Incorporation of resistance to angular leaf spot and bean common mosaic necrosis virus diseases into adapted common bean (Phaseolus vulgaris L.) genotype in Tanzania

Angular leaf spot (ALS) caused by the fungus Pseudocercospora griseola and Bean common mosaic and necrosis virus (BCMV/BCMNV) are important diseases of common bean in Tanzania that can cause severe yield reduction when uncontrolled. This study was conducted to incorporate resistant genes for ALS and BCMV/BCMNV diseases into adapted, market class and farmers and consumers preferred bean genotype using marker assisted selection. The parents Mexico 54 and UBR(25)95 donor of Phg-2 and I/bc-3 genes for ALS and BCMV/BCMNV, respectively were used for the recipient being Kablanketi. In selection, SCAR markers SNO2, ROC11 and SW13 linked to Phg-2, bc-3 and I gene, respectively were used. A parallel backcrossing (modified double cross) procedure was used. The F1, F2 and backcrosses from single crosses were characterized. The Chi square values for ALS were 0.081 (P<0.776) and 0.017 (P<0.896) and for BCMNV were 1.609 (P<0.205) and 1.2 (P<0.273) for molecular and phenotypic screening, respectively. The resistance to ALS and BCMNV was found to be monogenic and the genes involved are dominant and recessive, respectively. The heritability of ALS was found to be high (0.772) implying that selection for ALS can be done early in segregating populations. High correlation values, r = 0.741 and 0.624 for ALS and BCMNV, were obtained between phenotypic and molecular data, indicating high reliability for markers. In selection, it was possible to select lines with multiple disease resistances. This work signified the use of MAS for multiple gene screening.Keywords: Common bean, Pseudocercospora griseola, marker assisted selection, genotype, inheritanceAfrican Journal of Biotechnology Vol. 12(27), pp. 4343-435

Assessing the effect of sample storage time on viral detection using a rapid and cost-effective CTAB-based extraction method

Abstract Background Cassava leaf samples degrade quickly during storage and transportation from distant areas. Proper sampling and efficient, low-cost storage methods are critical for obtaining sufficient quality DNA and RNA for plant virus epidemiology and improving disease control understanding. This is useful when samples are collected from remote areas far from a laboratory or in developing countries where money and materials for virus diagnostics are scarce. Results The effect of sample storage duration on nucleic acid (N.A.) quality on virus detection was investigated in this study. A simple, rapid, and cost-effective CTAB-based approach (M3) for single N.A. extraction was optimized and tested alongside two existing CTAB-based methods (M1 and M2) for N.A. extraction from fresh and herbarium cassava leaves stored for; 1, 8, 26, and 56 months. The amount and quality of DNA and RNA were determined using Nanodrop 2000 c U.V.–vis Spectrophotometer and agarose gel electrophoreses. The sample degradation rate was estimated using a simple mathematical model in Matlab computational software. The results show no significant difference in mean DNA concentration between M1 and M2 but a significant difference between M3 and the other two methods at p < 0.005. The mean DNA concentration extracted using M3 was higher for 1 and 8 months of leave storage. M3 and M2 produced high concentrations at 26 and 56 months of leave storage. Using a developed scale for quality score, M3 and M2 produced high-quality DNA from fresh samples. All methods produced poor-quality DNA and RNA at 8 and 26 months of leave storage and no visual bands at the age of 56 months. Statistically, there was a significant difference in the mean DNA quality between M1 and M2, but there was no significant difference between M3 and the other two methods at p < 0.005. However, Cassava brown streak virus (CBSV) and Ugandan cassava brown streak virus (UCBSV) were readily detected by RT-PCR from RNA isolated using M3. The quality of DNA declined per storage time at 0.0493 and 0.0521/month, while RNA was 0.0678 and 0.0744/month. Compared to the existing two methods, modified CTAB extracted enough high-quality N.A. in one-third the time of the existing two methods. Conclusion Our method provides cost-effective, quick, and simple processing of fresh and dry samples, which will quicken and guide the decision on when and what type of sample to process for plant disease management and surveillance actions