Abstract：Abstract Betaine aldehyde dehydrogenase (BADH) is the key enzyme in the synthesis process of glycine betaine (GB). To explore the expression difference of BADH gene in different tissues of cassava (Manihot esculenta) and the expression under drought and salt stress, bioinformatics approaches were used to identify and characterize MeBADH genes. Using computational methods, these genes were localized on cassava genome chromosome and stress-responsive cis-elements within their 5' upstream regions were identified. Expression profiles of these genes in different tissues were detected by qRT-PCR, at the same time, the differential expression of MeBADH genes under PEG-6000 stress, salt stress and natural drought stress were investigated in cassava. The GB content in cassava was determined by ELISA. The results showed that MeBADHs genes expressed in all tissues (root, root tuber, stem, leaf). The expression in root tuber was significantly higher than that in root (P＜0.05). The expression of MeBADH1 and MeBADH2 in leaves, stems, root and root tuber in 1~2 h changed a little compared with 0 h under PEG- 6000 treatment. Under NaCl treatment, the expression of MeBADH1 in leaf at 8 h was the highest, and was extremely significantly higher than that at 0 h (P＜0.01); the expression of MeBADH2 did not change much at different times. In stems and roots, the expressions of MeBADH1 and MeBADH2 at 1 h were significantly higher than that at 0 h (P＜0.01). Under natural drought treatment, the expressions of MeBADH1 and MeBADH2 in stem and root at 6 and 18 d were significantly higher than that at 0 d (P＜0.01), respectively. Under PEG-6000 treatment, the GB content was fluctuant and slightly increased. The GB content under NaCl treatment showed stable expression in the early stage, and then occurred intense response at 8 h (appeared downward trend), and the GB content at 24 h was significantly lower than that at 0 h (P＜0.05). Under natrual drought stress, the GB content gradually increased at beginning, reached he peak at 9 and 15 d, and was higher than that st 0 d (P＜0.05). This study provides foundation for further research on the molecular mechanisms of cassava resistance to drought stress and breeding drought-resistant new cultivars.
[1] 黄世平, 曾幼玲. 2015. 植物醛脱氢酶在逆境胁迫中的研究进展[J]. 生物技术通报, 2015, 31(12): 8-14.(Huang S P, Zeng Y L.2015. Research Progress on Plant Aldehyde Dehydrogenase Under Adversity Stresses. Biotechnology Bulletin[J], 31(12): 8-14. )[2] GANESH P K R S. 2016. Effect of coir pith compost on growth and yield attributes of onion and cassava[J]. International Journal of Applied Research,2: 501-506.[3] 黄洁, 李开绵, 叶剑秋,等. 2006. 中国木薯产业化的发展研究与对策[J]. 中国农学通报, 22(5): 421-426.(Huang J, Li K M, Ye J Q, et al. 2006. The Developing Countermeasure for Cassava Commercialization in China[J]. Chinese Agricultural Science Bulletin, 22(5): 421-426.)[4] Bakayoko S, Tschannen A, Nindjin C,et al. 2009. Impact of water stress on fresh tuber yield and dry matter content of cassava (Manihot esculenta Crantz) in C?te d'Ivoire[J]. African Journal of Agricultural Research, 4: 21-27.[5] Pérez J C, Lenis J I, Calle F, et al. 2011. Genetic variability of root peel thickness and its influence in extractable starch from cassava[J]. Plant Breeding, 130: 688-693.[6] Laban T F, Kizito E B, Baguma Y, et al. 2013. Evaluation of Ugandan cassava germplasm for drought tolerance[J]. International Journal of Agriculture and Crop Sciences, 5: 212-226.[7] Hanson A D, May A M, Grumet R, et al. 1985. Betaine synthesis in chenopods: Localization in chloroplasts[J]. Proceedings of the National Academy of Sciences, 82: 3678-3682.[8] Arakawa K, Takebe T, Sugiyama T, et al. 1987. Purification of betaine-aldehyde dehydrogenase from spinach leaves and preparation of its antibody[J]. Journal of biochemistry, 101: 1485-1488.[9] Rhodes D, Hanson A D.1993. Quaternary Ammonium and Tertiary Sulfonium Compounds in Higher Plants[J]. Annual Review of Plant Biology, 44: 357-384.[10] Grieve C M, Maas E V. 1984. Betaine accumulation in salt-stressed sorghum[J]. Physiologia Plantarum, 61: 167-171.[11] Kamata T, Uemura M. 2004. Solute accumulation in heat seedlings during cold acclimation: contribution to increased freezing tolerance[J]. Cryo Letters, 25: 311-322.