苜蓿高产蛋白悬浮细胞系的筛选及其反应动力学研究
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摘要
为了系统全面的研究苜蓿悬浮细胞培养中代谢产物蛋白质的反应动力学,分别从苜蓿品种、外植体、培养条件、愈伤组织和悬浮细胞系等方面进行了优选,对苜蓿细胞生长过程中营养物质的消耗动力学和生长结构动力学模型进行了探讨,为大规模细胞培养生产苜蓿蛋白提供了基础理论研究依据。
根据亲缘关系较远的五种苜蓿中叶组织的蛋白酶活和蛋白质含量的测定,选择苜蓿王(WL215)作为苜蓿的供试品种。进行了苜蓿叶愈伤组织的诱导以及高产蛋白苜蓿悬浮细胞系的筛选。通过单因子试验,分别确定在600lx,12h循环光照条件下,采用0.3cmx0.3cm大小,苜蓿叶脉处外植体在MB+2.0mg/L 2,4-D+0.5mg/L 6-BA的优化培养基中可诱导出多量且较疏松的愈伤组织。筛选获得的苜蓿高产蛋白悬浮细胞系培养12d后生长指数及蛋白总酶活均达到最大值,分别为6.94和10.14x104U/L。
通过单因素变化考察了培养基中主要营养成分对悬浮培养苜蓿细胞生长和蛋白积累的影响。蔗糖是适合苜蓿细胞悬浮培养的碳源,不仅有利于细胞生长,并有较高的蛋白产量。在1%-6%范围内,苜蓿细胞生物量和蛋白积累量在3%蔗糖浓度下达到最高。MB培养基的基本氮源已经足够满足苜蓿细胞生长和蛋白合成的需要。NH_4~+和NO_3~-作为无机氮源提供的离子较易被苜蓿细胞吸收利用,尿素和酪蛋白水解物有助于苜蓿细胞的生长。磷酸盐对苜蓿细胞生长和蛋白合成都是必需的。细胞在无磷培养基中不生长,不同起始PO43-浓度对蛋白合成影响较大。1.25mM磷源浓度下苜蓿细胞生长和蛋白的合成最佳。
苜蓿细胞生长过程呈现较典型的“S”型曲线。低浓度时蔗糖成为细胞生长的限制性基质;提高蔗糖浓度可明显促进细胞生长,但浓度超过3%后则表现出对生长的底物抑制作用。不同NH_4~+/NO_3~-比例对苜蓿细胞生长的影响主要体现在高NH_4~+浓度对细胞生长的抑制作用。低浓度时磷酸盐成为苜蓿细胞生长的限制性因素。提高磷酸盐浓度细胞生长周期缩短。在基准培养条件下,苜蓿细胞的比生长速率约为0.31~0.34day-1,倍增时间约为3.4~3.8天。
不同起始蔗糖浓度下蛋白积累的趋势相似,在一定范围内,增加起始蔗糖浓度有利于蛋白的合成,但在高糖浓度下,蛋白总酶活下降。采用单一NH_4~+为氮源培养时蛋白总酶活最低,其他氮源配比下蛋白的总酶活均可达到一定的水平,当NH_4~+/NO_3~-为20:40时,蛋白总活力达到最大。初始磷酸盐的浓度对苜蓿细胞蛋白积累影响显著,1倍磷源浓度时蛋白积累可达到最大值。
不同培养条件下,蛋白的积累与苜蓿细胞的生长呈相对密切的偶联关系,蛋白的积累与苜蓿细胞生长呈正相关。
苜蓿细胞对糖的吸收速率和利用效率以及胞内糖的积累与培养基的组成有很大关系。通过线性回归计算了各种培养条件下苜蓿细胞及蛋白对糖的得率。
苜蓿细胞对NH_4~+和NO_3~-离子的吸收差异较大,NH_4~+离子在培养初期即被快速吸收,而NO_3~-离子的吸收较缓慢,细胞内NH_4~+和NO_3~-离子的积累不明显。
苜蓿细胞对磷酸盐吸收快速,起始磷源浓度越高,被细胞利用的速率越快。胞内磷酸盐的积累水平与培养基初始磷酸盐浓度有关。苜蓿细胞蛋白含量的积累与胞内磷酸盐的积累水平成反比。胞内磷酸盐的积累水平对苜蓿细胞的生长和蛋白的合成有重要的影响。
构建了苜蓿细胞悬浮培养的结构动力学模型,整个培养系统分为生物相和非物相两部分。生物相是苜蓿细胞内部,被分为四个部分:可溶性糖G、游离磷盐P、中间代谢产物M、代谢产物蛋白S;非生物相指苜蓿细胞外培养基,包括蔗糖Se、还原糖Ge和磷酸盐Pe三个部分。模型的模拟结果与测定值基本吻合,能应用于苜蓿细胞培养过程。
Induction Alfalfa suspension cell cultures in the kinetics of protein metabolism suspension cell line were studied. That are optimized from Alfalfa varieties, explants, culture conditions, callus and suspension cell lines, etc. The process of cell growth on the nutrition of Alfalfa material consumption and growth kinetics were discussed structural dynamics model for large-scale production of alfalfa protein in cell culture provide a basis for fundamental research.
According to the paternity far five kinds of Alfalfa are determinate the protein protease alive and protein content that choice the Alfalfa (WL215). Callus of the Alfalfa induction and high production protein of Alfalfa lines suspended selection. Selected the optimized of suspension cell culture the explants size with epidemus was 0.3cm X 0.3cm, more loose callus could be induced from the WL215 in the medium of MB+ 3.0 mg/L 2,4-D+0.5mg/L 6-BA under condition of 600lux 12hours circular illumination. The maximum growth indices was 6.49 while the maximum total Protein activity was 10.14x 104U/L selected Alfalfa cell line suspension after 12 days' culture.
The effects of major nutrients on Alfalfa cell growth and protein production were discussed. Sucrose was better for cell growth and protein production among different carbon sources. In the range of 1%~6% sucrose concentration,highest biomass and protein production were achieved on 3%. Standard nitrogen concentration in MB medium was enough for Alfalfa cell growth and protein production. Mineral nitrate source and urea were suitable for cell growth. Phosphate was necessary for cell growth and protein production. Different initial phosphate concentration had remarkable effect on protein production. The standard phosphate concentration appeared in 1.25mM more suitable for cell growth and protein production.
The suspension culture process of Alfalfa cell could be appeared“S”grow curve. Sucrose was growth limiting nutrient on low concentration condition. Increase of sucrose concentration stimulated cell growth, extended rapid growth phase and increased the final cell harvest. Substrate inhibition was observed when the concentration was above 3%. The influence of different NH_4~+ / N03- ratio on cell growth was mainly reflected by the inhibition of high ammonium. Phosphate was also the growth limiting nutrient on low concentration condition. Increase of phosphate concentration increased cell growth rate and shortened the culture period. Under the standard culture condition, the specific cell growth rate was 0.31~0.34 day-1, and the doubling time was 3.4~3.8 days.
The trends of protein production variation under different initial sucrose concentration were the same. Increase of sucrose concentration could also improve protein production to some extent, while higher sucrose concentration inhibited protein production. Protein production was the lowest when used ammonium as only nitrate source. The maximum total protein activity could be achieved while the ratio of NH_4~+ / N03- was 20/40. Initial phosphate concentration was most effective on protein accumulation; the maximum protein yield could be obtained under the standard phosphate concentration in 1.25mM.
Protein accumulation was partially-growth-associated, protein production by Alfalfa cell suspension was regard as synchronism with cell growth. Alfalfa cell rapidly hydrolyzed extra cellular sucrose before adsorb it. The hydrolyzing process could be described using substrate saturation kinetic equation. Glucose was absorbed more preferentially than fructose. The adsorbing rate, utilizing efficiency and intracellular accumulation of sugar were closely related to medium composition.
Ammonium was rapidly adsorbed at the early stage. The adsorption of nitrate was much more slowly. The intracellular accumulation of ammonium and nitrate was not obvious.
Phosphate was also rapidly adsorbed at the early stage. The higher the initial phosphate concentration was, the more rapid the utilization rate was. The intracellular phosphate accumulation level related to the initial phosphate concentration in the medium. Protein content in Alfalfa cell was decreased with the increasing of intracellular phosphate accumulation. The accumulation level showed strong influence on Alfalfa cell growth and protein production.
A structural kinetic model was constructed in this thesis. The whole culture system was separated into biotic phase and abiotic phase. The former was the inside of Alfalfa cell, which was divided into 4 compartments: soluble sugar G, free phosphate P, Middle metabolites M and products S. The abiotic phase was extra cellular environment, which included sucrose Se, reductive sugar Ge, and phosphate Pe. Intracellular phosphate acted as a regulator on the cell growth and protein accumulation. Calculated result coincided well with experimental data. The model could be used for good description of culture process.
