Assessment of genetic diversity of selected cowpea landraces from Nigeria based on simple sequence repeat markers

Understanding the genetic diversity of cowpea (Vigna unguiculata L. Walp.) landraces is useful for effective characterization and ex-situ conservation of germplasm. The analysis of genetic diversity of eighteen cowpea landraces collected from five agro-ecological zones in Nigeria was reported in this study. Five individuals per landrace were genotyped with six polymorphic microsatellite markers. Three to 5 alleles with a mean of 3.833 were detected. Mean Polymorphic information content (PIC) and observed heterozygosity of the markers were 0.5721 and 0.2433, respectively. Analysis of Molecular Variance (AMOVA) showed that variation due to agroecological zone constituted 24%, while variations among and within landraces as well as within individuals constituted 25%, 17% and 33%, respectively. Landraces collected from the humid rainforest zone showed high within landrace diversity and were not significantly different (P ≥ 0.001) from other landraces collected from the same zone. Landraces from the savannah zones showed low within landrace diversity and homozygous across all loci. Consequently, among landrace diversity was higher in the savannah zone with landraces collected from guinea savannah been the most diverse, followed by landraces from the derived savannah and Sudan savannah. Mantel test showed positive and significant correlation (r= 0.377, p= 0.01) between genetic and geographical distance of landrace collections. The findings are important for up-to-date characterizations of cowpea germplasm in Nigeria for improved breeding programs. Keywords: Genetic diversity, cowpea, Alleles, Polymorphic information content, AMOVA

Extraction of high quality DNA from Cola nitida and Cola acuminata

Cola spp. are highly valued in African traditional medicine because they are rich in common and rare phytochemicals. They are important raw material in both the food and pharmaceutical industries. It is often difficult to isolate DNA from many tropical species with high polyphenolic and alkaloid contents. Many published protocols for nucleic acid isolation failed to yield sufficiently good quality amounts of DNA from leaf samples of C. nitida and C. acuminata for analysis. In this research, a simple and fast sodium dodecyl sulphate (SDS) – based extraction protocol have therefore been developed for routine isolation of high-quality nucleic acids from these Cola species by modifying established protocols. Genomic DNA obtained was found suitable for PCR amplifications

Colchicine-induced variations in survival rate and morphological characteristics of water yam (Dioscorea alata)

The effects of 0.2% aqueous solution of colchicine on the survival rate and morphological features of five accessions of Dioscorea alata were investigated. Sprouting buds of two month old plants were treated with 0.2% colchicine and their performances were monitored until maturity. Survival of buds was lower in all the treated plants (ranging between 6.25% and 8.75%) compared to controls (between 13.25% and 15.25%). However, colchicine treated vines had higher survival rates when exposed to drought (13.8% - 24%) compared to controls (7.2% - 16.69%). Higher number of leaves, larger leaf width and fewer numbers of stomata were observed among the treated plants. The survival of buds between the treated plants and control plants was signifi cantly different at P ≤ 0.05. The treated and non-treated plants were also signifi cantly different at P ≤ 0.05 for leaf width in accessions TDa02/00246, TDa98/00116 and TDa99/00240, and for stomata number in accessions TDa02/00151 and TDa02/00246. Our results suggest that colchicine can be used to induce mutagenic changes in yam which may be of agronomic importance in the production of the crop

