is one of the largest tropical trees with a wide distribution in the Brazilian Atlantic rainforest. We investigated the Mendelian inheritance, genetic linkage, and genotypic disequilibrium at seven microsatellite loci specifically isolated for C. legalis, and at two previously developed heterologous microsatellite loci. Forty to 100 open-pollinated seeds were collected from 22 seed-trees in two populations. Using the Bonferroni correction, no remarkable deviations from the expected Mendelian segregation, linkage, or genotypic disequilibrium were detected in the nine loci studied. Only 3.7% of the tests were significant for investigations of the Mendelian proportions. On the other hand, only 2.8% of tests for linkage detection showed significance. In addition, among all pairwise tests used for investigating linkage disequilibrium, significance was found in 9.7% of the locus pairs. Our results show clear evidence that the nine simple sequence repeat loci can be used without restriction in genetic diversity, mating system, and parentage analyses.

Methodological approaches for the selection of genotypes in a progeny trial of <i>Dacryodes edulis</i> (G. Don) H. J. Lam in Cameroon

The World Agroforestry Centre (ICRAF) initiated a program in Cameroon for the selection of improved genotypes of Dacryodes edulis for domestication. There are major challenges in such selection because superior performance of individual trees may be based on site conditions rather than on genetic differences. Open-pollinated seeds were collected from mother trees in four provenances in Cameroon and planted in a progeny trial at Minkoameyos, Cameroon. Progeny from the same mother tree are referred to as family. After one year of growth in the field, growth traits were measured annually for five years and fruit traits were measured in two successive years. Three methodological approaches were investigated: family and within-family selection (combined selection), ranking for phenotypic selection and correlation for multi-trait and early selection. Variation in growth and fruit traits among families was not significant for most provenances, indicating that family selection would not be effective for the selection of genotypes in the trial. Phenotypic selection resulted in the retention of a small number of individuals of all families in the trial. There were strong positive correlations between fruit traits and between fruit and growth traits, while negative correlations were found between fruit traits and yield. The correlations indicate that selecting for one fruit trait may also improve the other fruit traits but reduce fruit yield. Positive and significant juvenile-mature correlations for growth were also found in this trial, indicating that early selection may be possible based ongrowth. Based on these results, phenotypic selection is recommended in the progeny trial for the domestication program in Cameroon and further research on early selection is needed as the trial advances in age.

Open-Pollinated Seed Exchange: Renewed Ozark Tradition as Agricultural Biodiversity Conservation

In situ conservation refers to the perpetuation of genetic resources in their original cultural and biophysical habitats and to the diverse strategies employed to sustain crop genetic diversity and the cultural milieus that support and maintain it. This article reviews in situ agrobiodiversity conservation in the Arkansas Ozarks, in the United States over a four-year period (2006–2010) through applied ethnographic and agroecological research. I examine the (re)introduction of “seed swaps” in the Ozark region as an in situ conservation strategy to connect diverse Ozark inhabitants, (re)institute a dynamic flow of agroecological knowledge, and conserve agricultural biodiversity in the region.

New insight into the history of domesticated apple: secondary contribution of the European wild apple to the genome of cultivated varieties.

The apple is the most common and culturally important fruit crop of temperate areas. The elucidation of its origin and domestication history is therefore of great interest. The wild Central Asian species Malus sieversii has previously been identified as the main contributor to the genome of the cultivated apple (Malus domestica), on the basis of morphological, molecular, and historical evidence. The possible contribution of other wild species present along the Silk Route running from Asia to Western Europe remains a matter of debate, particularly with respect to the contribution of the European wild apple. We used microsatellite markers and an unprecedented large sampling of five Malus species throughout Eurasia (839 accessions from China to Spain) to show that multiple species have contributed to the genetic makeup of domesticated apples. The wild European crabapple M. sylvestris, in particular, was a major secondary contributor. Bidirectional gene flow between the domesticated apple and the European crabapple resulted in the current M. domestica being genetically more closely related to this species than to its Central Asian progenitor, M. sieversii. We found no evidence of a domestication bottleneck or clonal population structure in apples, despite the use of vegetative propagation by grafting. We show that the evolution of domesticated apples occurred over a long time period and involved more than one wild species. Our results support the view that self-incompatibility, a long lifespan, and cultural practices such as selection from open-pollinated seeds have facilitated introgression from wild relatives and the maintenance of genetic variation during domestication. This combination of processes may account for the diversification of several long-lived perennial crops, yielding domestication patterns different from those observed for annual species.