根据亲缘关系较远的五种苜蓿中叶组织的蛋白酶活和蛋白质含量的测定,选择苜蓿王(WL215)作为苜蓿的供试品种。进行了苜蓿叶愈伤组织的诱导以及高产蛋白苜蓿悬浮细胞系的筛选。通过单因子试验,分别确定在600lx,12h循环光照条件下,采用0.3cmx0.3cm大小,苜蓿叶脉处外植体在MB+2.0mg/L 2,4-D+0.5mg/L 6-BA的优化培养基中可诱导出多量且较疏松的愈伤组织。筛选获得的苜蓿高产蛋白悬浮细胞系培养12d后生长指数及蛋白总酶活均达到最大值,分别为6.94和10.14x104U/L。
通过单因素变化考察了培养基中主要营养成分对悬浮培养苜蓿细胞生长和蛋白积累的影响。蔗糖是适合苜蓿细胞悬浮培养的碳源,不仅有利于细胞生长,并有较高的蛋白产量。在1%-6%范围内,苜蓿细胞生物量和蛋白积累量在3%蔗糖浓度下达到最高。MB培养基的基本氮源已经足够满足苜蓿细胞生长和蛋白合成的需要。NH_4~+和NO_3~-作为无机氮源提供的离子较易被苜蓿细胞吸收利用,尿素和酪蛋白水解物有助于苜蓿细胞的生长。磷酸盐对苜蓿细胞生长和蛋白合成都是必需的。细胞在无磷培养基中不生长,不同起始PO43-浓度对蛋白合成影响较大。1.25mM磷源浓度下苜蓿细胞生长和蛋白的合成最佳。
苜蓿细胞生长过程呈现较典型的“S”型曲线。低浓度时蔗糖成为细胞生长的限制性基质;提高蔗糖浓度可明显促进细胞生长,但浓度超过3%后则表现出对生长的底物抑制作用。不同NH_4~+/NO_3~-比例对苜蓿细胞生长的影响主要体现在高NH_4~+浓度对细胞生长的抑制作用。低浓度时磷酸盐成为苜蓿细胞生长的限制性因素。提高磷酸盐浓度细胞生长周期缩短。在基准培养条件下,苜蓿细胞的比生长速率约为0.31~0.34day-1,倍增时间约为3.4~3.8天。
不同起始蔗糖浓度下蛋白积累的趋势相似,在一定范围内,增加起始蔗糖浓度有利于蛋白的合成,但在高糖浓度下,蛋白总酶活下降。采用单一NH_4~+为氮源培养时蛋白总酶活最低,其他氮源配比下蛋白的总酶活均可达到一定的水平,当NH_4~+/NO_3~-为20:40时,蛋白总活力达到最大。初始磷酸盐的浓度对苜蓿细胞蛋白积累影响显著,1倍磷源浓度时蛋白积累可达到最大值。
不同培养条件下,蛋白的积累与苜蓿细胞的生长呈相对密切的偶联关系,蛋白的积累与苜蓿细胞生长呈正相关。
苜蓿细胞对糖的吸收速率和利用效率以及胞内糖的积累与培养基的组成有很大关系。通过线性回归计算了各种培养条件下苜蓿细胞及蛋白对糖的得率。
苜蓿细胞对NH_4~+和NO_3~-离子的吸收差异较大,NH_4~+离子在培养初期即被快速吸收,而NO_3~-离子的吸收较缓慢,细胞内NH_4~+和NO_3~-离子的积累不明显。
苜蓿细胞对磷酸盐吸收快速,起始磷源浓度越高,被细胞利用的速率越快。胞内磷酸盐的积累水平与培养基初始磷酸盐浓度有关。苜蓿细胞蛋白含量的积累与胞内磷酸盐的积累水平成反比。胞内磷酸盐的积累水平对苜蓿细胞的生长和蛋白的合成有重要的影响。
构建了苜蓿细胞悬浮培养的结构动力学模型,整个培养系统分为生物相和非物相两部分。生物相是苜蓿细胞内部,被分为四个部分:可溶性糖G、游离磷盐P、中间代谢产物M、代谢产物蛋白S;非生物相指苜蓿细胞外培养基,包括蔗糖Se、还原糖Ge和磷酸盐Pe三个部分。模型的模拟结果与测定值基本吻合,能应用于苜蓿细胞培养过程。
Induction Alfalfa suspension cell cultures in the kinetics of protein metabolism suspension cell line were studied. That are optimized from Alfalfa varieties, explants, culture conditions, callus and suspension cell lines, etc. The process of cell growth on the nutrition of Alfalfa material consumption and growth kinetics were discussed structural dynamics model for large-scale production of alfalfa protein in cell culture provide a basis for fundamental research.
According to the paternity far five kinds of Alfalfa are determinate the protein protease alive and protein content that choice the Alfalfa (WL215). Callus of the Alfalfa induction and high production protein of Alfalfa lines suspended selection. Selected the optimized of suspension cell culture the explants size with epidemus was 0.3cm X 0.3cm, more loose callus could be induced from the WL215 in the medium of MB+ 3.0 mg/L 2,4-D+0.5mg/L 6-BA under condition of 600lux 12hours circular illumination. The maximum growth indices was 6.49 while the maximum total Protein activity was 10.14x 104U/L selected Alfalfa cell line suspension after 12 days' culture.
The effects of major nutrients on Alfalfa cell growth and protein production were discussed. Sucrose was better for cell growth and protein production among different carbon sources. In the range of 1%~6% sucrose concentration,highest biomass and protein production were achieved on 3%. Standard nitrogen concentration in MB medium was enough for Alfalfa cell growth and protein production. Mineral nitrate source and urea were suitable for cell growth. Phosphate was necessary for cell growth and protein production. Different initial phosphate concentration had remarkable effect on protein production. The standard phosphate concentration appeared in 1.25mM more suitable for cell growth and protein production.
The suspension culture process of Alfalfa cell could be appeared“S”grow curve. Sucrose was growth limiting nutrient on low concentration condition. Increase of sucrose concentration stimulated cell growth, extended rapid growth phase and increased the final cell harvest. Substrate inhibition was observed when the concentration was above 3%. The influence of different NH_4~+ / N03- ratio on cell growth was mainly reflected by the inhibition of high ammonium. Phosphate was also the growth limiting nutrient on low concentration condition. Increase of phosphate concentration increased cell growth rate and shortened the culture period. Under the standard culture condition, the specific cell growth rate was 0.31~0.34 day-1, and the doubling time was 3.4~3.8 days.
The trends of protein production variation under different initial sucrose concentration were the same. Increase of sucrose concentration could also improve protein production to some extent, while higher sucrose concentration inhibited protein production. Protein production was the lowest when used ammonium as only nitrate source. The maximum total protein activity could be achieved while the ratio of NH_4~+ / N03- was 20/40. Initial phosphate concentration was most effective on protein accumulation; the maximum protein yield could be obtained under the standard phosphate concentration in 1.25mM.
Protein accumulation was partially-growth-associated, protein production by Alfalfa cell suspension was regard as synchronism with cell growth. Alfalfa cell rapidly hydrolyzed extra cellular sucrose before adsorb it. The hydrolyzing process could be described using substrate saturation kinetic equation. Glucose was absorbed more preferentially than fructose. The adsorbing rate, utilizing efficiency and intracellular accumulation of sugar were closely related to medium composition.
Ammonium was rapidly adsorbed at the early stage. The adsorption of nitrate was much more slowly. The intracellular accumulation of ammonium and nitrate was not obvious.
Phosphate was also rapidly adsorbed at the early stage. The higher the initial phosphate concentration was, the more rapid the utilization rate was. The intracellular phosphate accumulation level related to the initial phosphate concentration in the medium. Protein content in Alfalfa cell was decreased with the increasing of intracellular phosphate accumulation. The accumulation level showed strong influence on Alfalfa cell growth and protein production.
A structural kinetic model was constructed in this thesis. The whole culture system was separated into biotic phase and abiotic phase. The former was the inside of Alfalfa cell, which was divided into 4 compartments: soluble sugar G, free phosphate P, Middle metabolites M and products S. The abiotic phase was extra cellular environment, which included sucrose Se, reductive sugar Ge, and phosphate Pe. Intracellular phosphate acted as a regulator on the cell growth and protein accumulation. Calculated result coincided well with experimental data. The model could be used for good description of culture process.
引文
[1]欧阳平凯,陆永明,肖民耕等,生物工程进展,12(5):12,1992
[2]周立刚,郑光植,生物工程进展,11(1):29一35,1991
[3]郭勇,崔堂兵等主编.植物细胞培养技术与应用[ M] .北京:化学工业出版社, 2003. 12.
[4]韩军丽,李振秋,刘本叶等植物萜类代谢工程,生物工程学报2007,24(4): 561~569
[5] Powler M. W. Plnat cell culture. VolⅡ,edited by Russel G.E.,1984:41~67
[6]孙敬三,桂耀林主编.植物细胞工程试验技术.科学出版社,北京,1995, 49~72
[7]朱蔚华.人参的研究及报告.淑馨出版社,1989:121~168
[8]王文静,袁道强,高松洁植物组织培养的应用现状[ M] .河南师范大学学报, 2000, ( 3) : 137~139.
[9]孙志强,孙占育,植物组织培养技术在生产上的应用与进展,陕西农业科学,2010(3),98~103
[10]黄远新,玉永雄,胡艳,南方紫花苜蓿不同外植体离体培养的研究,中国草地,2003,25(3):43~47
[11]熊丽,吴丽芳.观赏花卉的组织培养与大规模生产[ M] .北京:北京化学工业出版社, 2003, 80~81.
[12]曾黎琼,段玉云,张云孙等,云南转基因农作物研究与产业化:现状及其发展,云南大学学报(自然科学版),2009,31(S2):479~488
[13]邝琦,余倩花,周厚高,植物细胞工程在菊花育种中的应用,北方园艺,2010(04):220~224
[14] JAMES C. Global status of commercialized Biotech /GMcrop s: 2006 [R ]. International Service for The Acquisi2tion ofAgri - Biotech App lications ( ISAAA). 2006.
[15] Goossens A ,Hakkinen ST,Laakso I , et al . A functional genomics approach toward the understanding of secondary metabolism in plant cells. Proc Natl Acad Sci USA ,2003 ,100 : 8595 - 8600.
[16] Winkel BSJ . Metabolic channeling in plants. Annu Rev Plant Biol ,2004 ,55 : 85 - 107.
[17] Kappers IF , Aharoni A , van Herpen TWJM, et al . Genetic engineering of terpenoid metabolism attracts bodyguards to Arabidopsis . Science ,2005 ,309 : 2070 - 2072.
