Transformation and Regeneration Protocol for Two Farmer Preferred Open Pollinated Tropical Maize (Zea Mays) Varieties

Article PurchasedAbstract: In vitro regeneration of open pollinated varieties (OPVs) Kakamega Striga Tolerant Population 94 (KSTP’94) and ‘Namba Nane’ alongside a tropical inbred line (CML144) was evaluated using immature zygotic embryos as explants. Four callus induction media (CIM) regimes; Murashige and Skoog (MS), Linsmaier and Skoog (LS), Chu (N6) and N6*(N6 medium fortified with 0.35 gL-1 L-proline and 0.8 mgL-1 AgNO3) were evaluated for their potential to induce callus in the three genotypes. All the media were supplemented with sucrose and five levels of 2, 4-Dichlorophenoxyacetic acid (2, 4-D) (0.5, 1.0, 1.5, 2.0 and 2.5 mgL-1). Resulting calli were matured on MS and N6 basal media supplemented with 60 g/L sucrose and similar concentration levels (0.5, 1.0, 1.5, 2.0 and 2.5 mgL-1) of 2, 4-D while the subsequent embryogenic calli were regenerated on hormone-free media. Transformability of these varieties was assessed via histochemical analysis of β-glucuronidase (GUS) reporter gene following Agrobacterium-mediated transformation. Statistical analyses were done using Statistical Analysis Software (SAS) and Graphpad Prism softwares with mean separations achieved at 95% confidence intervals. Of the 2 OPVs, KSTP’94 recorded the highest callus induction frequency (84.4%) while Namba Nane (45.6%) had the lowest. Similarly, KSTP, 94 had the highest mean of mature somatic embryos (59.7%) while Namba Nane recorded the lowest (16.4%). Assessment of regeneration frequencies from embryogenic calli revealed no significant differences among the 3 lines although CML 144 had the highest mean number of juvenile plantlets (36.7%). Analysis of transformation frequency (upon selection of calli on media with basta) showed that Namba Nane recorded the lowest transformation frequency (average 13.5%) some words missing. Transformation frequency (based on GUS positive calli) of these varieties ranged from 0.8 to 2.1%. This work therefore provides an empirical platform for potential introduction of useful genes into these varieties

Midgut bacterial diversity analysis of laboratory reared and wild Anopheles gambiae and Culex quinquefasciatus mosquitoes in Kenya

Open Access JournalMidgut symbiotic bacteria are known to play fundamental roles in the biology of mosquitoes, however knowledge of midgut bacterial communities associated with mosquitoes is scanty due to limitation of the isolation techniques based on culturing. In this study, the composition and diversity of midgut bacteria in field collected and lab reared adult female Anopheles gambiae and Culex quinquefasciatus mosquitoes was explored using the Illumina sequencing. Deoxyribonucleic acid was isolated from the pooled midgut extracts and their 16S rRNA gene sequenced using Illumina sequencing platform. Operational taxonomic units (OTUs) were analyzed using QIIME 1.8.0; taxonomy was assigned using BLASTn against SILVA 119 and hierarchical clustering was done using R program software. Out of the total number of sequence reads obtained, 145 OTUs were realized at 3% genetic distance. The 145 OTUs spanned 12 phyla; Proteobacteria, Firmicutes, Cyanobacteria, Euryarchaeota, Gemmatimonadetes, Spirochaetae, Archeabacteria Verrucomicrobia, Chloroflexi, Bacteriodetes, Acidobacteria and Actinobacteria. Microbial community composition based on OTUs showed significant difference between field collected and lab reared mosquitoes (ᵪ2 = 45.0799, p = 3.2 × 10-5). Similarly, there was a significant difference in community composition at OTU level between Anopheles gambiae and Culex quinquefasciatus (ᵪ2 = 31.2257, p = 7.7 × 10-4). The bacterial composition and diversity appeared to be influenced by the environment and the species of the mosquitoes

Screening for potential Striga hermonthica fungal and bacterial biocontrol agents from suppressive soils in Western Kenya

Published online: 18 July 2017Striga hermonthica is a hemiparasitic weed that causes huge grain yield losses to small-scale farmers in Africa. Effective biocontrol agents against S. hermonthica can sustainably mitigate these losses. This study characterized the biocontrol potential of culturable fungal and bacterial isolates from S. hermonthica suppressive soils of western Kenya. These isolates were screened for their ability to produce antibiotic compounds and extra cellular enzymes and also their ability to cause S. hermonthica seed decay. Genomic DNA of the selected bacterial and fungal isolates was extracted and partial characterization of 16S rRNA and 18S rRNA genes performed respectively. Analysis show that antibiosis and enzymatic properties of potential biocontrol isolates correlated positively. Isolate KY041696 recorded high antibiosis, enzymatic and seed decay values. This study also revealed that bioactive bacterial isolates belonged to Bacillus, Streptomyces and Rhizobium genera. In this study, no fungal isolate caused S. hermonthica seed decay. This study therefore provides baseline information on the potential biocontrol microbes against S. hermonthica in Western Kenya that could be exploited further in the management of the weed

