欢迎访问作物学报，今天是
作物学报

作物学报 ›› 2010, Vol. 36 ›› Issue (2): 228-232.doi: 10.3724/SP.J.1006.2010.00228

• 作物遗传育种·种质资源·分子遗传学 • 上一篇    下一篇

能以植酸为唯一磷源生长的转基因甘蓝型油菜

方小平1，王转1，陈茹梅2，李均1，范云六2，罗莉霞1，陈坤荣1，任莉1   

  1. 1 中国农业科学院油料作物研究所 / 农业部油料作物遗传改良重点开放实验室，湖北武汉 430062; 2 中国农业科学院生物技术研究所，北京 100081
  • 收稿日期:2009-07-14 修回日期:2009-10-02 出版日期:2010-02-10 网络出版日期:2009-12-21
  • 通讯作者: xpfang2008@163.com; xpfang@public.wh.hb.cn
  • 基金资助:

    本研究由国家自然科学基金项目（30270791）和国家高技术研究发展计划（863计划）项目（2002AA212011）资助。

Transgenic Brassica napus Growing with Phytate as a Sole Phosphorus Source

FANG Xiao-Ping1,WANG Zhuan1,CHEN Ru-Mei2,LI Jun1,FAN Yun-Liu2,LUO Li-Xia1,CHEN Kun-Ron1,REN Li1   

  1. 1 Key Laboratory of Oil Crops Genetic Improvement, Ministry of Agriculture / Oil Crops Research Institute, Chinese Academy of Agricultural Sciences, Wuhan 430062, China; 2 Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, Beijing 100081, China
  • Received:2009-07-14 Revised:2009-10-02 Published:2010-02-10 Published online:2009-12-21
  • Contact: xpfang2008@163.com; xpfang@public.wh.hb.cn

PDF

1276

被引次数

12

摘要：

由于难溶性矿物磷酸盐和有机磷的大量存在，土壤总磷虽高，但多数土壤缺乏有效磷，这是世界范围内农业生产的主要限制因素之一。土壤中有机磷的存在形式主要是植酸，为了提高土壤有效磷供给，利用根癌农杆菌菌株LBA4404，通过两步再生方法将植物表达载体pBINPR-phyI中含有的带胞外分泌信号肽序列的植酸酶基因转入油菜品种中双6号中，并获得转基因油菜56株，转化效率0.16%~9.20%。分子检测和酶活性检验表明，植酸酶基因已导入中双6号，转基因植株能有效表达植酸酶基因并向根外分泌植酸酶，转基因植株能以植酸为唯一磷源正常生长，非转基因植株则不能。结果表明，转基因植株根系分泌大量高活性植酸酶有助于土壤中有机磷释放出有效磷供植物利用；利用基因工程技术提高植物对土壤闭蓄态有机磷利用效率的前景是可期待的。

关键词: 油菜, 植酸酶, 转基因, 胞外分泌

Abstract:

Phosphorus (P) deficiency in soil is a major constraint in agricultural production worldwide. Most soils contain significant amounts of total soil P that occurs in insoluble inorganic and organic fractions, but lack available phosphorus. Phytic acid is the major storage form of organic phosphorus in soil. In this experiment, the phytase gene with the signal peptide sequence of extracellular secretion was introduced into Brassica napus cv. Zhongshuang 4 via Agrobacterium tumefaciens LBA4404. Fifty six plants of transgenic Brassica napus were obtained and checked by PCR and phytase activity, most of them gave the positive results. The transformation efficiency was 0.16–9.2%. When grown in MS medium with phytic acid as a sole phosphorus source under sterile conditions, transgenic Brassica napus plants were able to obtain inorganic phosphate from phytic acid and grew normally, but the wild-type plants not. These results show that extracellular phytase secreted from plant roots is a significant factor in the utilization of phosphorus from phytate and indicate that there exists a prospect for using gene technology to improve the ability of plants to utilize accumulated forms of soil organic phosphorus.

Key words: Brassica napus, Phytase, Transgene, Extracellular secretion

[1] Atkinson D. Some general effects of phosphorus deficiency on growth and development. New Phytol, 1973, 72: 101-111

[2] Institute of Soil Science of Chinese Academy of Sciences (中国科学院南京土壤研究所主编). Chinese Soil (中国土壤). Beijing: Science Press, 1978. pp 376-391(in Chinese)

[3] Bielesk R L.Phosphate pools, phosphate transport, and phosphate availability. Annu Rev Plant Physiol, 1973, 24: 225-252

[4] Dalal R C. Soil organic phosphorus. Adv Agron, 1977, 29: 83-117

[5] Anderson G. Assessing organic phosphorus in soil. In: Khasawneh F E, Sample E C Kamprath E J, eds. The Role of Phosphorus in Agriculture. Madison, USA: Amer. Soc. Agron. 1980. pp 411-432

[6] Turner B L, Mahieu N, Condron L M. Quantification of myoinositol hexaphosphate in alkaline soil extracts by solution 31P-NMR spectroscopy and spectral deconvolution. Soil Sci, 2003, 168: 469-478