[18]王兴春,李宏,王敏等。植物体细胞胚胎发生的调控网络,生物工程学报,2010,25;26(2):141~146
[19] Lange BM, Ketchum REB , Croteau RB. Isoprenoid biosynthesis. Metabolite profiling ofpeppermint oil gland secretory cells and application to herbicide target analysis. Plant Physiol ,2001 ,127 :305 - 314.
[20] Sandmann G, Romer S , Fraser PD. Understanding carotenoid metabolism as a necessity for genetic engineering of crop plants.Metabolic Engineering ,2006 ,8 : 291 - 302.
[21]于荣敏,金钱星,孙辉等。西洋参冠瘿组织悬浮培养及其人参皂苷类成分的分离,生物工程学报,2005,21(5):754~758
[22]钟建江,沈林南,王斯靖.植物细胞培养工程的最新进展.工业微生物,1995,25(l):25~29
[23]郑光植.植物细胞培养及其次级代谢.第1卷.云南大学出版社,昆明,1992:1~2
[24] Fujita Y, Hara Y, Suga C et al.Production of Shikonin derivatives by cell suspension cultures of Lithospermum erythrozhizon of shikonin derivatives.Plnat Cell Report, 1981,l:61~63
[25] Zenk M H. Plant tissue culture and its biotechnological application, W. Barz et al(Ed.), Springer-Verlag, New York, 1977:27~43
[26]罗建平,牛炳韬,贾敬芬等.云南红豆杉培养细胞系的建立.生物工程学报,1997,13(3):326~330
[27]沈征武,吴莲芬.紫杉醇研究进展.化学进展,1997,9(1):1~13
[28] Fowler M. W. Plant Biotechnology, Mantell(Ed.), Cambridge Univ. Press, 1983:3~7
[29]陈因良,陈志宏.细胞培养工程,华东化工学院出版社,上海,1992:275~276
[30] Drapeau D, Blanch H W, Wilke C R. Economic assessment of plant cell culture for the production of Agrnalicine. Biotechnol. Bioeng., 1987,30:946~953
[31] Tanaka H. Large-Scale cultuvition of plant cells at high density: A review. Process Biochemistry, 1987,8:106~113
[32] Dornenburg H, Knorr D. Strategies for the improvement of secondary metabolite production in plant cell cultures. Enzyme Microb.Technol.,1995,17:674~684
[33] Yoshikawa T., T.Furuya. Plant Cell Reports,1987,6:449~453
[34] Parr A J. Secondary products from plant cell cultures. Adv.Bio/technol Proc.,1988,9:l~34
[35] Brodelius P. The potential role of immobilization in plant cell biotechnolngy.Trends in Biotechnol.,1985,3:280~285
[36] Lindsey K., Yeoman M. M.The synthetic potential of immobilized cells of capsicum frutescens mill cv. Annuurm. Planta,1984,162:495~501
[37]韩迎山.植物细胞培养生产天然产物的研究进展。生物化学与生物物理学进展。1989,16(1):19~22
[38] Tsoulpha P, Doram P M. Solasodine production from self-immobilizated Solanum aviculare cells. J.Biotechnol.1991,19:99~110
[39] Hoedstra S S, Harkes P A A., Verpoorte R. et al. Effect of aucin on cytodifferentitation andproduction of quinoline alkaloids in compact globulas structures of Cinchona ledgeriana. Plant Cell Report, 1990,8:571~574
[40]王爱勤,何龙飞,林鉴钊。草本咖啡的组织培养和植株再生,广西农业生物科学,1999,18(2):6
[41]许建峰.高山红景天愈伤组织颗粒悬浮培养生产红景天甙动力学与过程控制.大连理工大学博士论文.1996
[42] Francois G, Christen P, Kapetanidis I. Prodution of valepotriates by hairy root cultures of Centranthus ruber DC. Plant Cell Report, 1995, 14:294~298
[43] Sang Y B, Henrik P. Two-phase airlift fermentor operation with elicitation for the enhanced production of Benzophenanthridine alkaloids in cell suspension of Escherichia Californica. Biotechnol. Bioeng., 1994,44:14~20
[44] Kim D J, Chang H N. Enhanced skikonin production from Lithospermam erythroihizon by in Situ extraction and Calcium alginate immobilization. Biotech. Bioeng., 1990,36:460~466
[45] Pyane G F, Pyane N N, Shuler M L et al. In Situ adsorption for enhanced alkaloid production by Catharanthus Roseus. Biotech. Letter, 1998,88:187~192
[46] Byun S Y, Pederson H, Chin C K. Two-phase culture for the enhanced production of benzophenanthridine alkaloids in cell suspensions of Eschancholtzia Californica. Phytochemistry, 1990,29(10):3135~3139
[47] Buitelaar R M, Langenhoff A A M, Heidstra R et al. Growth and thiophene production by hairy root cultures of Tagates patrla in various two-liquid-phase bioreactors. Enzyme Microb. Technol., 1991,13:487~494
[48]陈士云,候篙生.生产植物次生代谢物几项新技术研究进展.生物工程进展,1993,14(2):43~45
[49] Dicosmo F, Misawa M. Eliciting secondary metabolism in plant cell cultures. Trends in Biotechnol., 1985, 3:318~322
[50] Panda A K, Mishra, Saroj et al. Plant cell reactor- a perspective. Enzyme Micro. Technol., 1989,11:386~387
[51]陶璐璐,袁明静,徐锦堂.丹参愈伤组织细胞固定化及其转化产物的特征.生物工程学报,1990,6(3):218~223
[52]徐任生,天然产物化学,科学出版社,北京,1993,p856一893
[53] Fujita Y, Hara Y, Ogino T et al, Plant Cell Report, 1981,1:59~62
[54] Schlatmann. J. E, Mreno.P. R, Selles. M et al, Biotechnol. Bioeng., 1995,47:53~59
[55]甘烦远,郑光植,植物学通讯,1991,8(4):14~20
[56] Panda A K, Mishra, Saroj et al, Plant cell reactor - a perspective. Enzyme Micro.Technol.,1989,11:386~387
[57] Hashimoto. T, Pytochemistry, 1993,32:713~718
[58] Fesker. L. F, Plant Mol. Biol. 1993,23:11~21
[59] Phillipson J D. et al. The Chemistry and Biology of Isoquinoline, Alkaloids, springer-verlag, Berlin,1985,265
[60]刘涤.植物次生物质工业化生产研究的现况.生物工程学报,1987,3(1):9~15
[61]齐鸿雁编译,Jean Paul Schwitzguebal等著.从植物细胞到生物工程.生物工程进展,1992,12(5):24~28
[62]山根恒夫著,周斌译.生化反应工程,西北大学出版社,1992
[63]安东·莫泽尔著,曲音波等译.生物工艺技术学,湖南科学技术出版社,1994
[64] Shuler M L.Comprehensive biotechnology-The principles, application and regulations of biotechnology in industry, agriculture and medicine, Vol.2, Murray Moo-Young et al(Ed.), Cambridge Univ. Press, 1985,119~131
[65]戚义政,汪叔雄.生化反应动力学与反应器.化学工业出版社,1996,119
[66] L.拉皮德斯.化学反应器理论,石油工业出版社,1984,271~323.
[67]赵霞,叶勤,俞俊棠.影响重组工程菌发酵产率的因素.工业微生物,1995,25(2):26~32.
[68]郑穗平,郭勇.基因工程菌培养过程的动力学模型.微生物学通报,1998,25(l):46~48.
[69]俞俊棠、顾其丰、叶勤.生物化学工程,化学工业出版社,1991:68~73.
[70]白净.生理系统的仿真与建模,清华大学出版社,1994:211~240.
[71]杨冰.实用最优化方法及计算机程序,哈尔滨船舶工程学院出版社,1994.
[72] Brian S H, James M. L. Application of a new structured model to Tobacco cell cultures. Biotechnol. Bioeng.,2003,39:765~774
[73] Van Gulik W M, Ten Hoopen H J G, Heijnen J J. A structured model describing carbon and phosphate limited growth of catharanthus roses plant cell suspensions in batch and chemostat culture, Biotechnol. Bioeng,2005,41:771~780
[74] Bailey C M, Nicholsm H. A new structured model for plant cell culture. Biotechnol. Bioeng.,2003,34:1331~1336.
[75] Bailey C M, Nicholsm H. Optimal temperature control for a structured model of plant cell culture. Biotechnol. Bioeng.,2000,35:252~259
[76] Wei Zhang, Seki M., Furusaki S et al. A phase-segergated model for plant cell culture:the effect of cell volume fraction. Journal of Chemical Engineering of Japan, 2002:31(3):469~474
[77] Aynsley M, Ward A C., Wright A R. A mathematical model for the growth of mycelial Fungi in submerged culture.Biotechnol. Bioeng.,2005,35:820~830
[78] De Gunst M C M, Harkes P A A, van Zwet, Modelling the growth of a batch culture of plantcells; a corpuscular approach. Enzyme Micro. Technol., 2000,12:61~71
[79]颜日明,张志斌,邱晓芳等,杜仲细胞悬浮培养产黄酮及其动力学研究,中国生物工程杂志,2008,28(10):60~65
[80] Dohnal M. Fuzzy bioengineering models. Biotechnol. Bioeng., 2005,27:1146~1151
[81]王西娥,吴玉荷,烟草细胞悬浮培养生产CoQ10的动力学研究,仲恺农业技术学院学报,200619(4):9~12
[82]高正航,李卫东,长春花生物碱类药物研究概况,贵州农业科学,2005,33(6):94~96
[83] Schlatmann J E ,Moreno P R H , Ten Hoopen H J G, et al .Effect of oxygen and nut rient limitation on ajamalicine production and related enzyme actives in high density cutures of Catharant hus roseus[J ] . Biotechnol Bioeng ,1994 ,44 (4) :461~468.
