Unlocking the potential of tropical root crop biotechnology in east Africa by establishing a genetic transformation platform for local farmer-preferred cassava cultivars.
[en] Cassava genetic transformation capacity is still mostly restricted to advanced laboratories in the USA, Europe and China; and its implementation and maintenance in African laboratories has remained scarce. The impact of transgenic technologies for genetic improvement of cassava will depend largely on the transfer of such capabilities to researchers in Africa, where cassava has an important socioeconomic niche. A major constraint to the development of genetic transformation technologies for cassava improvement has been the lack of an efficient and robust transformation and regeneration system. Despite the success achieved in genetic modification of few cassava cultivars, including the model cultivar 60444, transgenic cassava production remains difficult for farmer-preferred cultivars. In this study, a protocol for cultivar 60444 developed at ETH Zurich was successfully implemented and optimized to establish transformation of farmer-preferred cassava cultivars popular in east Africa. The conditions for production and proliferation of friable embryogenic calli (FEC) and Agrobacterium-mediated transformation were optimized for three east African farmer-preferred cultivars (Ebwanatereka, Kibandameno and Serere). Our results demonstrated transformation efficiencies of about 14-22 independent transgenic lines per 100 mg of FEC for farmer-preferred cultivars in comparison to 28 lines per 100 mg of the model cultivar 60444. The presence, integration and expression of the transgenes were confirmed by PCR, Southern blot analysis and histochemical GUS assay. This study reports the establishment of a cassava transformation platform at International Institute of Tropical Agriculture (IITA) hosted by Biosciences eastern and central Africa (BecA) hub in Kenya and provides the basis for transferring important traits such as virus resistance and prolonged shelf-life to farmer-preferred cultivars in east Africa. We anticipate that such platform will also be instrumental to transfer technologies to national agricultural research systems (NARS) in sub-Saharan Africa.
Unlocking the potential of tropical root crop biotechnology in east Africa by establishing a genetic transformation platform for local farmer-preferred cassava cultivars.
Alicai, T., Omongo, C. A., Maruthi, M. N., Hillocks, R. J., Baguma, Y., Kawuki, R., et al. (2007). Re-emergence of cassava brown streak disease in Uganda. Plant Dis. 91, 24-29. doi: 10.1094/PD-91-0024
Baba, A. I., Nogueira, F. C. S., Pinheiro, C. B., Brasil, J. N., Jereissati, E. S., Juca, T. L., et al. (2008). Proteome analysis of secondary somatic embryogenesis in cassava (Manihot esculenta). Plant Sci. 175, 717-723. doi: 10.1016/j.plantsci. 2008.07.014
Barampuram, S., andZhang, Z. J. (2011). Recent advances in plant transformation. Methods Mol. Biol. 701, 1-35. doi: 10.1007/978-1-61737-957-4_1
Bull, S. E., Owiti, J. A., Niklaus, M., Beeching, J. R., Gruissem, W., and Vanderschuren, H. (2009). Agrobacterium-mediated transformation of friable embryogenic calli and regeneration of transgenic cassava. Nat. Protoc. 4, 1845-1854. doi: 10.1038/nprot.2009.208
Ceballos, H., Iglesias, C. A., Perez, J. C., and Dixon, A. G. O. (2004). Cassava breeding: opportunities and challenges. Plant Mol. Biol. 56, 503-516. doi: 10.1007/s11103-004-5010-5
Chetty, C. C., Rossin, C. B., Gruissem, W., Vanderschuren, H., and Rey, M. E. C. (2013). Empowering biotechnology in southern Africa: establishment of a robust transformation platform for the production of transgenic industry- preferred cassava. New Biotechnol. 30, 136-143. doi: 10.1016/j.nbt.2012.04.006
Donald Danforth Plant Science Center. (2006). News Release (Danforth Center Cassava Update, Friday May 26th, 2006). Available online at: www.danforthcenter.org/wordpress/?page_id=395&pid=1474 (Accessed November 10, 2012).