In vitro androgenesis in Dioscorea rotundata Poir

Conventional breeding takes several years to achieve homozygosity in yam. Anther culture technology is a rapid and viable alternative for the development of homozygous plants, therefore optimizing key parameters for the establishment of an efficient protocol for yam anther culture is indispensable. Influence of anther developmental stages, sterilization treatments, concentration and combination of growth hormones on in vitro androgenesis of Dioscorea rotundata was investigated. Anthers were grouped into five developmental stages from least to most matured, designated as Sml to Sm5. The criteria used for the grouping were anther size, color and location on the inflorescence. Three sterilization treatments were designed to produce contamination-free anther cultures. These are NaOCI (sodiumhypochlorite), NaOCI with 70% ethanol pretreatment and NaOCI with ethanol and Tween 20. Sterilization of spikes was carried out using 2%, 3%, 5%,10%, 15% and 20% NaOCI concentrations for 5, 10, 15 and 20 minutes. Anthers were cultured on MS media supplemented with 2, 4-D or NAA at three concentrations (1.0, 1.5 and 2.0 mg/l) combined with Kinetin or 6-BAP at two concentrations (0.5 1.0 mg/l). Viable and contamination-free anthers were recorded two weeks after culture while callus production was recorded four weeks after culture. Anthers of Sm2 had100% viabitity under dark condition. Fifteen percent NaOCI sterilization for 20 minutes with ethanol gave 100% contamination-free anthers. Optimum callus (15%) was produced when 1.5 mg/I 2, 4-D was combined with 1.0 mg/I 6-BAP. The optimal conditions derived for yam anther culture would facilitate the production of haploid genotypes

Analysis of genetic diversity of African yam bean using SSR markers derived from cowpea

African yam bean, AYB (Sphenostylis stenocarpa Hochst. ex. A. Rich Harms), is a tuberous legume of tropical Africa. AYB has the potential to significantly boost food security due to its considerable nutritional qualities. However, the crop is underutilized. To efficiently utilize AYB genetic resources for its improvement, it is necessary to understand the crop’s diversity. This study investigated the amplification ability of 36 cowpea simple sequence repeat (SSR) primers across AYB genomic DNA, extracted from 67 accessions. Thirteen (36%) of the cowpea SSRs showed transferability in AYB. Eight of these SSRs amplified above 60% of AYB accessions and generated 55 polymorphic fragments with an average of 6.9 per primer. Polymorphic information content ranged from 0.6691 to 0.8857 with an average of 0.7791. This study also assessed the genetic diversity within 67 AYB accessions using eight cowpea (Vigna unguiculata L. Walp)-derived SSR primers. The result revealed a high level of genetic diversity with simple matching coefficient ranging from 0.458 to 1.000. A dendrogram depicting three main clusters was generated based on unweighted pair group method with arithmetic average. Cluster 1 was the most diverse with a dissimilarity range of 0.517–1.000. The level of genetic diversity revealed in this study indicates that the studied AYB germplasm can be exploited for genetic improvement. Additionally, the transferable markers will aid AYB genome research and also make possible the comparative mapping between AYB and cowpea.African yam bean, AYB (Sphenostylis stenocarpa Hochst. ex. A. Rich Harms), is a tuberous legume of tropical Africa. AYB has the potential to significantly boost food security due to its considerable nutritional qualities. However, the crop is underutilized. To efficiently utilize AYB genetic resources for its improvement, it is necessary to understand the crop’s diversity. This study investigated the amplification ability of 36 cowpea simple sequence repeat (SSR) primers across AYB genomic DNA, extracted from 67 accessions. Thirteen (36%) of the cowpea SSRs showed transferability in AYB. Eight of these SSRs amplified above 60% of AYB accessions and generated 55 polymorphic fragments with an average of 6.9 per primer. Polymorphic information content ranged from 0.6691 to 0.8857 with an average of 0.7791. This study also assessed the genetic diversity within 67 AYB accessions using eight cowpea (Vigna unguiculata L. Walp)-derived SSR primers. The result revealed a high level of genetic diversity with simple matching coefficient ranging from 0.458 to 1.000. A dendrogram depicting three main clusters was generated based on unweighted pair group method with arithmetic average. Cluster 1 was the most diverse with a dissimilarity range of 0.517–1.000. The level of genetic diversity revealed in this study indicates that the studied AYB germplasm can be exploited for genetic improvement. Additionally, the transferable markers will aid AYB genome research and also make possible the comparative mapping between AYB and cowpea.African yam bean, AYB (Sphenostylis stenocarpa Hochst. ex. A. Rich Harms), is a tuberous legume of tropical Africa. AYB has the potential to significantly boost food security due to its considerable nutritional qualities. However, the crop is underutilized. To efficiently utilize AYB genetic resources for its improvement, it is necessary to understand the crop’s diversity. This study investigated the amplification ability of 36 cowpea simple sequence repeat (SSR) primers across AYB genomic DNA, extracted from 67 accessions. Thirteen (36%) of the cowpea SSRs showed transferability in AYB. Eight of these SSRs amplified above 60% of AYB accessions and generated 55 polymorphic fragments with an average of 6.9 per primer. Polymorphic information content ranged from 0.6691 to 0.8857 with an average of 0.7791. This study also assessed the genetic diversity within 67 AYB accessions using eight cowpea (Vigna unguiculata L. Walp)-derived SSR primers. The result revealed a high level of genetic diversity with simple matching coefficient ranging from 0.458 to 1.000. A dendrogram depicting three main clusters was generated based on unweighted pair group method with arithmetic average. Cluster 1 was the most diverse with a dissimilarity range of 0.517–1.000. The level of genetic diversity revealed in this study indicates that the studied AYB germplasm can be exploited for genetic improvement. Additionally, the transferable markers will aid AYB genome research and also make possible the comparative mapping between AYB and cowpea.African yam bean, AYB (Sphenostylis stenocarpa Hochst. ex. A. Rich Harms), is a tuberous legume of tropical Africa. AYB has the potential to significantly boost food security due to its considerable nutritional qualities. However, the crop is underutilized. To efficiently utilize AYB genetic resources for its improvement, it is necessary to understand the crop’s diversity. This study investigated the amplification ability of 36 cowpea simple sequence repeat (SSR) primers across AYB genomic DNA, extracted from 67 accessions. Thirteen (36%) of the cowpea SSRs showed transferability in AYB. Eight of these SSRs amplified above 60% of AYB accessions and generated 55 polymorphic fragments with an average of 6.9 per primer. Polymorphic information content ranged from 0.6691 to 0.8857 with an average of 0.7791. This study also assessed the genetic diversity within 67 AYB accessions using eight cowpea (Vigna unguiculata L. Walp)-derived SSR primers. The result revealed a high level of genetic diversity with simple matching coefficient ranging from 0.458 to 1.000. A dendrogram depicting three main clusters was generated based on unweighted pair group method with arithmetic average. Cluster 1 was the most diverse with a dissimilarity range of 0.517–1.000. The level of genetic diversity revealed in this study indicates that the studied AYB germplasm can be exploited for genetic improvement. Additionally, the transferable markers will aid AYB genome research and also make possible the comparative mapping between AYB and cowpea

