亚硫酸氢钠测序法检测水稻FIE基因CpG岛甲基化状态

建立了适用于水稻基因组特定基因甲基化检测的亚硫酸氢钠测序法,并利用此方法对FIE2A基因CpG岛部分片段的甲基化差异进行了研究。采用CTAB法提取水稻叶片和胚乳细胞的基因组DNA,经亚硫酸氢钠化学修饰后,针对已修饰的FIE基因序列设计特异引物并结合巢式PCR扩增,TA载体克隆、测序,最后对测序结果进行分析。结果表明巢式PCR能够增加特异性产物的产生,FIE基因CpG岛在对称的CG和CNG位点甲基化水平较高,而在非对称CNN位点甲基化水平最低,此外在叶片中的平均甲基化水平较高。由此表明本研究建立的亚硫酸氢钠测序法适用于水稻基因组特定基因甲基化状态的检测。

Analysis of Methylation at CpG Island of FIE in Rice by Bisulfite Sequencing

To test the feasibility of bisulfite sequencing in methylation analysis at a specific gene of rice,we analyzed DNA methylation at fragment of CpG island of FIE2A.In the present paper,genomic DNA was extracted from leaf and endosperm of rice with the CTAB method.After treated with bisulfite sodium,we amplified the CpG island of FIE gene with designed primers using nest-PCR,sequencing results after cloning in TA-vector.Results showed that nest-PCR increased the specificity of the PCR products,and symmetric se...