The isolation of early nuclear endosperm of Oryza sativa to facilitate gene expression analysis and screening imprinted genes.
The isolation of early nuclear endosperm of Oryza sativa to facilitate gene expression analysis and screening imprinted genes.
复制标题
水稻早期核胚乳的分离以利于基因表达分析和筛选印记基因
DOI：
10.1186/s13007-015-0092-4
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发表时间：
2015
期刊：
影响因子：
5.1
通讯作者：
Sun MX
中科院分区：
文献类型：
--
作者：
Kuang Q;Yu X;Peng X;Sun MX
背景：由于水稻生产的品质和产量取决于胚乳的发育，以往的研究主要集中在调节胚乳发育过程的分子机制上。最近，这一过程如何受到表观遗传的调控已成为一个重要的话题。然而，由于早期胚乳的分离是不可能的，因此早期胚乳发育过程中的基因表达分析和印记基因的筛选仍然是具有挑战性的。在这里，我们报道了一种在胚乳发育的游离核阶段，在24或48 HAP(授粉后数小时)分离胚乳的方法。结果：该技术可以快速、方便地收集纯的游离核胚乳。然后，可以使用动态胚乳从分离的胚乳细胞中提取早期胚乳RNA。结果表明，提取的RNA质量较好，可用于早期胚乳的基因表达分析和亲本特异性基因的筛选。结论：为研究早期胚乳发育的分子机制提供了一种可靠的方法。我们的方法可以用于准确的基因表达分析和印记基因的筛选，并有助于在胚乳发育的非常早期阶段确认胚乳特异的基因表达。该方法也可用于其他物种的早期游离核胚乳的采集。
Background:Since the quality and yield of rice production depends on endosperm development, previous studies have focused on the molecular mechanism that regulates this developmental process. Recently, how this process is epigenetically regulated has become an important topic. However, the gene expression analysis and screening imprinted genes during early endosperm development remain challenging since the isolation of early endosperm has not been possible. Here, we report a procedure for the isolation of endosperm at 24 or 48 HAP (hours after pollination) during the free nuclear stage of endosperm development.Results:This technique allows for rapid and convenient collection of pure free nuclear endosperm. Early endosperm RNA can then be extracted from the isolated endosperm cells using dynabeads. Our results showed that the quality of RNA is satisfactory for gene expression analysis and screening the parental-of-origin specific genes in early endosperm.Conclusions:Thus, we offer a reliable method to overcome one of the major obstacles in the investigation of the molecular mechanisms of early endosperm development. Our approach can be used for accurate gene expression analysis and screening of imprinted genes, and facilitates the confirmation of endosperm-specific gene expression at the very early stages of endosperm development. This method could also be used in other species to collect early free nuclear endosperm.