[7] Turner B L, Papha´zy M J, Haygarth P M, McKelvie I D. Inositol phosphates in the environment. Philo Rans Royal Soc, Series B, 2002, 357: 449-469

[8] Raboy V. Progress in breeding low phytate crops. -505J Nutr, 2002, 132: 503

[9] Yao B(姚斌), Fan Y-L(范云六). Molecular biology and gene engineering of phytase. Chin J Biotechnol生物工程学报), 2000, 16: 1-5 (in Chinese with English abstract) (

[10] Reddy N R, Sathe S K. Occurrence, Distribution, Content and Dietary Intake of Phytate. In: Food Phytates. Boca Raton: CRC press, 2002. pp 25-52

[11] Sun H-G(孙海国), Zhang F-S(张福锁). Effect of phosphorus deficiency on activity of acid phosphates exuded by wheat roots. Chin J Appl Ecol (应用生态学报), 2002, 13(3): 379-381 (in Chinese with English abstract)

[12] Irving G C J. Phytase. In: Cosgrove D J ed. Inositol Phosphates: Their Chemistry, Biochemistry and Physiology (Studies in Organic Chemistry 4). 1980. pp 85-186

[13] Hayes J E, Richardson A E, Simpson R J. Components of organic phosphorus in soil extracts that are hydrolysed by phytase and acid phosphatase. Biol Fertil Soils, 2000, 32: 279-286

[14] Hayes J E, Richardson A E, Simpson R J. Phytase and acid phosphatase activities in extracts from roots of temperate pasture grass and legume seedlings. Aust J Plant Physiol, 1999, 26: 801-809

[15] Richardson A E, George T S, Hens M, Simpson R J. Utilisation of soil organic phosphorus by higher plants. In: Turner B L, Frossard E, Baldwin D, eds. Organic Phosphorus in the Environment. Wallingford: CABI Publishing. 2004. pp 165-184

[16] Kong F-L(孔凡利), Ling W-L(林文量), Yan X-L(严小龙), Liao H(廖红). Phytate-phosphorus uptake and utilization by transgenic tobacco carrying Bacillus subtilis phytase gene. Chin J Appl Ecol (应用生态学报), 2005, 16(12): 2225-2478 (in Chinese with English abstract)

[17] Zou L-K邹立扣)，Wang H-N王红宁). Phytase and its plant genetic engineering. (((Microbiology微生物学通报), 2005, 32(6): 128-132 (in Chinese with English abstract)

[18] Chan W L, Lung S C, Lim B L.-106 Properties of beta-propeller phytase expressed in transgenic tobacco. Protein Exp Purif, 2006, 46: 100

[19] Shen Y-O(沈亚欧), Peng H-W(彭焕伟), Pan G-T(潘光堂). Research study on expressed phytase in transgenic plants. Chin J Biotechnol (中国生物工程杂), 2005, 25(1): 29-32 (in Chinese with English abstract)

[20] Xiao K, Harrison M J, Wang Z Y.-36 Transgenic expression of a novel M. truncatula phytase gene results in improved acquisition of organic phosphorus by Arabidopsis. Planta, 2005, 222: 27

[21] Hong C Y, Cheng K J, Tseng T H, Wang C S, Liu L F, Yu S M.-39 Production of two highly active bacterial phytases with broad pH optima in germinated transgenic rice seeds. Transgenic Res, 2004, 13: 29

[22] Richardson A E, Hadobas P A, Hayes J E. Extracellular secretion of Aspergillus phytase from Arabidopsis roots enables plants to obtain phosphorus from phytate. Plant J, 2001, 25: 641-649

[23] Mudge S R, Smith F W, Richardson A E. Root-specific and phosphate-regulated expression of phytase under the control of a phosphate transporter promoter enables Arabidopsis to grow on phytate as a sole P source. Plant Sci, 2003, 165: 871-878

[24] Yan X-L(阎秀兰), Duan H-Y(段海燕), Wang Y-H(王运华). Phosphorus efficiency of different rape (Brassica napus L.) genotypes. Chin J Oil Crop Sci(中国油料作物学报), 2002, 24(2): 47-49 (in Chinese with English abstract)

[25] Ponstein A S, Bade J B, Verwoerd T C, MolendijkStable expression of Phytase (phyA) in canola (Brassica napus) seeds: Towards a commercial product. Mol Breed, 2002, 10: 31-44 L, Storms J, Beudeker R F and Pen J.