Ezezika, O. C., Daar, A. S., Barber, K., Mabeya, J., Thomas, F., Deadman, J., et al. (2012). Factors influencing agbiotech adoption and development in sub-Saharan Africa. Nat. Biotechnol. 30, 193. doi: 10.1038/nbt0212-193a
Fregene, M., and Puonti-Kaerlas, J. (2002). "Cassava biotechnology" in Cassava: Biology, Production and Utilization, eds R. J. Hillocks, J. M. Thresh, and A. C. Bellotti (Wallingford: CAB International), 179-207. doi: 10.1079/9780851995243.0179
Gelvin, S. B. (2003). Agrobacterium-mediated plant transformation: the biology behind the "gene jockeying" tool. Microbiol. Mol. Biol. Rev. 67, 16-37. doi: 10.1128/MMBR.67.1.16-37.2003
Gonzalez, A. E., Schopke, C., Taylor, N. J., Beachy, R. N., and Fauquet, C.M.(1998). Regeneration of transgenic cassava plants (Manihot esculenta Crantz) through Agrobacterium-mediated transformation of embryogenic suspension cultures. PlantCellRep. 17, 827-831. doi: 10.1007/s002990050492
Gresshoff, P., and Doy, C. (1974). Derivation of a haploid cell line from Vitis vinifera and the importance of the stage of meiotic development of anthers for haploid culture of this and other genera. Zeitschrift fur Pflanzenphysiologie 73, 132-141. doi: 10.1016/S0044-328X(74)80084-X
Hankoua, B. B., Ng, S. Y. C., Puonti-Kaerlas, J., Fawole, I., Dixon, A. G. O., and Pillay, M. (2005). Regeneration of a wide range of African cassava genotypes via shoot organogenesis from cotyledon of maturing somatic embryos and conformity of the field-established regenerants. Plant Cell Tissue Organ Cult. 81, 200-211. doi: 10.1007/s11240-005-0514-5
Jansson, C., Westerbergh, A., Zhang, J., and Xinwen Hu, S. C. (2009). Cassava, a potential biofuel crop in (the) People's Republic of China. Appl. Energy 86, S95-S99. doi: 10.1016/j.apenergy.2009.05.011
Jefferson, R. A. (1987). Assaying chimeric genes in plants: the GUS gene fusion system. PlantMol. Biol. Rep. 5, 387-405. doi: 10.1007/BF02667740
Kawuki, R. S., Pariyo, A., Amuge, T., Nuwamanya, E., Ssemakula, G., Tumwesigye, S., et al. (2011). A breeding scheme for local adoption of cassava (Manihot esculenta Crantz). J. PlantBreed. Crop Sci. 3, 120-130.
Koehorst-van Putten, H. J. J., Sudarmonowati, E., Herman, M., Pereira-Bertram, I. J., Wolters, A. M. A., Meima, H., et al. (2012). Field testing and exploitation of genetically modified cassava with low-amylose or amylose- free starch in Indonesia. Transgenic Res. 21, 39-50. doi: 10.1007/s11248-011- 9507-9
Lacombe, S., Rougon-Cardoso, A., Sherwood, E., Peeters, N., Dahlbeck, D., van Esse, H. P., et al. (2010). Interfamily transfer of a plant pattern-recognition receptor confers broad-spectrum bacterial resistance. Nat. Biotechnol. 28, 365-369. doi: 10.1038/nbt.1613
Legg, J. P., Jeremiah, S. C., Obiero, H. M., Maruthi, M. N., Ndyetabula, I., Okao- Okuja, G., et al. (2011). Comparing the regional epidemiology of the cassava mosaic and cassava brown streak virus pandemics in Africa. Virus Res. 159, 161-170. doi: 10.1016/j.virusres.2011.04.018
Li, H. Q., Sautter, C., Potrykus, I., and Pounti-Kaerlas, J. (1996). Genetic transformation ofcassava (Manihotesculenta Crantz). Nat. Biotechnol. 14, 736-740. doi: 10.1038/nbt0696-736
Liu, J., Zheng, Q., Ma, Q., Gadidasu, K. K., and Zhang, P. (2011). Cassava genetic transformation and its application in breeding. J. Integr. PlantBiol. 53, 552-569. doi: 10.1111/j.1744-7909.2011.01048.x
Machuka, J. (2001). Agricultural Biotechnology for Africa. African scientists and farmers must feed their own people. Plant Physiol. 126, 16-19. doi: 10.1104/pp.126.1.16
Mendes, B. M. J., Cardoso, S. C., Boscariol-Camargo, R. L., Cruz, R. B., Mourao Filho, F. A. A., and Bergamin Filho, A. (2010). Reduction in susceptibility to Xanthomonas axonopodis pv. citri in transgenic Citrus sinensis expressing the rice Xa21 gene. PlantPathol. 59, 68-75. doi: 10.1111/j.1365-3059.2009.02148.x
Namukwaya, B., Tripathi, L., Tripathi, J. N., Arinaitwe, G., Mukasa, S. B., and Tushemereirwe, W. K. (2012). Transgenic banana expressing Pflp gene confers enhanced resistance to XanthomonasWilt Disease. TransgenicRes. 12, 855-865. doi: 10.1007/s11248-011-9574-y
National CassavaProgramme-UgandaReport. (2005). Integration ofGender, SocioEconomic and Market Factors for Expanded Cassava Production in the Teso and Lango Farming Systems. NARO/DFID CORF3029 Project Terminal Report.
