An optimized protocol using Steedman’s wax for high-sensitivity RNA in situ hybridization in shoot apical meristems and flower buds of cucumber

作     者：WANG Cui SUN Jin-jing YANG Xue-yong WAN Li ZHANG Zhong-hua ZHANG Hui-min WANG Cui;SUN Jin-jing;YANG Xue-yong;WAN Li;ZHANG Zhong-hua;ZHANG Hui-min

作者机构：College of HorticultureQingdao Agricultural University/Engineering Laboratory of Genetic Improvement of Horticultural Crops of Shandong Province/Laboratory of Quality&Safety Risk Assessment for Fruit(Qingdao)Ministry of Agriculture and Rural AffairsQingdao 266109P.R.China Institute of Vegetables and FlowersChinese Academy of Agricultural Sciences/Key Laboratory of Biology and Genetic Improvement of Horticultural Crops of the Ministry of Agriculture and Rural Affairs/Sino-Dutch Joint Laboratory of Horticultural GenomicsBeijing 100081P.R.China 

出 版 物：《Journal of Integrative Agriculture》 (农业科学学报（英文版）)

年 卷 期：2023年第22卷第2期

页      面：464-470页

核心收录：

学科分类：09[农学] 0902[农学-园艺学] 090202[农学-蔬菜学] 

基　　金：supported by the National Natural Science Foundation of China(32002036) 

主　　题：cucumber in situ hybridization Steedman’s wax paraffin wax 

摘      要：In situ mRNA hybridization(ISH)is a powerful tool for examining the spatiotemporal expression of genes in shoot apical meristems and flower buds of *** most common ISH protocol uses paraffin wax;however,embedding tissue in paraffin wax can take a long time and might result in RNA degradation and decreased ***,we developed an optimized protocol to simplify the process and improve RNA *** combined embedding tissue in low melting-point Steedman’s wax with processing tissue sections in solution,as in the whole-mount ISH method in the optimized *** the optimized protocol,we examined the expression patterns of the CLAVATA3(CLV3)and WUSCHEL(WUS)genes in shoot apical meristems and floral meristems of Cucumis sativus(cucumber)and Arabidopsis thaliana(Arabidopsis).The optimized protocol saved 4–5 days of experimental period compared with the standard ISH protocol using paraffin ***,the optimized protocol achieved high signal *** optimized protocol was successful for both cucumber and Arabidopsis,which indicates it might have general applicability to most plants.