[26] Murray M G, Thompson W F. Rapid iso1ation of high mo1ecular weight plant.DNA Nucl Acid Res, 1980, 8: 4321-4325

[27] Sambrook J, Fritsch E F, Maniatis T. Molecular Cloning: A Laboratory Manual. New York: Cold Spring Harbor Laboratory Press, 2002

[28] Yan X-P(闫新甫). Transgenic Plants (转基因植物). Beijing: Science Press, 2003 (in Chinese)

[29] Moloney M M, Walker J M, Sharma K K. High efficiency transformation of Brassica napus using Agrobacterium vectors. Plant Cell Rep, 1989, 8: 238-242

[30] Xie J-K(谢建坤), Xiong H-J(熊焕金), Wan Y(万勇), Xiao Y-Q(肖叶青). Advances in plant gene transformation techniques of Brassica mediated by Agrobacterium tumefaciens. Acta Agric Jiangxi (江西农业学报), 2002, 14(2): 38-44 (in Chinese with English abstract)
[1] 陈松余, 丁一娟, 孙峻溟, 黄登文, 杨楠, 代雨涵, 万华方, 钱伟. 甘蓝型油菜BnCNGC基因家族鉴定及其在核盘菌侵染和PEG处理下的表达特性分析[J]. 作物学报, 2022, 48(6): 1357-1371.
[2] 秦璐, 韩配配, 常海滨, 顾炽明, 黄威, 李银水, 廖祥生, 谢立华, 廖星. 甘蓝型油菜耐低氮种质筛选及绿肥应用潜力评价[J]. 作物学报, 2022, 48(6): 1488-1501.
[3] 单露英, 李俊, 李亮, 张丽, 王颢潜, 高佳琪, 吴刚, 武玉花, 张秀杰. 转基因玉米NK603基体标准物质研制[J]. 作物学报, 2022, 48(5): 1059-1070.
[4] 黄伟, 高国应, 吴金锋, 刘丽莉, 张大为, 周定港, 成洪涛, 张凯旋, 周美亮, 李莓, 严明理. 芥菜型油菜BjA09.TT8和BjB08.TT8基因调节类黄酮的合成[J]. 作物学报, 2022, 48(5): 1169-1180.
[5] 雷新慧, 万晨茜, 陶金才, 冷佳俊, 吴怡欣, 王家乐, 王鹏科, 杨清华, 冯佰利, 高金锋. 褪黑素与2,4-表油菜素内酯浸种对盐胁迫下荞麦发芽与幼苗生长的促进效应[J]. 作物学报, 2022, 48(5): 1210-1221.
[6] 石育钦, 孙梦丹, 陈帆, 成洪涛, 胡学志, 付丽, 胡琼, 梅德圣, 李超. 通过CRISPR/Cas9技术突变BnMLO6基因提高甘蓝型油菜的抗病性[J]. 作物学报, 2022, 48(4): 801-811.
[7] 袁大双, 邓琬玉, 王珍, 彭茜, 张晓莉, 姚梦楠, 缪文杰, 朱冬鸣, 李加纳, 梁颖. 甘蓝型油菜BnMAPK2基因的克隆及功能分析[J]. 作物学报, 2022, 48(4): 840-850.
[8] 黄成, 梁晓梅, 戴成, 文静, 易斌, 涂金星, 沈金雄, 傅廷栋, 马朝芝. 甘蓝型油菜BnAPs基因家族成员全基因组鉴定及分析[J]. 作物学报, 2022, 48(3): 597-607.
[9] 王瑞, 陈雪, 郭青青, 周蓉, 陈蕾, 李加纳. 甘蓝型油菜白花基因InDel连锁标记开发[J]. 作物学报, 2022, 48(3): 759-769.
[10] 赵改会, 李书宇, 詹杰鹏, 李晏斌, 师家勤, 王新发, 王汉中. 甘蓝型油菜角果数突变体基因的定位及候选基因分析[J]. 作物学报, 2022, 48(1): 27-39.
[11] 王渭霞, 赖凤香, 胡海燕, 何佳春, 魏琪, 万品俊, 傅强. 超低温11年保存期对转基因作物基体标准样品核酸检测的影响[J]. 作物学报, 2022, 48(1): 238-248.
[12] 娄洪祥, 姬建利, 蒯婕, 汪波, 徐亮, 李真, 刘芳, 黄威, 刘暑艳, 尹羽丰, 王晶, 周广生. 种植密度对油菜正反交组合产量与倒伏相关性状的影响[J]. 作物学报, 2021, 47(9): 1724-1740.
[13] 张建, 谢田晋, 尉晓楠, 王宗铠, 刘崇涛, 周广生, 汪波. 无人机多角度成像方式的饲料油菜生物量估算研究[J]. 作物学报, 2021, 47(9): 1816-1823.
[14] 王艳花, 刘景森, 李加纳. 整合GWAS和WGCNA筛选鉴定甘蓝型油菜生物产量候选基因[J]. 作物学报, 2021, 47(8): 1491-1510.
[15] 李杰华, 端群, 史明涛, 吴潞梅, 柳寒, 林拥军, 吴高兵, 范楚川, 周永明. 新型抗广谱性除草剂草甘膦转基因油菜的创制及其鉴定[J]. 作物学报, 2021, 47(5): 789-798.
Viewed
Full text


Abstract

Cited
  Shared   
  Discussed   
No Suggested Reading articles found!
京ICP备15008789号-2