Rs old from two independent study groups, including 705 obese cases and
Rs old from two independent study groups, including 705 obese cases and 1325 nonobese controls recruited from the urban regions of Beijing, China. The first study group came from the study on Adolescent Lipids, Insulin Resistance, and candidate genes (ALIR). The second study group was from the Comprehensive Prevention project for Overweight and Obese Adolescents (CPOOA). All obese individuals in the selected schools were recruited with their voluntary participation. The method of cluster sampling was adopted to recruit non-obese subjects from some classes of each grade in the same schools. The ALIR subjects were ascertained from adolescents aged 14?7 years in nine middle schools of Dongcheng District of Beijing, including 386 obese adolescents and 551 non-obese adolescents. The CPOOA subjects were recruited from children and adolescents aged 7?8 years old in five elementary and middle schools of the Haidian District of Beijing, comprising 319 obese children and adolescents and 774 non-obese children and adolescents. The ascertainment strategies for the two study groups have been described in detail previously [15, 16]. We used the uniform BMI percentile criteria for obese and non-obese children, which were determined in a representative Chinese population [17]. According to the criteria, the children and adolescents with an age- and gender-specific BMI 95th percentile are defined as obese, whereas those with a BMI between 15th and 95th percentile are nonobese. The individuals with any cardiovascular or metabolic disease were excluded. Anthropometric measurements, including height and weight, were measured at school according to standard protocols. Fasting venous blood samples were taken for GS-9620 web detection of ALT. Methylation data were collected from 110 severely obese children and 110 normal-weight age- and gendermatched controls, which were chosen from the CPOOA study. We chose those with an age- and gender-specific BMI 97th percentile as the severely obese cases, and those with BMI between 15th and 85th percentile as non-obese controls. We point the reader to our prior work, where we have described the study design for the methylation detection [14]. Studies were approved by the Ethic committee of Peking University Health Science Center. Written informed consent was provided by all participants and, in PubMed ID:https://www.ncbi.nlm.nih.gov/pubmed/26266977 the case of minors, by their parents.Wang et al. BMC Medical Genetics (2017) 18:Page 3 ofSNP genotypingThe HIF3A rs3826795 polymorphism was genotyped using genomic DNAs extracted from blood leukocytes by the phenol-chloroform extraction method. Genotyping was conducted on MassARRAY System (Sequenom, San Diego, CA, USA). Primers, including a pair of amplification primers and an extension primer, were designed with Sequenom MassArray Assay Design Suite. A multiplex polymerase chain reaction was performed, and unincorporated double stranded nucleotide triphosphate bases were dephosphorylated with shrimp alkaline phosphatase followed by primer extension. The purified primer extension reaction was spotted on to a 384-element silicon chip (SpectroCHIP, Sequenom) and analyzed in the Matrix assisted laser desorption ionization time of flight mass Spectrometry (MALDI-TOF MS, Sequenom). The resulting spectra were processed with MassArray Typer (Sequenom) (http://www.sequenom.com). The genotyping call rate of the HIF3A rs3826795 polymorphism was 97.9 . All the experiments were done by investigators who were blind to the phenotypes.DNA methylation det.