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Open Access Research article

Screening significantly hypermethylated genes in fetal tissues compared with maternal blood using a methylated-CpG island recovery assay-based microarray

Aihua Yin12, Xiangzhong Zhang3, Jing Wu1, Li Du1, Tianwen He1 and Xiaozhuang Zhang2*

Author Affiliations

1 Prenatal Diagnosis Centre, Guangdong Women and Children Hospital, Guangzhou, Guangdong, 510010, China

2 Maternal and Children Metabolic-Genetic Key Laboratory, Guangdong Women and Children Hospital, Guangzhou, Guangdong, 510010, China

3 Department of Hematology, The Third Affiliated Hospital of Sun Yat-sen University, Guangzhou, Guangdong, 510630, China

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BMC Medical Genomics 2012, 5:26  doi:10.1186/1755-8794-5-26

Published: 18 June 2012

Abstract

Background

The noninvasive prenatal diagnosis procedures that are currently used to detect genetic diseases do not achieve desirable levels of sensitivity and specificity. Recently, fetal methylated DNA biomarkers in maternal peripheral blood have been explored for the noninvasive prenatal detection of genetic disorders. However, such efforts have covered only chromosomal aneuploidy, and fetal methylated DNA biomarkers in maternal whole blood for detecting single-gene diseases remain to be discovered.

Methods

To address this issue, we systematically screened significantly hypermethylated genes in fetal tissues and compared them with maternal peripheral blood potential in an attempt to detect fetal genes in maternal peripheral blood. First, the methylated-CpG island recovery assay combined with a CpG island array was performed for four fetus-toward placental tissues and the corresponding maternal peripheral bloods. Subsequently, direct bisulfite sequencing and combined bisulfite restriction analysis (COBRA) were carried out to validate the methylation status of the hypermethylated genes that were identified by the microarray analysis.

Results

Three hundred and ten significantly hypermethylated genes in the placental tissues were detected by microarray. From the top 15 hypermethylated genes detected by microarray, two were selected for sequencing validation in placental tissue and chorionic villus samples and four were selected for COBRA validation in four placental tissues, ten amniotic fluids and five chorionic villus samples. The six selected genes were confirmed to be hypermethylated in placental tissue and chorionic villus samples, but methylation of the genes could not be detected in the amniotic fluids.

Conclusions

Of the many hypermethylated genes and methylation sites that were found in the fetal tissues, some have great potential to be developed into molecular markers for noninvasive prenatal diagnosis of monogenic disorders. Further clinical studies are warranted to confirm these findings.

Keywords:
CpG islands methylation; Methyl-CpG binding protein; Microarray; Combined bisulfite restriction analysis