Genetic engineering approach in the control of cassava brown streak and bacterial blight diseases
Njiru, Joshua Mugendi
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Cassava is the fifth most important food crop in the world. The importance of cassava in Africa cannot be overstated. As a drought-tolerant crop that does well in poor soils, it enhances household food security and is a source of income that provides livelihood to 800 million people globally. However cassava is faced by numerous challenges of which pests and diseases are the greatest problems to its production across East and Central Africa (ECA) sub-regions. Cassava is vulnerable to at least 20 different viral diseases, cassava brown streak disease (CBSD), being one of the most damaging viral diseases in East Africa. Cassava bacterial blight (CBB) caused by Xanthomonas axonopodis pv. manihotis (Xam) on the other hand is the most destructive bacterial disease in all cassava growing areas of the world. The overall objective of this study was to develop cassava cultivars that would be resistant to CBSD and CBB through genetic engineering approaches'. A robust reproducible genetic transformation and regeneration protocol is a prerequisite for any plant transformation system. Therefore this study optimized a cultivar independent transformation protocol using friable embryogenic callus (FEC) as the explant. The optimized protocol was then used to transform selected cultivars for CBB and CBSD resistance. RNA interference (RNAi) gene construct p5001 expressing inverted repeats of the full length coat proteins (FL-CP) from UCBSV and CBSV was used to transform farmer preferred cultivar TME 14 for CBSD resistance. Hypersensitivity response assisting protein (Hrap) gene was used to transform cultivar cv. 60444 for CBB resistance. The transformation frequency for Mkombozi was 20.8%, 32.3% for Albert and 39.6% for cv. 60444. The developed protocol was repeated in· TME 14 and a transformation frequency of 47.5% was achieved. Through top graft inoculation, transgenic TME 14 lines were evaluated for CBSD resistance and 85.7% of the screened plants were immune to CBSD and did not develop disease symptoms. After inoculation of Xam in cv. 60444 Hrap transgenic lines, 58.3% of screened events were totally immune to CBB showing no disease symptoms at 60 days post inoculation and 33.3% had delayed symptoms with reduced disease severity when compared to the control non-transgenic plants. Post-transcriptional gene silencing (PTGS) offer significant potential for controlling RNA plant viruses. In this study, RNAi approach was used to developing CBSD resistant lines by accumulating transgenically derived siRNAs in the plants. The constitutive expression of the Hrap in cv. 60444 generated durable resistance against CBB. These findings show that CBSD and CBB can be effectively controlled using genetic engineering approaches' .