[84] Drapeau D ,Blanch H W ,Wilke C R. Ajmalicine , serpentine and catharant hine accumulation in Cat harant hus roseus bioreactor cultures [J ] . Planta Med ,1987 (53) :373~376.
[85] Wei Zhang, Seki M., Furusaki S et al. A phase-segregated model for plant cell culture: the effect of cell volume fraction. Journal of Chemical Engineering of Japan, 1998:31(3):469~474
[86]艾江宁,贾景明,高山红景天细胞悬浮培养生长和产物积累动力学,沈阳药科大学学报,2009 8,26(8):653~656
[87] Aynsley M, Ward A C,. Wright A R. A mathematical model for the growth of mycelial Fungi in submerged culture. Biotechnol. Bioeng.,2008,35:820~830
[88]许建峰.高山红景天愈伤组织颗粒悬浮培养生产红景天甙动力学与过程控制。大连理工大学博士论文. 1996
[89] Gunst M C M, Harkes P A A, van Zwet. Modelling the growth of a batch culture of plant cells; a corpuscular approach. Enzyme Micro. Technol., 1999,12:61~71
[90] Dohnal M. Fuzzy bioengineering model. Biotechnol. Bioeng., 2005,27:1146~1151
[91] Baraniak B, Karas M.Antioxidative properties of chloroplast concentratesobtained by various methods from Lucerne juice [J].Journal of Animal andFeed Science, 2000, 9( 2) : 397～405.
[92]曹致中,席亚丽,赵波.苜蓿叶蛋白提取及深加工[A].首届中国苜蓿发展大会(论文集) [C].北京:中国草原学会, 2001.100～115.
[93]董宽虎,朱慧森.苜蓿产品的开发应用现状[A].第二届中国苜蓿发展大会(论文集) [C].北京:中国草原学会, 2003.210～213.
[94]郭佩玉.苜蓿叶蛋白工业化提取概述[A].第二届中国苜蓿发展大会(论文集) [C].北京:中国草原学会, 2003.129～130.
[95]强莉.苜蓿叶蛋白的加工及开发利用[J].饲料博览, 2002, 4: 43～51.
[96]薛琳,田丽萍,刘青广.苜蓿叶蛋白提取后副产品的开发与利用[J].江西饲料, 2006, 2: 18～19.
[97]杨丽娥,胡雪华.叶蛋白提取方法研究[J].上海交通大学学报(农业科学版) , 2003, 21( 12) : 66～70.
[98]李斌.剑麻组织和细胞培养诱导蛋白酶的研究.华南理工大学博士学位论文,1998
[99]马晖玲,卢欣石,曹致中.苜蓿原生质培养及体细胞杂交技术的应用[J ] .草业科学, 2007 ,24 (8) :64~68
[100] Johnson L B , Stuteville D L , Higgns R K, et al . Regeneration of alfalfa plant s from protoplast s of regen′s clone [ J ] . Plant Science Letters , 1981 , 20 : 297~304
[101] Lu D Y, Davey M R , Cocking E C. A comparison of t he cultural behaviour of protoplasts of leaves , cotyledonous and root of Medicago sati va [J ] . Plant Science Letters , 1983 , 31 : 87~89
[102] Gilmour D E , Davey M R , Cooling E C. Plant regeneration f rom cotyledon of wild Medicago species [ J ] . Plant Science ,1987 , 48 : 107~112
[103]王娟,李玉珠,师尚礼。苜蓿愈伤组织原生质体游离于培养[J].草地学报,2010,3(18):258~262
[104]赵红娟,张博,陈爱坪,等.酶解对苜蓿子叶原生质体分离效果的影响[J ] .草地学报, 2008 , 16 (1) : 24~29
[105]白静仁,何茂泰,袁清,等.野生黄花苜蓿叶肉原生质体培养和植株再生[J ] .草地学报, 1994 , 2 (1) : 59~63
[106]舒文华,耿华珠,孙勇如.紫花苜蓿原生质体培养与植株再生[J ] .草地学报, 1994 , 2 (1) : 40~44
[107]黄绍兴,王慧叶,占德扬.紫花苜蓿根原生质体植株再生[J ] .科技通报, 4 (11) : 35~39
[108]张相岐,王献平,安利佳,等.天蓝苜蓿原生质体培养再生植株[J ] .植物学报, 1996 , 38 (3) : 241~244
[109]刘明志.大叶紫花苜蓿愈伤组织原生质体再生植株[J ] .武汉植物学研究, 1996 ,14 (4) : 329~333
[110]王金,马永祥,李娟.紫花苜蓿营养成分及主要生物学特性[J ] .草业科学, 2003 ,20 (10) : 39~40
[111]黄新,叶红霞,舒小丽.紫花苜蓿耐逆诱变和转基因研究进展[ J ].核农学报, 2008, 22 (5) : 630 ~ 634
[112] Webb K J. Transformation of forage legumes using Agrobacteriumtum efactions[ J ]. TheorApp l Genet, 1986, 72 (1) : 53 ~ 58
[113]赵桂琴,慕平,张勃.紫花苜蓿基因工程研究进展[ J ].草业学报, 2006, 15 (6) : 9~18
[114] Saunders J M, Bingham E T. Production of alfalfa p lants from tissue culture[ J ]. Crop science, 1972, (12) : 804 ~ 809
[115] Song J S, Sorensen E L, Geoge H L. Direct embryogenesis from single mesophyll p rotop lasts in alfalfa[ J ]. Plant cell reports, 1990, (9) : 21 ~25.
[116]王瑛,朱宝成,孙毅,张琳宇,罗建平.外源lea3基因转化紫花苜蓿的研究[ J ].核农学报, 2007, 21 (3) : 249 ~ 252
[117]金湘,张建军,毛培宏.离子注入介导麻黄和甘草总DNA转化苜蓿的初步研究[ J ].生物技术, 2007, 17 (4) : 53~ 55
[118]包爱科,王强龙,张金林.苜蓿基因工程研究进展[ J ].分子植物育种, 2007, 5 (6) : 160 ~168
[119] Maqbool S B, Christou P. Multip le traits of agronomic importance in transgenic indica rice plants: analysis of transgene integration patterns, expression levels and stability [ J ]. Molecular breeding, 1999, 5 (5) : 471 ~ 480
[120] Akama K, Shiraishi H, Ohta S. Efficient transformation of A. thaliana: comparison of the efficiencies with various organs, p lant ecotypes and Agrobacterium strains[ J ]. Plant cell reports, 2004, 12(1) : 7 ~11
[121] Sugita K, Matsunage E, Ebinuma H. Effective selection system for generating marker - free transgenic p lants independent of sexual crossing[ J ]. Plant cell report, 1999, 18 (11) : 941~947
[122] Sugita K, Kasahara T, Matsunage E. A transformation vector for thep roduction ofmarker - free transgenic p lants containing a single copy transgene at high frequency[ J ]. Plant journal, 2000, 22 ( 5) : 461~ 469
[123] VaeckM, ReynaertsA, Hofte H. Transgenic p lants p rotected from insect attack[ J ]. Nature, 1987, 328: 33 ~ 37
[124] Sandhu J S, Qsadjan M D, Krasnyanski S F. Enhanced exp ression of the human resp iratory syncytial virus - F gene in app le leaf protoplasts[ J ]. Plant Cell Rep, 1999, 18: 394~ 397
[125] Rose A B, Beliakoff J A. Intron - mediated enhancement of gene expression independent of unique intron sequences and sp licing[ J ].Plant Physiology, 2000, 122: 535~ 542
[126] Ali S, TaylorW C. Quantitative regulation of the FlaveriaMe1 gene is controlled by the 3’- untranslated region and sequence near the amino terminus [ J ]. PlantMoleucular Biology, 2001, 46: 251 ~261
[127] Iannacone R, Grieco P D, Cellini F. Specific sequence modification of a cry3B endotoxin gene result in high levels of exp ression and insect resistance[ J ]. Plant molecular biology, 1997, 34 ( 3 ) : 485~ 496
[128]董江丽.利用转基因苜蓿生产轮状病毒可食用疫苗研究[D ].北京:中国农业大学, 2005
[129] Perlak F J, Deaton R W, Armstrong T A. Insect resistant cotton plants[ J ]. Naturebiotechnology, 1990, 8: 939~ 943
[130] Allen G C, Sp iker S, Thomp son W F. Use of matrix attachment regions tominimize transgene silencing[ J ]. Plantmolecular biology, 2000, 43 (2 -3) : 361~376
[131] Allen G C, Hall G E, Childs J. Scaffold attachment regions increase reporter gene exp- ression in stably transformed p lant cells[ J ]. Plant Cell, 1993, 5: 603~ 613
[132] BaucherM, VailheM A, Chabbert B. Down regulation of cinnamyl alcohol dehydrogenase in transgenic alfalfa and the effect on lignin composition and digestibility [ J ]. PlantMolecularBiology , 1999 ,39 : 437~447
[133] Guo D, Chen F, Inoue K. Down regulation of caffeic acid 3 - O -methyltransferase and caffeoyl CoA 3 - O - methyltransferase in transgenic alfalfa : impacts on lignin structure and imp- lications for the biosysthesis of G and S lignin[ J ]. The Plant Cell, 2001, 13 :73~ 88
[134] Jackson L A, Shadle GL, Zhou R, Nakashima J, Chen F, Dixon R A. Improving saccharification efficiency of alfalfa stems through modification of the terminal stages of monolignol biosynthesis [ J ]. Bioenergy research, 2008, 1 (3 - 4) : 180– 192
[135] GruberM Y, Ray H, Auser P. Genetic system for condensed tannin biotechnology [A ]. In : Gross G G, Hemingway R , Yoshida T.Plant Polyphenols [M ]. New York : Plenum Press , 1999: 315~341