Contemporary pollen flow, mating patterns and effective population size inferred from paternity analysis in a small fragmented population of the Neotropical tree Copaifera langsdorffii Desf. (Leguminosae-Caesalpinioideae)

Pollen flow and dispersal patterns were investigated in a small, isolated forest fragment of the Neotropical insect pollinated tree Copaifera langsdorffii, using paternity analysis and eight microsatellite loci. We also investigated the coancestry and effective population size of progeny arrays for conservation and environmental restoration purposes. Open-pollinated seeds were collected from 15 seed trees within the forest fragment, in which all adult trees were mapped, measured and genotyped. Twenty seeds were also collected from the nearest neighbor tree located 1.2 km from the forest fragment. Our results show that levels of genetic diversity were significantly higher in adults than offspring and significant levels of inbreeding were detected in offspring (F = 0.226). From paternity analysis, we observed low levels of selfing (s = 8%) and pollen immigration (m = 8%) in the fragment, but very high levels were detected for the isolated tree (s = 20%; m = 75%), indicating that the population and the tree are not reproductively isolated and are connected by patterns of long distance pollen dispersal (maximum detected 1,420 m). Within the forest fragment, the pattern of pollen dispersal was a near neighbor pattern with 49% of the pollen being dispersed within 50 m. The effective population size of the progeny array was low, indicating the need to collect seeds from a large number of seed trees (at least 76) for conservation purposes. The results show that the spatial isolation of the population and isolated tree due to forest fragmentation has not disrupted genetic connectivity; however, spatial isolation does seem to increase selfing and correlated mating.

Linking Phenology to Mating System: Exploring the Reproductive Biology of the Threatened Palm Species Butia eriospatha

The reproductive biology of the vulnerable palm species Butia eriospatha was studied to provide important information that contributes to our understanding and conservation of the species. In order to determine when and how B. erisopatha reproduces, we combined data from 7 nuclear microsatellite loci with ecological data on flowering and fruiting phenology collected between 2009 and 2011 from a population (N = 515) in the Atlantic Rainforest, Southern Brazil. Periods of flowering and fruit production were seasonal and variable across reproductive events. Mating system analyses indicate that B. eriospatha is a predominantly outcrossing species, ((m) = 0.961), since a certain degree of biparental inbreeding does occur. The species is self-compatible and reproduction may also occur by geitonogamy, indicating the ability of isolated populations to survive and persist. Open-pollinated seeds varied in relatedness, including mainly half-sibs and full-sibs. The effective population size was lower than that expected for panmictic populations. Hence, seeds for conservation programs must be collected from a large number of seed-trees to ensure an adequate effective population in the sample. The collection of germplasm is a high-priority strategy that should be employed to maintain the genetic variability that remains.

GENETIC CHARACTERIZATION OF SELECTED MACADAMIA GERMPLASM AND THE IMPLICATIONS ON BREEDING AND CONSERVATION IN KENYA

Macadamia is the most important nut crop in Kenya with an estimated annual production of about 13,510 metric tones with an estimated value of $ 4.8M. The cultivated macadamia is derived from two species; the rough-shelled M. tetraphylla and the smooth shelled M. integrifolia. The Kenyan macadamia industry relies on selection of superior trees from germplasm derived from open-pollinated seeds of the two species planted between 1946 and 1968. Twenty six macadamia accessions sampled from five populations; Bob Harries, Thika, Kirinyaga, Embu and Meru in central and eastern Kenya were analyzed using 110 polymorphic markers of six AFLP primer combinations. The highest percentage of polymorphic loci of 80.0% was observed in the Bob Harries population and the least, 67.3%, from Thika population. Expected heterozygosity was also highest in the Bob Harries population. Phylogenetic analysis based on UPGMA of genetic distances of the five populations revealed the Bob Harries population, the source of macadamia germplasm in Kenya to be closely related only to the Kirinyaga population and distantly related to the Thika population, a preservation block of selected superior germplasm. Cluster analysis based on the 26 accessions did not distinctly separate the species indicating a high level of heterozygosity and genetic mixture of the original gene pool used in Kenya. Analysis of molecular variance further indicated higher genetic variation within the populations, 96%, than among the populations, 4%, and high genetic differentiation among the accessions (theta=0.1909). The results of this study indicate high genetic diversity within the Kenyan macadamia germplasm. This information can immediately be used for enhanced breeding and more effective sampling of populations for conservation.