Morphological diversity of farmers’ and improved potato (Solanum tuberosum) cultivars growing in Eritrea

Farmers’ and improved potato (Solanum tuberosum L.) cultivars growing in Eritrea are main sources of food and income to many growers. The current study was proposed to characterize 17 farmers’ and 4 imported cultivars of potato using 33 morphological descriptors. Planting was done in two geographically distinct locations, HAC and Asmara, Eritrea. The experiment was laid out in a randomized complete block design with three replications having 18 plants per plot. Plants grown at HAC emerged early (24.52 days) and reach maturity (94.84 days) while at Asmara it took 43.77 and 123.59 days, respectively. However, yield was higher in Asmara (0.49 kg/plant) compared to HAC (0.37 kg/plant). An accession having many and longer stems was associated with more tuber production, but inversely related to yield. Similarly, accessions with higher stem thickness and tuber size were associated with high yields. The PCA analysis indicated that the first four components explained about 85% of the total variability among the studied materials. The PCA clustered the materials in to four main groups (GI, GII, GIII, GIV) mainly explained by flowering patterns and yield related descriptors. The work has provided useful information on morphological characteristics of the farmer’s potato to avoid duplication of resources and identify promising materials for future breeding program

Genetic diversity assessment of farmers’ and improved potato (Solanum tuberosum) cultivars from Eritrea using simple sequence repeat (SSR) markers.

Sixty three potato clones (51 farmers’ and 12 varieties) from Eritrea, 18 and 12 varieties from Kenya and Rwanda, respectively were characterized using 12 highly polymorphic simple sequence repeat (SSR) markers. The study was designed to assess the genetic diversity and varietal distinctness among the different samples. In total, 91 alleles ranging between 2 (STM1053) to 13 (STM0031) alleles per marker were scored. All but 97.8 SSR markers were highly polymorphic with an average PIC value of 0.87 (0.51 to 0.98). All of the 51 farmers’ cultivars were clearly distinct from each other. Samples from Eritrea showed the highest genetic diversity as explained by the diversity index (h). The principal coordinate analysis (PCoA) revealed that the local farmers’ Eritrean samples are different from the Kenyan, Rwandese and even the imported varieties. Genetic distance analysis generated three clusters correlating with the PCoA findings. Cluster I consisted of 45 samples with 6 sub-clusters; Cluster II consisted of 29 samples with a majority (26) from Eritrea while cluster III consisted of 19 samples. Potato materials from Eritrea appeared to cluster separately from the other samples, which reflects a contribution from the Tuberosum germplasm prominent in temperate regions, unlike from the Andigenum germplasm for Kenyan and Rwandan potato materials. Most of the Eritrean samples in cluster I are farmers’ cultivars with intermediate maturity, good performance and better tuber quality characteristics. Cluster II contains mainly the imported variety from Eritrea characterized by late emergence and late maturity. The Kenyan and Rwandese were grouped mainly in Cluster III. In summary, the farmers’ cultivars are distinct from the Kenyan and Rwandese materials and represent more genetic diversity than the varieties imported into Eritrea. This finding is of interest to national breeding program to use the farmer’s materials as source of genetic variation for traits of interest

Genetic diversity and population structure of Eritrean pepper (Capsicum species) as revealed by SSR markers

Pepper (Capsicum spp.) is one of the most important vegetable crops and the most widely used spice worldwide including Eritrea. Diversity studies are an essential step for crop breeding and improvement. Therefore, the objectives of the study were to determine the diversity and population structure of local Eritrean pepper collected from farmers and research institutions and to evaluate the relatedness of the Eritrean pepper with accessions obtained from five other countries. A total of 407 individual pepper plants from 150 seed samples were evaluated using 28 SSR markers. The results showed that varieties maintained in situ by farmers were heterogeneous. Diversity parameters indicated extensive genetic variation among the Eritrea genotypes. The 28 markers revealed a total of 352 alleles with an average of 13 alleles per marker. Mean Polymorphic Information Content was 0.62 and, mean Observed Heterozygosity was 0.41. The analysis of molecular variance showed only 10% variation was among populations, 30% among individuals within populations and 60% within individuals. This can be explained by the high mean number of effective migrants (2.25) that ranged from 1.01 to 10.45 among populations indicating movement of germplasm among farmers in different geographic and agro-ecological regions. A factor analysis, neighbour joining clustering and the model based clustering (Structure) classified the 407 individuals into 3 groups. However, in the model based clustering; increasing the number of populations to 4 (K=4 ) caused all non-Eritrean genotypes to fall in a separate cluster suggesting availability of potentially rich diversity within the Eritrean populations justified by the large number of private alleles observed