[136] Tabe L M, Wardley - Richardson T, Ceriotti A. A biotechnological approach to improving the nutritive value of alfalfa[ J ]. J Anim Sci,1995, 73: 2752~2759
[137] Avaham T, Badani H, Galili S. Enhanced levels of methionine and cysteine in transgenic alfalfa p lants over - exp ression the Arabidop siscystathionine syntheses gene [ J ]. PlantBiotech J, 2005, 3 (1) : 71~79
[138]吕德扬,范云六,俞梅敏.苜蓿高含硫氨基酸蛋白转基因植株再生[ J ].遗传学报, 2000, 27 (4) : 331~337
[139] Winicov I, Bastola C R. Transgenic over - exp ression of the transcrip tion factor Alfin- enhances expression of the endogenous MsPRP- gene in alfalfa and improves salinity tolerance of plants[ J ].Plant Phys, 1999, 120: 473~480
[140] McKersie B D, Bowley S R, Jones K S. Winter survival of transgenic alfalfa over - exp ressing superoxide dismutase [ J ]. Plant Phys, 1999, 119: 839~847
[141] McKersie B D, Murnaghan J. Iron - superoxide dismutase exp ression in transgenic alfalfa increases winter survival without a detectable increase in photosynthetic oxidative stress tolerance [ J ]. Plant Physiol, 2000, 122 (4) : 1427~1437
[142] Zhang J Y, Broeckling C D. Overexp ression ofWXP1, a putative Medicago truncatula AP2 domain - containing transcrip tion factor gene, increases cuticular wax accumulation andenhances drought tolerance in transgenic alfalfa (M edicago sativa ) [ J ]. Plant J,2005, 42 (5) : 689~707
[143]韦晓阳.拟南芥镁离子转运蛋白基因A tMGT1对紫花苜蓿的遗传转化[D ].重庆:西南农业大学, 2008
[144] Guo Y, Biochemical Engineering for 2001, Springer-Verlag, Tokyo: 1992, 242~ 245
[145]张玉墀,阮茜华南理工大学学报(自然科学版) , 1993, 21: 70~ 73
[146]蔡燕璇,华南理工大学学报(自然学版) , 1991, 20( 4) : 94~ 99
[147] YE M,DAI J G,GUO H Z, et al1 Glucosylation of cinobufagin by cul2 tured suspension cells of Catharanthus roseus [J ]1 Tetrahedron Lett ,2002 ,43 (47) :8535~8538
[148] DAI J G,CUI YJ ,ZHU W H , et al1 Biotransformation of 2alpha ,5alpha , 10beta , 14beta2tetraacetoxy24 ( 20) , 112taxadieneby cell suspension cultures of Catharanthus roseus [ J ] Planta Med ,2002 ,68(12) :1113~1117
[149] HAMADA H,TANAKA T,FURUYA T, et al1 Hydroxylation of benzylic and allylic sites by plant cultured suspension cells [J ]1 Tetrahedron Lett ,2001 ,42(4) :909~911
[150] LIU Y, CHENG K D , ZHU P , et al1 Biotransformation of dehy-droepiandrosterone by hairy root cultures of Anisodus tanguticus [J ] Acta Pharm Sin (药学学报) , 2004 , 39(6) : 445~448
[151] Islam M O, Ichihashi S. Effects of sucrose, maltose and sorbitol on callus growth and plantlet regeneration in Phalaenosis, Doritaenop sis and Neofinetia [ J ]. J Japan Soc Hort Sci, 1999,68 (6) : 1124 ~ 1131.
[152] Wickremesinne ERM ,Arteca RN , Taxus Cell Cultures : Initiation , Growth Optimization , Characterization and Taxol Production ,Plant Cell ,Tissue and Organ Culture ,1993 ,35 :181~191
[153] Luis J . Pestchenker , Susan C. Roberts , and Michael L.Shuler ,kinetics of taxol production and nutrient use in suspension cultures of Taxus cuspidata in shake flasks and a wilson - type bioreactor. Enzyme Microbe. technol. ,1996 ,19 :256~162
[154] J . Guardiola ,J . L. lborra , and M. Canovas ,A Model That Links Growth and Secondary Metabolite Production in Plant Cell Suspension Cultures ,Biotech. & Bioeng. 1995 ,46 :291~301
[155]郑穗平.悬浮培养玫瑰茄细胞产生花青素的动力学研究〔D〕.广州:华南理工大学, 1998
[156]李弘剑.黄花蒿细胞培养过程中细胞生长及青蒿素生物合成的调节控制:〔D〕.广州:华南理工大学, 1994.
[157]王霜.大蒜细胞培养及产超氧化物歧化酶的动力学规律研究:〔M〕.广州:华南理工大学,1998.
[158] A shihara H, Tokoro T . Metabolic rate of inorganic phosphate absorbed by suspensioncultured cells of Catha-ranthus roseus. J P lant P hy siol, 1985, 118 (3) : 227 237.
[159] J.L. Fox, Turning plants into protein factories, Nature Biotechnology 2006 (24)1191~1193.
[160] J.C. Yin, G.X. Li, X.F. Ren, G. Herrler, Select what you need: a comparative evaluation of the advantages and limitations of frequently used expression systems for foreign genes, Journal of Biotechnology 2007 (127) 335~347.
[161] L.M. Houdebine, Production of pharmaceutical proteins by transgenic animals,Sang Thrombose Vaisseaux 2008 (20) 43–50.
[162] S. Lukjan, V. Srinivasan, G. Tous, S. Parekh, C. Swindell, M. Horn, Large-scale production of pharmaceutical proteins using plant cell suspension cultures,In Vitro Cellular & Developmental Biology-Animal 2007 (43) S50–S51.
[163] H.P. Sorensen, K.K. Mortensen, Advanced genetic strategies for recombinant protein expression in Escherichia coli, Journal of Biotechnology 2005 (115)113–128.
[164] L.M. Barnes, A.J. Dickson, Mammalian cell factories for efficient and stable protein expression, Current Opinion in Biotechnology 17 (2006) 381–386.
[165] V. Gomord, L. Faye, Posttranslational modification of therapeutic proteins in plants, Current Opinion in Plant Biology 2004 (7) 171–181.
[166] R.M. Twyman, E. Stoger, S. Schillberg, P. Christou, R. Fischer,Molecular farming in plants: hostsystems and expression technology, Trends in Biotechnology 2003 (21) 570–578.
[167]李志勇,悬浮培养大蒜细胞产生SOD的动力学研究〔D〕.广州:华南理工大学, 1998
[168] Ramachandra Rao S, Ravishankar GA. Biotransformation of protocatechuic aldehyde and caffeic acid to vanillin and capsaicin in freely suspended and immobilized cell cultures of Capsicum frutescens. J Biotechnol 2000a;76:137–46.
[169] Ramachandra Rao S, Ravishankar GA. Vanilla flavour: production by conventional and biotechnological routes.J Sci Food Agric 2000b;80:289–304.
[170] Ramachandra Rao S, Usha T, Ravishankar GA. Biotransformation of digitoxin to digoxin in cell cultures of Capsicum frutescens in the presence of b-cyclodextrin. Biocatal Biotransform 2002.
[171] Ravishankar GA, Ramachandra Rao S. Biotechnological production of phyto-pharmaceuticals. J Biochem, Mol .Biol Biophys 2000;4:73–102.
[172]郑穗平,玫瑰茄细胞悬浮培养过程的动力学研究[D],广州:华南理工大学,1998
[173] 172.林丽,范海延,潘野等,发根农杆菌Ri质粒及其在植物次生代谢物质生产中的应用[J],北方园艺,2007,11(5):65-68
[174]刘佳佳,高产黄酮苷和萜内酯银杏细胞系选育及黄酮苷积累调控的研究[D],广州:华南理工大学,1999
[175]黄伦云,小克银汉菌发酵产γ-亚麻酸的动力学研究[M],长沙:华中科技大学,2005
[176]魏明,霍山石斛类原球茎悬浮培养细胞生长和多糖合成的动力学研究[D],合肥:合肥工业大学,2006
[177]罗焕亮,胡桐细胞培养生产红厚壳素的细胞工程研究[D],广州:华南理工大学,2003
[178]李红,邝炎华,彭新湘,主要营养成分对甘蔗叶悬浮细胞生长影响的研究[J],华南农业大学学报,2002,4(30):66-69
[179]朱薿,红法夫酵母发酵合成虾青素工艺研究[M],长沙:华中科技大学,2007
[2]周立刚,郑光植,生物工程进展,11(1):29一35,1991
[3]郭勇,崔堂兵等主编.植物细胞培养技术与应用[ M] .北京:化学工业出版社, 2003. 12.
[4]韩军丽,李振秋,刘本叶等植物萜类代谢工程,生物工程学报2007,24(4): 561~569
[5] Powler M. W. Plnat cell culture. VolⅡ,edited by Russel G.E.,1984:41~67
[6]孙敬三,桂耀林主编.植物细胞工程试验技术.科学出版社,北京,1995, 49~72
[7]朱蔚华.人参的研究及报告.淑馨出版社,1989:121~168
[8]王文静,袁道强,高松洁植物组织培养的应用现状[ M] .河南师范大学学报, 2000, ( 3) : 137~139.
[9]孙志强,孙占育,植物组织培养技术在生产上的应用与进展,陕西农业科学,2010(3),98~103
[10]黄远新,玉永雄,胡艳,南方紫花苜蓿不同外植体离体培养的研究,中国草地,2003,25(3):43~47
[11]熊丽,吴丽芳.观赏花卉的组织培养与大规模生产[ M] .北京:北京化学工业出版社, 2003, 80~81.
