北京大学学报自然科学版 ›› 2022, Vol. 58 ›› Issue (2): 195-200.DOI: 10.13209/j.0479-8023.2022.019

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水稻整体原位杂交实验技术方法的改进

王东辉   

  1. 生物学国家级实验教学示范中心, 蛋白质与植物基因研究国家重点实验室, 北京大学生命科学学院, 北京 100871
  • 收稿日期:2021-07-06 修回日期:2021-09-15 出版日期:2022-03-20 发布日期:2022-03-20
  • 通讯作者: 王东辉, E-mail: wangdh(at)pku.edu.cn
  • 基金资助:
    蛋白质与植物基因研究国家重点实验室开放运行费(8102000374/006)资助

Improvement of Experimental Techniques of Whole Mount in situ Hybridization Technology in Rice

WANG Donghui   

  1. School of Life Sciences, National Teaching Center for Experimental Biology, The State Key Laboratory of Protein and Plant Gene Research, Peking University, Beijing 100871
  • Received:2021-07-06 Revised:2021-09-15 Online:2022-03-20 Published:2022-03-20
  • Contact: WANG Donghui, E-mail: wangdh(at)pku.edu.cn

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摘要：

以检测水稻SUPERWOMAN1 (SPW1)/OsMADS16基因的表达模式为例, 通过对杂交探针的制备、材料的固定及解离通透、显色等方面进行优化, 获得背景值低、特异性高的SPW1/OsMADS16基因特异性表达结果。建立一套针对水稻的整体原位杂交技术, 方法简单, 费用低廉, 可在离心管中多种材料同时进行。主要实验步骤包括探针制备、材料固定和解离、杂交反应、杂交后处理和检测显色等。提出的方法可为高通量检测水稻基因表达的时空模式及其功能分析提供技术条件。

关键词: 整体原位杂交技术(WISH), 水稻, SUPERWOMAN1/OsMADS16 基因

Abstract:

Taking rice SUPERWOMAN1 (SPW1)/OsMADS16 gene as an example, by optimizing the preparation of hybridization probes, material immobilization, dissociation, permeabilization and color development, the SPW1/OsMADS16 gene expression patterns with high specificity but low backgrounds were obtained. The procedure of whole mount in situ hybridization (WISH) technology for rice is simple and inexpensive, and it can be performed simultaneously with several species in the centrifuge tube. The main experimental steps include probe preparation, material immobilization and dissociation, hybridization reaction, post-hybridization treatment and detection and color development. The proposed method provides a technological basement for a protentional high throughput detection of spatiotemporal gene expression patterns and functional analysis in rice.

Key words: whole mount in situ hybridization (WISH), rice, SUPERWOMAN1/OsMADS16

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