Ntawuruhunga, P., and Legg, J. P. (2007). New Spread ofCassava Brown Streak Virus Disease and its Implications for the Movement of Cassava Germplasm in the East and Central African Region, 2007. 1-6. Available online at: http://c3project.iita.org/Doc/A25-CBSDbriefMay26.pdf
Ogunjobi, A. A., and Dixon, A. (2006). Molecular variation in population structure of Xanthomonas axonopodis pv. manihotis in the southeastern Nigeria. Afr. J. Biotechnol. 5, 1868-1872.
Ogwok, E., Odipio, J., Halsey, M., Gaitan-Solis, E., Bua, A., Taylor, N., et al. (2012). Transgenic RNA interference (RNAi)-derived field resistance to cassava brown streak disease. Mol. Plant Pathol. 13, 1019-1031. doi: 10.1111/j.1364- 3703.2012.00812.x
Ombori, O., Muoma, J. V. O., and Machuka, J. (2013). Agrobacterium-mediated genetic transformation of selected tropical inbred and hybrid maize (Zea mays L.) lines. Plant Cell, Tissue Organ Cult. 113, 11-23. doi: 10.1007/s11240-012- 0247-1
Owiti, J., Grossmann, J., Gehrig, P., Dessimoz, C., Laloi, C., Hansen, M. B., Gruissem, W., and Vanderschuren, H. (2011). iTRAQ-based analysis of changes in the cassava root proteome reveals pathways associated with postharvest physiological deterioration. Plant J. 67, 145-156. doi: 10.1111/j.1365- 313X.2011.04582.x
Raemakers, C. J. J. M., Schreuder, M., Pereira, I., Munyikwa, T., Jacobsen, E., and Visser, R. G. F. (2001). Progress made in FEC transformation of cassava. Euphytica 120, 15-24. doi: 10.1023/A:1017574713880
Reilly, K., Bernal, D., Cortes, D. F., Gomez-Vasquez, R., Tohme, J., and Beeching, J. R. (2007). Towards identifying the full set of genes expressed during cassava post-harvest physiological deterioration. PlantMol. Biol. 64, 187-203. doi: 10.1007/s11103-007-9144-0
Reilly, K., Buschmann, R., Tohme, J., and Beeching, J. R. (2004). Oxidative stress responses during post-harvest physiological deterioration. Plant Mol. Biol. 56, 625-641. doi: 10.1007/s11103-005-2271-6
Restrepo, S., Velez, C. M., Duque, M. C., Verdier, V. (2004). Genetic structure and population dynamics of Xanthomonas axonopodis pv. Manihotis in Colombia from 1995 to 1999. Appl. Environ. Microbiol. 70, 255-261. doi: 10.1128/AEM.70.1.255-261.2004
Rudi, N., Norton, G. W., Alwang, J., and Asumugha, G. (2010). Economic impact analysis of marker-assisted breeding for resistance to pests and postharvest deterioration in cassava. Afr. J. Agric. Resour. Econ. 4, 110-122.
Sayre, R., Beeching, J., Cahoon, E., Egesi, C., Fauquet, C., Fellman, J., et al. (2011). The BioCassava Plus Program: biofortification of cassava for sub-Saharan Africa. Annu. Rev. Plant Biol. 62, 251-272. doi: 10.1146/annurev-arplant- 042110-103751
Schopke, C., Taylor, N. J., Carcamo, R., Konan, N. K. K., Marmey, P., Henshaw, G. G., et al. (1996). Regeneration of transgenic cassava plants (Manihot escu- lenta Crantz) from microbombarded embryogenic suspension cultures. Nat. Biotechnol. 14, 731-735. doi: 10.1038/nbt0696-731
Shibata, D. and Liu, Y. G. (2000). Agrobacterium-mediated plant transformation with large DNA fragments. Trends Plant Sci. 5, 354-357. doi: 10.1016/S1360- 1385(00)01689-7
Soni, R., and Murray, J. A. (1994). Isolation of intact DNA and RNA from plant tissues. Anal. Biochem. 218, 474-476. doi: 10.1006/abio.1994.1214
Taylor, N. J., Chavarriaga, P., Raemakers, K., Siritunga, D., and Zhang, P. (2004). Development and application of transgenic technologies in cassava. Plant Mol. Biol. 56, 671-688. doi: 10.1007/s11103-004-4872-x
Taylor, N. J., Kiernan, R. J., Henshaw, G. G., and Blakesley, D. (1997). Improved procedures for producing embryogenic tissues of African cassava genotypes: Implications for genetic transformation. Afr. J. Roots Tuber Crops 2, 200-204.
Taylor, N. J., Masona, M. V., Schopke, C., and Fauquet, C. M. (2002). "Transgenic cassava for food security and economic development," in Transgenic Plants and Crops, eds G. G. Khachatourians, A. McHughen, R. Scorza, W. K. Nip, and Y. H. Hui (NewYork, NY: CRC press), 523-546.
