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中国生物工程杂志

China Biotechnology
China Biotechnology
China Biotechnology  2010, Vol. 30 Issue (08): 31-36    DOI: Q786
    
Expression Profiling of Genes Involved in Taxuyunnanine C Biosynthesis in Cell Suspension Cultures of Taxus chinensis by Repeated Elicitation with a Newly Synthesized Jasmonate,in situ Absorption and Sucrose Feeding
GAO Ming-bo1,2,ZHANG Wei3,LI Xing-tai2,RUAN Cheng-jiang1,2,FAN Sheng-di1
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Abstract  

Taxus suspension cell culture has the potential to provide a sustainable source of anticancer drug paclitaxel (Taxol) and other taxoids. In the cell culture of Taxus chinensis, taxuyunnanine c (Tc) is the primary taxoid. To design a rational strategy for redirecting the precursor fluxes from other taxoids into paclitaxel production, this study employed quantitative real-time-PCR (RQ-PCR) to understand the dynamic profiling of key biosynthetic pathway genes of palcitaxel and taxoids during the culture process. Six genes —TASY, TDAT, T5αH, TαH, TDAT and T10βH were quantified under a process condition of double elicitation by 100μM 2,3-dihydroxylpropanyl jasmonate(DHPJA) on day 7 and day 12, 20g/L sucrose feeding and 100g/L XAD-7HP absorption on day 7 . This process treatment led to a high accumulation of Tc at 1517±37mg/L on day 30, 11.1 folds of that of control and 1.7 folds of that of double elicitation with sucrose feeding. The RQ-PCR results showed that with DHPJA elicitation, the expression of all the six genes was greatly increased, but sharply declined after 12 hours. This rapid decline suggested that the second DHPJA elicitation was required to sustain gene expression. Simultaneous absorbents addition would delay the improvement of gene expression but maintain the gene expression at a higher level, resulting in higher gene expression over treatments without absorbents. TαH expression was somewhat different which was markedly inhibited by XAD-7HP absorption while absorbants still maintained its expression.

Key words： Taxus chinensis      Taxuyunnanine C      DHPJA      XAD-7HP      RQ-PCR     
Received: 29 April 2010      Published: 25 August 2010
Corresponding Authors: Gao Mingbo     E-mail: lily@dlnu.edu.cn
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GAO Meng-Bei
ZHANG Wei
LI Xin-Tai
RUAN Cheng-Jiang
FAN Ku-Di
Cite this article:

GAO Meng-Bei, ZHANG Wei, LI Xin-Tai, RUAN Cheng-Jiang, FAN Ku-Di. Expression Profiling of Genes Involved in Taxuyunnanine C Biosynthesis in Cell Suspension Cultures of Taxus chinensis by Repeated Elicitation with a Newly Synthesized Jasmonate,in situ Absorption and Sucrose Feeding. China Biotechnology, 2010, 30(08): 31-36.

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https://manu60.magtech.com.cn/biotech/Q786     OR     https://manu60.magtech.com.cn/biotech/Y2010/V30/I08/31

［1］ Baloglu E, Kingston D G I. The taxane diterpenoids. J Nat Prod, 1999, 62: 14481472. 
［2］ Itokawa H. Taxoids occuring in the genus Taxus. In: Itokawa H, Lee K H. TaxusThe Genus Taxus. London: Taylor & Francis, 2003.3578. 
［3］ Kikuchi Y,Yatagai M. The commercial cultivation of Taxus species and production of taxoids. In: Itokawa H, Lee K H. TaxusThe Genus Taxus. London: Taylor & Francis,2003.151178. 
［4］ Takeya K. Plant tissue culture of taxoids. In: Itokawa H, Lee K H. TaxusThe Genus Taxus.London: Taylor & Francis, 2003.134150. 
［5］ Zhao Z J，Xu Y F，Qian Z G, et al. Novel fluoro and hydroxylcontaining jasmonate derivatives as highly efficient elicitors in suspension cultures of Taxus chinensis. Bioorganic & Medicinal Chemistry Letters, 2004, 14:47554758. 
［6］ Qian Z G，Zhao Z J，Xu Y F, et al. Novel chemically synthesized hydroxylcontaining jasmonates as powerful inducing signals for plant secondary metabolism. Biotechnology and Bioengineering, 2004, 86( 7): 809816. 
［7］ Qian Z G, Zhao Z J, Xu Y F, et al. Highly efficient strategy for enhancing taxoid production by repeated elicitation with a newly synthesized jasmonate in fedbatch cultivation of Taxus chinensis cells. Biotechnol Bioeng, 2005, 90(6): 516521. 
［8］ Croteau R. Cyclization of geranylgeranyl diphosphate to taxa4(5),11(12)diene is the committed step of Taxol biosynthesis in Pacific yew. J Biol Chem, 1995, 270:86868690. 
［9］ Hezari M, Croteau R. Taxol biosynthesis: an update. Planta Med, 1997, 63: 291295. 
［10］ Hefner J, Ketchum R E, Croteau R. Cloning and functional expression of a cDNA encoding geranylgeranyl diphosphate synthase from Taxus canadensis and assessment of the role of this prenyltransferase in cells induced for taxol production . Arch Biochem Biophys, 1998, 360(1) :6274. 
［11］ Ketchum R E B, Christopher D R, Qiu D Y, et al. Taxus metabolomics: methyl jasmonate preferentially induces production of taxoids oxygenated at C13 in Taxus x media cell cultures. Phytochem, 2003, 62: 901909. 
［12］ Croteau R, Ketchum R E B, Long R M, et al. Taxol biosynthesis and molecular genetics. Phytochem Rev, 2006, 5: 7597. 
［13］ Jennewein S, Long R M, Williams R M, et al. Cytochrome P450 taxadiene 5ahydroxylase, a mechanistically unusual monooxygenase catalyzing the first oxygenation ste Pof Taxol biosynthesis. Chem Biol, 2004, 11: 379387. 
［14］ Jennewein S, Rithner C D, Williams R M, et al. Taxoid metabolism: taxoid 14bhydroxylase is a cytochrome P450dependent monooxygenase. Arch Biochem Biophys, 2003, 413: 262270. 
［15］ Croteau R. Administering cultured Taxus cells with early precursors reveals bifurcations in the taxoid biosynthetic pathway. Phytochem, 2007, 68:335341. 
［16］ 高明波，张卫，虞星炬. 原位吸附和茉莉酸甲酯联合使用显著提高中国红豆杉细胞云南紫杉烷c的生产，生物工程学报，2010，26（2）：223229. Gao M B, Zhang W, Yu X J.Chin J Biotech, 2010, 26(2): 223229. 
［17］ 徐德昌，纪巍，程大友,等. TaqMan探针荧光定量PCR检测甜菜低温胁迫相关miRNAs，中国甜菜糖业，2008，4：36. Xu D C, Ji W, Cheng D Y, et al.China Beet & Sugar, 2008, 4: 36. 
［18］ Ezekiel N, Camille P D, Susan C R, et al. Expression profiling of genes involved in paclitaxel biosynthesis for targeted metabolic engineering. Metab Eng, 2006, 8:385–394.
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