Candidate gene sequencing and validation of SNP markers linked to carotenoid content in cassava (Manihot esculenta Crantz)

Cassava is a widely grown staple in Sub-Saharan Africa and consumed as a cheap source of calories, but the crop is deficient in micronutrients including pro-vitamin A carotenoids. This challenge is currently being addressed through biofortification breeding that relies on phenotypic selection. Genebased markers linked to pro-vitamin A content variation are expected to increase the rate of genetic gain for this critical trait. We sequenced four candidate carotenoid genes from 167 cassava accessions representing the diversity of elite breeder lines from IITA. Total carotenoid content was determined using spectrophotometer and total β-carotene was quantified by highperformance liquid chromatography. Storage root yellowness due to carotenoid pigmentation was assessed. We carried out candidate gene association analysis that accounts for population structure and kinship using genome-wide single nucleotide polymorphisms (SNPs) generated through genotyping-by-sequencing. Significant SNPs were used to design competitive allele-specific PCR assays and validated on the larger population for potential use in markerassisted selection breeding. Candidate gene sequencing of the genes β-carotene hydroxylase (crtRB), phytoene synthase (PSY2), lycopene epsilon cyclase (lcyE), and lycopene beta cyclase (lcyB) yielded a total of 37 SNPs. Total carotenoid content, total β-carotene, and color parameters were significantly associated with markers in the PSY2 gene. The SNPs from lcyE were significantly associated with color while those of lcyB and crtRB were not significantly associated with carotenoids or color parameters. These validated and breeder-friendly markers have potential to enhance the efficiency of selection for high β-carotene cassava, thus accelerating genetic gain