Taylor, T., Gaitan-SoHs, E., Moll, T., Trauterman, B., Jones, T., Pranjal, A., et al. (2012). A High-throughput platform for the production and analysis of transgenic cassava (Manihot esculenta) plants. Trop. Plant Biol. 5, 127-139. doi: 10.1007/s12042-012-9099-4
Toennissen, G. H., Toole, J. C. O., and DeVries, J. (2003). Advances in plant biotechnology and its adoption in developing countries. Curr. Opin. Plant Biol. 6, 191-198. doi: 10.1016/S1369-5266(03)00002-5
Tripathi, J. N., Muwonge, A., and Tripathi, L. (2012). Efficient regeneration and transformation protocol for plantain cultivar 'Gonja Manjaya' (Musa spp. AAB) using embryogenic cell suspension. In vitro Cell Dev. Biol. Plant 48, 216-224. doi: 10.1007/s11627-011-9422-z
Tripathi, L., Mwaka, H., Tripathi, J. N., and Tushemereirwe W. K. (2010). Expression of sweet pepper hrap gene in banana enhances resistance to Xanthomonas campestris pv. musacearum. Mol. Plant Pathol. 11, 721-731. doi: 10.1111/j.1364-3703.2010.00639.x
Vanderschuren, H. (2012). Strengthening African R&D through effective transfer of tropical crop biotech to African Institutions. Nat. Biotechnol. 30, 3-4. doi: 10.1038/nbt.2405
Vanderschuren, H., Alder, A., Zhang, P., and Gruissem, W. (2009). Dosedependent RNAi-mediated geminivirus resistance in the tropical root crop cassava. Plant Mol. Biol. 70, 265-272. doi: 10.1007/s11103- 009-9472-3
Vanderschuren, H., Moreno, I., Anjanappa, R. B., Zainuddin, I. M., and Gruissem, W. (2012). Exploiting the combination of natural and genetically engineered resistance to cassava mosaic and cassava brown streak viruses impacting cassava production in Africa. PLoS ONE 7:e45277. doi: 10.1371/journal.pone. 0045277
Vanderschuren, H., Stupak, M., Futterer, J., Gruissem, W., and Zhang, P. (2007). Engineering resistance to geminiviruses-review and perspectives. Plant Biotechnol. J. 5, 207-220. doi: 10.1111/j.1467-7652.2006.00217.x
Verdier, V., Restrepo, S., Mosquera, G., Jorge, V., and Lopez, C. (2004). Recent progress in the characterization of molecular determinants in the Xanthomonas axonopodis pv. manihotis-cassavainteraction. PlantMol. Biol. 56, 573-584. doi: 10.1007/s11103-004-5044-8
Wheatley, C., Lozano, C. and Gomez, G. (1985). "Post-harvest deterioration ofcassava roots," in Cassava: Research, Production and Utilization, eds J. H. Cock and J. A. Reyes. (Cali: UNDP-CIAT), 655-671.
Xu, J., Duan, X., Yang, J., Beeching, J. R., and Zhang, P. (2013). Enhanced reactive oxygen species scavenging by overproduction of superoxide dis- mutase and catalase delays postharvest physiological deterioration of cassava storage roots. Plant Physiol. 161, 1517-1528. doi: 10.1104/pp. 112.212803
Yadav, J. S., Ogwok, E., Wagaba, H., Patil, B. L., Bagewadi, B., Alicai, T., et al. (2011). RNAi-mediated resistance to cassava brown streak Uganda virus in transgenic cassava. Mol. Plant Pathol. 12, 677-687. doi: 10.1111/j.1364-3703.2010. 00700.x
Zainuddin, I., Schlegel, K., Gruissem, W., and Vanderschuren, H. (2012). Robust transformation procedure for the production of transgenic farmer- preferred cassava cultivars. Plant Methods 8, 24. doi: 10.1186/17464811-8-24
Zhang, P., Vanderschuren, H., Futterer, J., and Gruissem, W. (2005). Resistance to cassava mosaic disease in transgenic cassava expressing antisense RNAs targeting virus replication genes. Plant Biotechnol. J. 3, 385-399. doi: 10.1111/j.1467- 7652.2005.00132.x
Zhang, P., Wang, W. Q., Zhang, G. L., Kaminek, M., Dobrev, P., Xu, J., and Gruissem, W. (2010). Senescence-inducible expression of isopentenyl transferase extends leaf life, increases drought stress resistance and alters cytokinin metabolism in cassava. J. Integr. Plant Biol. 52, 653-669. doi: 10.1111/j.1744- 7909.2010.00956.x
Zidenga, T., Leyva-Guerrero, E., Moon, H., Siritunga, D., and Sayre, R. (2012). Extending cassava root shelf life via reduction of reactive oxygen species production. Plant Physiol. 154, 1396-1407. doi: 10.1104/pp.112. 200345