[12]曾黎琼,段玉云,张云孙等,云南转基因农作物研究与产业化:现状及其发展,云南大学学报(自然科学版),2009,31(S2):479~488
[13]邝琦,余倩花,周厚高,植物细胞工程在菊花育种中的应用,北方园艺,2010(04):220~224
[14] JAMES C. Global status of commercialized Biotech /GMcrop s: 2006 [R ]. International Service for The Acquisi2tion ofAgri - Biotech App lications ( ISAAA). 2006.
[15] Goossens A ,Hakkinen ST,Laakso I , et al . A functional genomics approach toward the understanding of secondary metabolism in plant cells. Proc Natl Acad Sci USA ,2003 ,100 : 8595 - 8600.
[16] Winkel BSJ . Metabolic channeling in plants. Annu Rev Plant Biol ,2004 ,55 : 85 - 107.
[17] Kappers IF , Aharoni A , van Herpen TWJM, et al . Genetic engineering of terpenoid metabolism attracts bodyguards to Arabidopsis . Science ,2005 ,309 : 2070 - 2072.
[18]王兴春,李宏,王敏等。植物体细胞胚胎发生的调控网络,生物工程学报,2010,25;26(2):141~146
[19] Lange BM, Ketchum REB , Croteau RB. Isoprenoid biosynthesis. Metabolite profiling ofpeppermint oil gland secretory cells and application to herbicide target analysis. Plant Physiol ,2001 ,127 :305 - 314.
[20] Sandmann G, Romer S , Fraser PD. Understanding carotenoid metabolism as a necessity for genetic engineering of crop plants.Metabolic Engineering ,2006 ,8 : 291 - 302.
[21]于荣敏,金钱星,孙辉等。西洋参冠瘿组织悬浮培养及其人参皂苷类成分的分离,生物工程学报,2005,21(5):754~758
[22]钟建江,沈林南,王斯靖.植物细胞培养工程的最新进展.工业微生物,1995,25(l):25~29
[23]郑光植.植物细胞培养及其次级代谢.第1卷.云南大学出版社,昆明,1992:1~2
[24] Fujita Y, Hara Y, Suga C et al.Production of Shikonin derivatives by cell suspension cultures of Lithospermum erythrozhizon of shikonin derivatives.Plnat Cell Report, 1981,l:61~63
[25] Zenk M H. Plant tissue culture and its biotechnological application, W. Barz et al(Ed.), Springer-Verlag, New York, 1977:27~43
[26]罗建平,牛炳韬,贾敬芬等.云南红豆杉培养细胞系的建立.生物工程学报,1997,13(3):326~330
[27]沈征武,吴莲芬.紫杉醇研究进展.化学进展,1997,9(1):1~13
[28] Fowler M. W. Plant Biotechnology, Mantell(Ed.), Cambridge Univ. Press, 1983:3~7
[29]陈因良,陈志宏.细胞培养工程,华东化工学院出版社,上海,1992:275~276
[30] Drapeau D, Blanch H W, Wilke C R. Economic assessment of plant cell culture for the production of Agrnalicine. Biotechnol. Bioeng., 1987,30:946~953
[31] Tanaka H. Large-Scale cultuvition of plant cells at high density: A review. Process Biochemistry, 1987,8:106~113
[32] Dornenburg H, Knorr D. Strategies for the improvement of secondary metabolite production in plant cell cultures. Enzyme Microb.Technol.,1995,17:674~684
[33] Yoshikawa T., T.Furuya. Plant Cell Reports,1987,6:449~453
[34] Parr A J. Secondary products from plant cell cultures. Adv.Bio/technol Proc.,1988,9:l~34
[35] Brodelius P. The potential role of immobilization in plant cell biotechnolngy.Trends in Biotechnol.,1985,3:280~285
[36] Lindsey K., Yeoman M. M.The synthetic potential of immobilized cells of capsicum frutescens mill cv. Annuurm. Planta,1984,162:495~501
[37]韩迎山.植物细胞培养生产天然产物的研究进展。生物化学与生物物理学进展。1989,16(1):19~22
[38] Tsoulpha P, Doram P M. Solasodine production from self-immobilizated Solanum aviculare cells. J.Biotechnol.1991,19:99~110
[39] Hoedstra S S, Harkes P A A., Verpoorte R. et al. Effect of aucin on cytodifferentitation andproduction of quinoline alkaloids in compact globulas structures of Cinchona ledgeriana. Plant Cell Report, 1990,8:571~574
[40]王爱勤,何龙飞,林鉴钊。草本咖啡的组织培养和植株再生,广西农业生物科学,1999,18(2):6
[41]许建峰.高山红景天愈伤组织颗粒悬浮培养生产红景天甙动力学与过程控制.大连理工大学博士论文.1996
[42] Francois G, Christen P, Kapetanidis I. Prodution of valepotriates by hairy root cultures of Centranthus ruber DC. Plant Cell Report, 1995, 14:294~298
[43] Sang Y B, Henrik P. Two-phase airlift fermentor operation with elicitation for the enhanced production of Benzophenanthridine alkaloids in cell suspension of Escherichia Californica. Biotechnol. Bioeng., 1994,44:14~20
[44] Kim D J, Chang H N. Enhanced skikonin production from Lithospermam erythroihizon by in Situ extraction and Calcium alginate immobilization. Biotech. Bioeng., 1990,36:460~466
[45] Pyane G F, Pyane N N, Shuler M L et al. In Situ adsorption for enhanced alkaloid production by Catharanthus Roseus. Biotech. Letter, 1998,88:187~192
[46] Byun S Y, Pederson H, Chin C K. Two-phase culture for the enhanced production of benzophenanthridine alkaloids in cell suspensions of Eschancholtzia Californica. Phytochemistry, 1990,29(10):3135~3139
[47] Buitelaar R M, Langenhoff A A M, Heidstra R et al. Growth and thiophene production by hairy root cultures of Tagates patrla in various two-liquid-phase bioreactors. Enzyme Microb. Technol., 1991,13:487~494
[48]陈士云,候篙生.生产植物次生代谢物几项新技术研究进展.生物工程进展,1993,14(2):43~45
[49] Dicosmo F, Misawa M. Eliciting secondary metabolism in plant cell cultures. Trends in Biotechnol., 1985, 3:318~322
[50] Panda A K, Mishra, Saroj et al. Plant cell reactor- a perspective. Enzyme Micro. Technol., 1989,11:386~387
[51]陶璐璐,袁明静,徐锦堂.丹参愈伤组织细胞固定化及其转化产物的特征.生物工程学报,1990,6(3):218~223
[52]徐任生,天然产物化学,科学出版社,北京,1993,p856一893
[53] Fujita Y, Hara Y, Ogino T et al, Plant Cell Report, 1981,1:59~62
[54] Schlatmann. J. E, Mreno.P. R, Selles. M et al, Biotechnol. Bioeng., 1995,47:53~59
[55]甘烦远,郑光植,植物学通讯,1991,8(4):14~20
[56] Panda A K, Mishra, Saroj et al, Plant cell reactor - a perspective. Enzyme Micro.Technol.,1989,11:386~387
[57] Hashimoto. T, Pytochemistry, 1993,32:713~718
[58] Fesker. L. F, Plant Mol. Biol. 1993,23:11~21
[59] Phillipson J D. et al. The Chemistry and Biology of Isoquinoline, Alkaloids, springer-verlag, Berlin,1985,265
[60]刘涤.植物次生物质工业化生产研究的现况.生物工程学报,1987,3(1):9~15
[61]齐鸿雁编译,Jean Paul Schwitzguebal等著.从植物细胞到生物工程.生物工程进展,1992,12(5):24~28
[62]山根恒夫著,周斌译.生化反应工程,西北大学出版社,1992
[63]安东·莫泽尔著,曲音波等译.生物工艺技术学,湖南科学技术出版社,1994
[64] Shuler M L.Comprehensive biotechnology-The principles, application and regulations of biotechnology in industry, agriculture and medicine, Vol.2, Murray Moo-Young et al(Ed.), Cambridge Univ. Press, 1985,119~131
[65]戚义政,汪叔雄.生化反应动力学与反应器.化学工业出版社,1996,119
[66] L.拉皮德斯.化学反应器理论,石油工业出版社,1984,271~323.
[67]赵霞,叶勤,俞俊棠.影响重组工程菌发酵产率的因素.工业微生物,1995,25(2):26~32.
[68]郑穗平,郭勇.基因工程菌培养过程的动力学模型.微生物学通报,1998,25(l):46~48.
[69]俞俊棠、顾其丰、叶勤.生物化学工程,化学工业出版社,1991:68~73.
[70]白净.生理系统的仿真与建模,清华大学出版社,1994:211~240.
[71]杨冰.实用最优化方法及计算机程序,哈尔滨船舶工程学院出版社,1994.
[72] Brian S H, James M. L. Application of a new structured model to Tobacco cell cultures. Biotechnol. Bioeng.,2003,39:765~774
[73] Van Gulik W M, Ten Hoopen H J G, Heijnen J J. A structured model describing carbon and phosphate limited growth of catharanthus roses plant cell suspensions in batch and chemostat culture, Biotechnol. Bioeng,2005,41:771~780
[74] Bailey C M, Nicholsm H. A new structured model for plant cell culture. Biotechnol. Bioeng.,2003,34:1331~1336.
