| 利用 HPLC 分析菠萝愈伤组织 DNA 甲基化水平 |
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| 引用本文： | 林文秋,肖熙鸥,张红娜 刘胜辉,孙伟生,李运合,吴青松.利用 HPLC 分析菠萝愈伤组织 DNA 甲基化水平[J].热带作物学报,2018,39(11):2220-2224. |
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| 作者姓名： | 林文秋 肖熙鸥 张红娜 刘胜辉 孙伟生 李运合 吴青松 |
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| 作者单位： | 1. 中国热带农业科学院南亚热带作物研究所，广东湛江 524091；
2. 海南省菠萝种质创新与利用工程技术研究中心，广东 湛江 524091；
3. 农业部热带果树生物学重点实验室，广东湛江 524091 |
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| 摘 要： | 建立菠萝 DNA 甲基化水平的 HPLC 测定方法，分析菠萝愈伤组织 DNA 甲基化水平变化，为进一步研究菠萝 体细胞无性系变异机理奠定基础。通过对流动相和水解温度等条件的优化，建立菠萝 DNA 甲基化水平的检测方法。结 果表明，分离 C 和 5m-C 的最佳流动相为甲醇∶磷酸二氢钾∶三乙胺为 10∶90∶0.2（V/V），pH 3.0，DNA 的最佳水解 温度为 90 ℃。利用此体系分析菠萝愈伤组织和胚性愈伤组织的 DNA 甲基化变化，结果表明，菠萝愈伤组织在分化过 程中 DNA 总甲基化水平呈动态变化，变化范围为 5.14%~96.86%。此外，胚性愈伤组织甲基化水平低于非胚性愈伤组 织。推测 DNA 甲基化影响菠萝愈伤组织的分化及胚性愈伤组织的形成。
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| 关 键 词： | 菠萝 愈伤组织 DNA甲基化 HPLC |
Analysis of DNA Methylation Levels in Pineapple Callus by HPLC |
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| LIN Wenqiu,XIAO Xi’ou ZHANG Hongna LIU Shenghui,SUN Weisheng,LI Yunhe,WU Qingsong.Analysis of DNA Methylation Levels in Pineapple Callus by HPLC[J].Chinese Journal of Tropical Crops,2018,39(11):2220-2224. |
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| Authors: | LIN Wenqiu XIAO Xi’ou ZHANG Hongna LIU Shenghui SUN Weisheng LI Yunhe WU Qingsong |
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| Institution: | 1. South Subtropical Crop Research Institute, Chinese Academy of Tropical Agricultural Sciences, Zhanjiang, Guangdong 524091, China;
2. Hainan Provincial Engineering Research Center for Pineapple Germplasm Innovation and Utilization, Zhanjiang, Guangdong 524091, China;
3. Key Laboratory of Tropical Fruit Biology, Ministry of Agriculture, Zhanjiang, Guangdong 524091, China |
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| Abstract: | In order to lay a foundation of the mechanism of somaclonal variation in pineapple, the analysis of the DNA methylation levels of callus was established by HPLC . A method for the determination of the DNA methylation levels was established by optimized the mobile phase and hydrolysis temperature. The results showed that the optimum mobile phase was the combination A (10% V/V methanol, 90% V/V KH2PO4, 0.2% V/V triethylamine in H3PO4 adjusted to pH 3.0). The hydrolysis temperature was 90 ℃. The analysis of the DNA methylation levels in pineapple callus indicated that the DNA methylation levels showed dynamic changes with pineapple callus which ranged from 5.14% to 96.86%. In addition, the DNA methylation level of embryogenic callus was lower than non-embryogenic callus. It is speculated that the differentiation of callus and formation of embryogenic callus were influenced by DNA methylation. |
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| Keywords: | pineapple callus DNA methylation HPLC |
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