拟南芥草酸不敏感突变体的筛选与分析

陈晓婷1,…

生物工程学报 Chin J Biotech 2008, February 25; 24(2): 203-208

journals.im.ac.cn Chinese Journal of Biotechnology ISSN 1000-3061

Received: June 8, 2007; Accepted: July 23, 2007

Supported by: the National Natural Sciences Foundation of China (Nos. 30671347 and 30471178).

Corresponding author: Zonghua Wang. Tel: +86-591-83790312; Fax: +86-591-83727618; E-mail: wangzh@fjau.edu.cn

国家自然科学基金资助项目(Nos. 30671347, 30471178)。

研究报告

拟南芥草酸不敏感突变体的筛选与分析

陈晓婷1, 2, 陈芳芳2, 陈立群1, 郑麟1, 鲁国东2, 王宗华1, 2

1 福建农林大学生命科学学院, 福州 350002 2 生物农药与化学生物学教育部重点实验室, 福州 350002

摘要: 草酸是多种真菌的致病因子。在含1.2 mmol/L 草酸和10 μmol/L 雌二醇的MS缺钙培养基上, 从大约含6000个独立株系的拟南芥化学诱导突变体库中筛选草酸不敏感的突变体。初筛获得的可能的草酸不敏感突变体单株收种后, 进一步复筛获得5株较抗草酸的突变体D33、D74、D154、D282和D630。对它们的TAIL-PCR的第三步产物回收、测序、比对的结果表明：D33的T-DNA插入位点位于At2g39720 (Zinc finger ) and At2g39730 (Rubisco activase) 之间, D74、D154、D282和D630都插在At5g10450 (14-3-3 protein GF14 lambda) 的第一个内含子上。突变体后继的遗传分析与分子分析正在进行中。

关键词: 拟南芥, 突变体筛选, 草酸不敏感, TAIL-PCR

Isolation and Analysis of Oxalic Acid Insensitive Mutant of Arabidopsis thaliana

Xiaoting Chen1, 2, Fangfang Chen2, Liqun Chen1, Lin Zheng1, Guodong Lu2, and Zonghua Wang1, 2

1 College of Life Sciences, Fujian Agriculture and Forestry University, Fuzhou 350002, China 2 Key Laboratory of Bio-pesticide and Chemistry Biology, Ministry of Education, Fuzhou 350002, China

Abstract: Oxalic acid (OA) is inhibitory to many fungal plant pathogens. To further characterize the molecular mechanism of OA involved in fungal pathogenesis, OA insensitive mutants were screened from a chemical inducible Arabidopsis mutant library (about 6000 lines) using MS medium (calcium free) containing 1.2 mmol/L OA and 10 μmol/L estradiol. Harvested putative mutants were collected separately. Individual lines of mutants were screened again on modified MS medium containing OA. Mutants D33, D74, D154, D282 and D630 with enhanced OA resistance were obtained. The T-DNA flanking sequences were amplified by TAIL-PCR. The sequences were blasted against TAIR database. The result indicated that the T-DNA of mutant D33 was inserted between At2g39720 (zinc finger) and At2g39730 (Rubisco activase), and the T-DNA junctions of the other four mutants were the same, all inserted in the same site of the first intron of At5g10450 (14-3-3 protein GF14 lambda).

Keywords: Arabidopsis thaliana, mutant screening, OA insensitive, TAIL-PCR

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