[75] Bailey C M, Nicholsm H. Optimal temperature control for a structured model of plant cell culture. Biotechnol. Bioeng.,2000,35:252~259
[76] Wei Zhang, Seki M., Furusaki S et al. A phase-segergated model for plant cell culture:the effect of cell volume fraction. Journal of Chemical Engineering of Japan, 2002:31(3):469~474
[77] Aynsley M, Ward A C., Wright A R. A mathematical model for the growth of mycelial Fungi in submerged culture.Biotechnol. Bioeng.,2005,35:820~830
[78] De Gunst M C M, Harkes P A A, van Zwet, Modelling the growth of a batch culture of plantcells; a corpuscular approach. Enzyme Micro. Technol., 2000,12:61~71
[79]颜日明,张志斌,邱晓芳等,杜仲细胞悬浮培养产黄酮及其动力学研究,中国生物工程杂志,2008,28(10):60~65
[80] Dohnal M. Fuzzy bioengineering models. Biotechnol. Bioeng., 2005,27:1146~1151
[81]王西娥,吴玉荷,烟草细胞悬浮培养生产CoQ10的动力学研究,仲恺农业技术学院学报,200619(4):9~12
[82]高正航,李卫东,长春花生物碱类药物研究概况,贵州农业科学,2005,33(6):94~96
[83] Schlatmann J E ,Moreno P R H , Ten Hoopen H J G, et al .Effect of oxygen and nut rient limitation on ajamalicine production and related enzyme actives in high density cutures of Catharant hus roseus[J ] . Biotechnol Bioeng ,1994 ,44 (4) :461~468.
[84] Drapeau D ,Blanch H W ,Wilke C R. Ajmalicine , serpentine and catharant hine accumulation in Cat harant hus roseus bioreactor cultures [J ] . Planta Med ,1987 (53) :373~376.
[85] Wei Zhang, Seki M., Furusaki S et al. A phase-segregated model for plant cell culture: the effect of cell volume fraction. Journal of Chemical Engineering of Japan, 1998:31(3):469~474
[86]艾江宁,贾景明,高山红景天细胞悬浮培养生长和产物积累动力学,沈阳药科大学学报,2009 8,26(8):653~656
[87] Aynsley M, Ward A C,. Wright A R. A mathematical model for the growth of mycelial Fungi in submerged culture. Biotechnol. Bioeng.,2008,35:820~830
[88]许建峰.高山红景天愈伤组织颗粒悬浮培养生产红景天甙动力学与过程控制。大连理工大学博士论文. 1996
[89] Gunst M C M, Harkes P A A, van Zwet. Modelling the growth of a batch culture of plant cells; a corpuscular approach. Enzyme Micro. Technol., 1999,12:61~71
[90] Dohnal M. Fuzzy bioengineering model. Biotechnol. Bioeng., 2005,27:1146~1151
[91] Baraniak B, Karas M.Antioxidative properties of chloroplast concentratesobtained by various methods from Lucerne juice [J].Journal of Animal andFeed Science, 2000, 9( 2) : 397～405.
[92]曹致中,席亚丽,赵波.苜蓿叶蛋白提取及深加工[A].首届中国苜蓿发展大会(论文集) [C].北京:中国草原学会, 2001.100～115.
[93]董宽虎,朱慧森.苜蓿产品的开发应用现状[A].第二届中国苜蓿发展大会(论文集) [C].北京:中国草原学会, 2003.210～213.
[94]郭佩玉.苜蓿叶蛋白工业化提取概述[A].第二届中国苜蓿发展大会(论文集) [C].北京:中国草原学会, 2003.129～130.
[95]强莉.苜蓿叶蛋白的加工及开发利用[J].饲料博览, 2002, 4: 43～51.
[96]薛琳,田丽萍,刘青广.苜蓿叶蛋白提取后副产品的开发与利用[J].江西饲料, 2006, 2: 18～19.
[97]杨丽娥,胡雪华.叶蛋白提取方法研究[J].上海交通大学学报(农业科学版) , 2003, 21( 12) : 66～70.
[98]李斌.剑麻组织和细胞培养诱导蛋白酶的研究.华南理工大学博士学位论文,1998
[99]马晖玲,卢欣石,曹致中.苜蓿原生质培养及体细胞杂交技术的应用[J ] .草业科学, 2007 ,24 (8) :64~68
[100] Johnson L B , Stuteville D L , Higgns R K, et al . Regeneration of alfalfa plant s from protoplast s of regen′s clone [ J ] . Plant Science Letters , 1981 , 20 : 297~304
[101] Lu D Y, Davey M R , Cocking E C. A comparison of t he cultural behaviour of protoplasts of leaves , cotyledonous and root of Medicago sati va [J ] . Plant Science Letters , 1983 , 31 : 87~89
[102] Gilmour D E , Davey M R , Cooling E C. Plant regeneration f rom cotyledon of wild Medicago species [ J ] . Plant Science ,1987 , 48 : 107~112
[103]王娟,李玉珠,师尚礼。苜蓿愈伤组织原生质体游离于培养[J].草地学报,2010,3(18):258~262
[104]赵红娟,张博,陈爱坪,等.酶解对苜蓿子叶原生质体分离效果的影响[J ] .草地学报, 2008 , 16 (1) : 24~29
[105]白静仁,何茂泰,袁清,等.野生黄花苜蓿叶肉原生质体培养和植株再生[J ] .草地学报, 1994 , 2 (1) : 59~63
[106]舒文华,耿华珠,孙勇如.紫花苜蓿原生质体培养与植株再生[J ] .草地学报, 1994 , 2 (1) : 40~44
[107]黄绍兴,王慧叶,占德扬.紫花苜蓿根原生质体植株再生[J ] .科技通报, 4 (11) : 35~39
[108]张相岐,王献平,安利佳,等.天蓝苜蓿原生质体培养再生植株[J ] .植物学报, 1996 , 38 (3) : 241~244
[109]刘明志.大叶紫花苜蓿愈伤组织原生质体再生植株[J ] .武汉植物学研究, 1996 ,14 (4) : 329~333
[110]王金,马永祥,李娟.紫花苜蓿营养成分及主要生物学特性[J ] .草业科学, 2003 ,20 (10) : 39~40
[111]黄新,叶红霞,舒小丽.紫花苜蓿耐逆诱变和转基因研究进展[ J ].核农学报, 2008, 22 (5) : 630 ~ 634
[112] Webb K J. Transformation of forage legumes using Agrobacteriumtum efactions[ J ]. TheorApp l Genet, 1986, 72 (1) : 53 ~ 58
[113]赵桂琴,慕平,张勃.紫花苜蓿基因工程研究进展[ J ].草业学报, 2006, 15 (6) : 9~18
[114] Saunders J M, Bingham E T. Production of alfalfa p lants from tissue culture[ J ]. Crop science, 1972, (12) : 804 ~ 809
[115] Song J S, Sorensen E L, Geoge H L. Direct embryogenesis from single mesophyll p rotop lasts in alfalfa[ J ]. Plant cell reports, 1990, (9) : 21 ~25.
[116]王瑛,朱宝成,孙毅,张琳宇,罗建平.外源lea3基因转化紫花苜蓿的研究[ J ].核农学报, 2007, 21 (3) : 249 ~ 252
[117]金湘,张建军,毛培宏.离子注入介导麻黄和甘草总DNA转化苜蓿的初步研究[ J ].生物技术, 2007, 17 (4) : 53~ 55
[118]包爱科,王强龙,张金林.苜蓿基因工程研究进展[ J ].分子植物育种, 2007, 5 (6) : 160 ~168
[119] Maqbool S B, Christou P. Multip le traits of agronomic importance in transgenic indica rice plants: analysis of transgene integration patterns, expression levels and stability [ J ]. Molecular breeding, 1999, 5 (5) : 471 ~ 480
[120] Akama K, Shiraishi H, Ohta S. Efficient transformation of A. thaliana: comparison of the efficiencies with various organs, p lant ecotypes and Agrobacterium strains[ J ]. Plant cell reports, 2004, 12(1) : 7 ~11
[121] Sugita K, Matsunage E, Ebinuma H. Effective selection system for generating marker - free transgenic p lants independent of sexual crossing[ J ]. Plant cell report, 1999, 18 (11) : 941~947
[122] Sugita K, Kasahara T, Matsunage E. A transformation vector for thep roduction ofmarker - free transgenic p lants containing a single copy transgene at high frequency[ J ]. Plant journal, 2000, 22 ( 5) : 461~ 469
[123] VaeckM, ReynaertsA, Hofte H. Transgenic p lants p rotected from insect attack[ J ]. Nature, 1987, 328: 33 ~ 37
[124] Sandhu J S, Qsadjan M D, Krasnyanski S F. Enhanced exp ression of the human resp iratory syncytial virus - F gene in app le leaf protoplasts[ J ]. Plant Cell Rep, 1999, 18: 394~ 397
[125] Rose A B, Beliakoff J A. Intron - mediated enhancement of gene expression independent of unique intron sequences and sp licing[ J ].Plant Physiology, 2000, 122: 535~ 542
[126] Ali S, TaylorW C. Quantitative regulation of the FlaveriaMe1 gene is controlled by the 3’- untranslated region and sequence near the amino terminus [ J ]. PlantMoleucular Biology, 2001, 46: 251 ~261
[127] Iannacone R, Grieco P D, Cellini F. Specific sequence modification of a cry3B endotoxin gene result in high levels of exp ression and insect resistance[ J ]. Plant molecular biology, 1997, 34 ( 3 ) : 485~ 496
[128]董江丽.利用转基因苜蓿生产轮状病毒可食用疫苗研究[D ].北京:中国农业大学, 2005
[129] Perlak F J, Deaton R W, Armstrong T A. Insect resistant cotton plants[ J ]. Naturebiotechnology, 1990, 8: 939~ 943
[130] Allen G C, Sp iker S, Thomp son W F. Use of matrix attachment regions tominimize transgene silencing[ J ]. Plantmolecular biology, 2000, 43 (2 -3) : 361~376
[131] Allen G C, Hall G E, Childs J. Scaffold attachment regions increase reporter gene exp- ression in stably transformed p lant cells[ J ]. Plant Cell, 1993, 5: 603~ 613