[12] 惠红霞, 许兴, 李前荣. 2004. NaCl胁迫对枸杞叶片甜菜碱、叶绿素荧光及叶绿素含量的影响[J]. 干旱地区农业研究, 22(3): 109-114.(Hui H X, Xu X, Li Q R. 2004. Effects of N aCl stress on betaine, chlorophyll fluorescence and chloroplast pigment of leaves of Lycium Barbarum L[J]. Agricultural Research in the Arid Areas, 22(3): 109-114.) [13] Perozich J, Nicholas H, Wang B C. 1999. Relationships within the aldehyde dehydrogenase extended family[J]. Protein Science, 8: 137-146.[14] Chang S, Puryear J, Cairney J. 1993. A simple and efficient method for isolating RNA from pine trees[J]. Plant Molecular Biology Reporter, 11: 113-116.[15] Wang F W, Wang M L, Guo C ,et al. 2016. Cloning and characterization of a novel betaine aldehyde dehydrogenase gene from Suaeda corniculata[J]. Genetics & Molecular Research Gmr, 15: gmr.15027848.[16] 马建华, 郑海雷. 植物耐盐的分子生物学基础[J]. 2007.生物学杂志,24(1): 5-8.(Ma J H, Zheng H L. 2007. Molecular biological basis of salt tolerance in plants[J]. Journal of Biology, 24: 5-8.[17] 于同泉, 秦岭,王有年. 1996. 渗透胁迫板栗苗可溶性糖的积累及组分变化的研究[J]. 北京农学院学报, 11(1): 43-47.(Yu T Q, Qin L, Wang Y J.1996. The accumulation and fluctuation of soluble sugar compositions under water strees of chestnut seedlings[J]. Journal of Beijing Agricultura College, 11: 43-47.)[18] 杨成龙, 刘姣, 周扬,等. 2014. 海马齿甜菜碱醛脱氢酶基因在大肠杆菌中表达[J]. 分子植物育种, 12:1275-1280. (Yang C L, Liu J, Zhou Y, et al. 2014. Expression of Recombinant Betaine Aldehyde Dehydrogenase of Sesuvium portulacastrum in Escherichia coli[J].Molecular Plant Breeding, 12:1275-1280.)[19] Hsiao T C. 1973. Plant response to water stress[J]. Annu Rev Plant Physiol, 24:519-570.[20] Al Hakimi A, Monneveux P, Galiba G. 1996. Soluble sugars, proline and relative water content (RCW) as traits for improving drought tolerance and divergent selection for RCW from Triticum polonicum into Triticum durum[J]. Journal of Genetics & Breeding,49(13):237-244.[21] Diab A A, El-Sadi Y S, Ageez A, et al. 2012. Cloning and Expression Analysis of Betaine Aldehyde Dehydrogenase from Pseudomonas fluorescens[J]. Egyptjgenetcytol, 40:161-173.[22] Li P, Liu Z, Fu X , et al. 2006. cDNA cloning of FMDV structural protein VP2-3-1 gene and its prokaryotic expression[J]. Chin J Vet Sci, 26:232-234.[23] Weigel P, Weretilnyk E A, Hanson A D. 1986. Betaine Aldehyde Oxidation by Spinach Chloroplasts[J]. Plant Physiology, 82: 753-759.[24] Nakamura T, Nomura M, Mori H, et al. 2001. An isozyme of betaine aldehyde dehydrogenase in barley[J]. Plant & Cell Physiology, 42: 1088-1092.[25] 金杭霞, 董德坤, 杨清华,等. 2016. 碱蓬SgBADH的克隆与分析及植物表达载体构建[J]. 核农学报,30(2): 246-251.(Jin H X,Dong D K, Yang Q H, et al. 2016. Cloning and Analysis of SgBADH in Suaeda glauca and Construction of Its Plant Expression Vector[J]. Journal of Nuclear Agricultural Sciences, 30: 0246-0251.)[26] Hibino T, Meng Y L, Kawamitsu Y, et al. 2001. Molecular cloning and functional characterization of two kinds of betaine-aldehyde dehydrogenase in betaine-accumulating mangrove Avicennia marina (Forsk.) Vierh[J]. Plant Molecular Biology, 45: 353-363.[27] Mu?oz-Clares R A, Riveros-Rosas H, Garza-Ramos G, et al. 2014. Exploring the evolutionary route of the acquisition of betaine aldehyde dehydrogenase activity by plant ALDH10 enzymes: implications for the synthesis of the osmoprotectant glycine betaine[J]. BMC plant biology, 14: 149.[28] 刘振林, 曹华雯, 夏新莉, 等. 2009. 甘菊BADH基因cDNA的克隆及在盐胁迫下的表达[J]. 植物科学学报, 27(1): 1-7.( Liu Z L, Cao H W, Xia X L, et al. 2009. Cloning and Expression Analysis of Betaine Aldehyde Dehydrogenase Gene from Dendranthema lavandulifolium on Salinity[J]. Journal of Wuhan Botanical Research, 27: 1-7.)[29] Fan W J, Zhang M, Zhang H X, et al. 2012. Improved Tolerance to Various Abiotic Stresses in Transgenic Sweet Potato (Ipomoea batatas) Expressing Spinach Betaine Aldehyde Dehydrogenase[J]. Plos One, 7: e37344.[30] Missihoun T D, Schmitz J, Klug R, et al. 2011. Betaine aldehyde dehydrogenase genes from Arabidopsis with different sub-cellular localization affect stress responses[J]. Planta, 233: 369-382.[31] Metris A, George S M, Mulholland F, et al. 2014. Metabolic Shift of Escherichia coli under Salt Stress in the Presence of Glycine Betaine[J]. Applied & Environmental Microbiology, 80:4745-4756.[32] Paleg L G, Aspinall D. 1981.The Physiology and biochemistry of drought resistance in plants[M]. Sydney: Academic Press, pp:171-204.
