Lambertini Luca, Diplas Andreas I, Lee Men-Jean, Sperling Rhoda, Chen Jia, Wetmur James
Department of Community and Preventive Medicine, Mount Sinai School of Medicine, New York, New York 10029, USA.
Epigenetics. 2008 Sep;3(5):261-9. doi: 10.4161/epi.3.5.6755. Epub 2008 Sep 7.
Loss of imprinting (LOI) is the gain of expression from the silent allele of an imprinted gene normally expressed from only one parental copy. LOI has been associated with neurodevelopmental disorders and reproductive abnormalities. The mechanisms of imprinting are varied, with DNA methylation representing only one. We have developed a functional transcriptional assay for LOI that is not limited to a single mechanism of imprinting. The method employs allele-specific PCR analysis of RT-PCR products containing common readout polymorphisms. With this method, we are able to measure LOI at the sensitivity of 1%. The method has been applied to measurement of LOI in human placentas. We found that RNA was stable in placentas stored for more than one hour at 4 degrees C following delivery. We analyzed a test panel of 26 genes known to be imprinted in the human genome. We found that 18 genes were expressed in placenta. Fourteen of the 18 expressed genes contained common readout polymorphisms in the transcripts with a minor allele frequency >20%. We found that 5 of the 14 genes were not imprinted in placenta. Using the remaining nine genes, we examined 93 heterozygosities in 27 samples. The range of LOI was 0%-96%. Among the 93 heterozygosities, we found 23 examples (25%) had LOI >3% and eight examples (9%) had LOI 1-3%. Our results indicate that LOI is common in human placentas. Because LOI in placenta is common, it may be an important new biomarker for influences on prenatal epigenetics.
印记丢失（LOI）是指印记基因的沉默等位基因获得表达，该基因通常仅从一个亲本拷贝表达。LOI与神经发育障碍和生殖异常有关。印记机制多种多样，DNA甲基化只是其中之一。我们开发了一种用于LOI的功能性转录测定法，该方法不限于单一的印记机制。该方法采用对含有常见读出多态性的逆转录聚合酶链反应（RT-PCR）产物进行等位基因特异性PCR分析。通过这种方法，我们能够以1%的灵敏度测量LOI。该方法已应用于人类胎盘LOI的测量。我们发现，分娩后在4摄氏度下储存超过一小时的胎盘中RNA是稳定的。我们分析了一组已知在人类基因组中被印记的26个基因的测试面板。我们发现18个基因在胎盘中表达。18个表达基因中的14个在转录本中含有常见读出多态性，次要等位基因频率>20%。我们发现14个基因中的5个在胎盘中没有印记。使用其余9个基因，我们检查了27个样本中的93个杂合性。LOI的范围为0%-96%。在93个杂合性中，我们发现23个例子（25%）的LOI>3%，8个例子（9%）的LOI为1%-3%。我们的结果表明，LOI在人类胎盘中很常见。由于胎盘中的LOI很常见，它可能是影响产前表观遗传学的一个重要新生物标志物。