[132] BaucherM, VailheM A, Chabbert B. Down regulation of cinnamyl alcohol dehydrogenase in transgenic alfalfa and the effect on lignin composition and digestibility [ J ]. PlantMolecularBiology , 1999 ,39 : 437~447
[133] Guo D, Chen F, Inoue K. Down regulation of caffeic acid 3 - O -methyltransferase and caffeoyl CoA 3 - O - methyltransferase in transgenic alfalfa : impacts on lignin structure and imp- lications for the biosysthesis of G and S lignin[ J ]. The Plant Cell, 2001, 13 :73~ 88
[134] Jackson L A, Shadle GL, Zhou R, Nakashima J, Chen F, Dixon R A. Improving saccharification efficiency of alfalfa stems through modification of the terminal stages of monolignol biosynthesis [ J ]. Bioenergy research, 2008, 1 (3 - 4) : 180– 192
[135] GruberM Y, Ray H, Auser P. Genetic system for condensed tannin biotechnology [A ]. In : Gross G G, Hemingway R , Yoshida T.Plant Polyphenols [M ]. New York : Plenum Press , 1999: 315~341
[136] Tabe L M, Wardley - Richardson T, Ceriotti A. A biotechnological approach to improving the nutritive value of alfalfa[ J ]. J Anim Sci,1995, 73: 2752~2759
[137] Avaham T, Badani H, Galili S. Enhanced levels of methionine and cysteine in transgenic alfalfa p lants over - exp ression the Arabidop siscystathionine syntheses gene [ J ]. PlantBiotech J, 2005, 3 (1) : 71~79
[138]吕德扬,范云六,俞梅敏.苜蓿高含硫氨基酸蛋白转基因植株再生[ J ].遗传学报, 2000, 27 (4) : 331~337
[139] Winicov I, Bastola C R. Transgenic over - exp ression of the transcrip tion factor Alfin- enhances expression of the endogenous MsPRP- gene in alfalfa and improves salinity tolerance of plants[ J ].Plant Phys, 1999, 120: 473~480
[140] McKersie B D, Bowley S R, Jones K S. Winter survival of transgenic alfalfa over - exp ressing superoxide dismutase [ J ]. Plant Phys, 1999, 119: 839~847
[141] McKersie B D, Murnaghan J. Iron - superoxide dismutase exp ression in transgenic alfalfa increases winter survival without a detectable increase in photosynthetic oxidative stress tolerance [ J ]. Plant Physiol, 2000, 122 (4) : 1427~1437
[142] Zhang J Y, Broeckling C D. Overexp ression ofWXP1, a putative Medicago truncatula AP2 domain - containing transcrip tion factor gene, increases cuticular wax accumulation andenhances drought tolerance in transgenic alfalfa (M edicago sativa ) [ J ]. Plant J,2005, 42 (5) : 689~707
[143]韦晓阳.拟南芥镁离子转运蛋白基因A tMGT1对紫花苜蓿的遗传转化[D ].重庆:西南农业大学, 2008
[144] Guo Y, Biochemical Engineering for 2001, Springer-Verlag, Tokyo: 1992, 242~ 245
[145]张玉墀,阮茜华南理工大学学报(自然科学版) , 1993, 21: 70~ 73
[146]蔡燕璇,华南理工大学学报(自然学版) , 1991, 20( 4) : 94~ 99
[147] YE M,DAI J G,GUO H Z, et al1 Glucosylation of cinobufagin by cul2 tured suspension cells of Catharanthus roseus [J ]1 Tetrahedron Lett ,2002 ,43 (47) :8535~8538
[148] DAI J G,CUI YJ ,ZHU W H , et al1 Biotransformation of 2alpha ,5alpha , 10beta , 14beta2tetraacetoxy24 ( 20) , 112taxadieneby cell suspension cultures of Catharanthus roseus [ J ] Planta Med ,2002 ,68(12) :1113~1117
[149] HAMADA H,TANAKA T,FURUYA T, et al1 Hydroxylation of benzylic and allylic sites by plant cultured suspension cells [J ]1 Tetrahedron Lett ,2001 ,42(4) :909~911
[150] LIU Y, CHENG K D , ZHU P , et al1 Biotransformation of dehy-droepiandrosterone by hairy root cultures of Anisodus tanguticus [J ] Acta Pharm Sin (药学学报) , 2004 , 39(6) : 445~448
[151] Islam M O, Ichihashi S. Effects of sucrose, maltose and sorbitol on callus growth and plantlet regeneration in Phalaenosis, Doritaenop sis and Neofinetia [ J ]. J Japan Soc Hort Sci, 1999,68 (6) : 1124 ~ 1131.
[152] Wickremesinne ERM ,Arteca RN , Taxus Cell Cultures : Initiation , Growth Optimization , Characterization and Taxol Production ,Plant Cell ,Tissue and Organ Culture ,1993 ,35 :181~191
[153] Luis J . Pestchenker , Susan C. Roberts , and Michael L.Shuler ,kinetics of taxol production and nutrient use in suspension cultures of Taxus cuspidata in shake flasks and a wilson - type bioreactor. Enzyme Microbe. technol. ,1996 ,19 :256~162
[154] J . Guardiola ,J . L. lborra , and M. Canovas ,A Model That Links Growth and Secondary Metabolite Production in Plant Cell Suspension Cultures ,Biotech. & Bioeng. 1995 ,46 :291~301
[155]郑穗平.悬浮培养玫瑰茄细胞产生花青素的动力学研究〔D〕.广州:华南理工大学, 1998
[156]李弘剑.黄花蒿细胞培养过程中细胞生长及青蒿素生物合成的调节控制:〔D〕.广州:华南理工大学, 1994.
[157]王霜.大蒜细胞培养及产超氧化物歧化酶的动力学规律研究:〔M〕.广州:华南理工大学,1998.
[158] A shihara H, Tokoro T . Metabolic rate of inorganic phosphate absorbed by suspensioncultured cells of Catha-ranthus roseus. J P lant P hy siol, 1985, 118 (3) : 227 237.
[159] J.L. Fox, Turning plants into protein factories, Nature Biotechnology 2006 (24)1191~1193.
[160] J.C. Yin, G.X. Li, X.F. Ren, G. Herrler, Select what you need: a comparative evaluation of the advantages and limitations of frequently used expression systems for foreign genes, Journal of Biotechnology 2007 (127) 335~347.
[161] L.M. Houdebine, Production of pharmaceutical proteins by transgenic animals,Sang Thrombose Vaisseaux 2008 (20) 43–50.
[162] S. Lukjan, V. Srinivasan, G. Tous, S. Parekh, C. Swindell, M. Horn, Large-scale production of pharmaceutical proteins using plant cell suspension cultures,In Vitro Cellular & Developmental Biology-Animal 2007 (43) S50–S51.
[163] H.P. Sorensen, K.K. Mortensen, Advanced genetic strategies for recombinant protein expression in Escherichia coli, Journal of Biotechnology 2005 (115)113–128.
[164] L.M. Barnes, A.J. Dickson, Mammalian cell factories for efficient and stable protein expression, Current Opinion in Biotechnology 17 (2006) 381–386.
[165] V. Gomord, L. Faye, Posttranslational modification of therapeutic proteins in plants, Current Opinion in Plant Biology 2004 (7) 171–181.
[166] R.M. Twyman, E. Stoger, S. Schillberg, P. Christou, R. Fischer,Molecular farming in plants: hostsystems and expression technology, Trends in Biotechnology 2003 (21) 570–578.
[167]李志勇,悬浮培养大蒜细胞产生SOD的动力学研究〔D〕.广州:华南理工大学, 1998
[168] Ramachandra Rao S, Ravishankar GA. Biotransformation of protocatechuic aldehyde and caffeic acid to vanillin and capsaicin in freely suspended and immobilized cell cultures of Capsicum frutescens. J Biotechnol 2000a;76:137–46.
[169] Ramachandra Rao S, Ravishankar GA. Vanilla flavour: production by conventional and biotechnological routes.J Sci Food Agric 2000b;80:289–304.
[170] Ramachandra Rao S, Usha T, Ravishankar GA. Biotransformation of digitoxin to digoxin in cell cultures of Capsicum frutescens in the presence of b-cyclodextrin. Biocatal Biotransform 2002.
[171] Ravishankar GA, Ramachandra Rao S. Biotechnological production of phyto-pharmaceuticals. J Biochem, Mol .Biol Biophys 2000;4:73–102.
[172]郑穗平,玫瑰茄细胞悬浮培养过程的动力学研究[D],广州:华南理工大学,1998
[173] 172.林丽,范海延,潘野等,发根农杆菌Ri质粒及其在植物次生代谢物质生产中的应用[J],北方园艺,2007,11(5):65-68
[174]刘佳佳,高产黄酮苷和萜内酯银杏细胞系选育及黄酮苷积累调控的研究[D],广州:华南理工大学,1999
[175]黄伦云,小克银汉菌发酵产γ-亚麻酸的动力学研究[M],长沙:华中科技大学,2005
[176]魏明,霍山石斛类原球茎悬浮培养细胞生长和多糖合成的动力学研究[D],合肥:合肥工业大学,2006
[177]罗焕亮,胡桐细胞培养生产红厚壳素的细胞工程研究[D],广州:华南理工大学,2003
[178]李红,邝炎华,彭新湘,主要营养成分对甘蔗叶悬浮细胞生长影响的研究[J],华南农业大学学报,2002,4(30):66-69
[179]朱薿,红法夫酵母发酵合成虾青素工艺研究[M],长沙:华中科技大学,2007