Highly sensitive and reliable human sex determination using multiplex PCR

Gender validation is indispensable in data verification for demographic information particularly in large-scale population studies and also as part of the quality assurance of biospecimen repositories. Gender validation is also critical as part of quality control processes before specimens are subjected to sequencing analysis to identify germline and somatic mutations. The SRY gene is considered a useful signature gene marker that differentiates male from female. The aim of the study was to validate the SRY gene marker for use in gender determination in large cohort studies. SRY gene-specific sequences were amplified by PCRCR, electrophoresed on agarose gels and the 254 bp band was visualized for male samples. A series of DN A template concentrations were tested for sensitivity determination. The reaction was validated on 48 gender-blinded samples obtained from the UMBI BioBank to determine the specificity. ATL1 gene-specific sequence on X chromosome was used as the internal control. This PCR method has demonstrated 100% gender specificity. The sensitivity of the reaction was demonstrated with as low as 0.1 ng male DN A. The findings had suggested that SRY analysis by Multiplex PCR is a highly sensitive and specific method for gender determination and can be extremely useful for large-scale samples

Exome sequencing identifies SLC26A4, GJB2, SCARB2 and DUOX2 mutations in 2 siblings with Pendred syndrome in a Malaysian family

List of PCR primers used. (XLSX 9 kb

First reported case of haemoglobin-M Hyde Park in a Malay family living in Malaysia

Haemoglobin (Hb)-M Hyde Park, also known as Hb-M Akita is a rare type of hereditary Hb M due to autosomal dominant mutation of CAC>TAC on codon 92 of β globin gene resulting in the replacement of histidine by tyrosine on β globin chain. This variant Hb has a tendency to form methaemoglobin (metHb). The iron ion in metHb is oxidized to ferric (Fe3+) which is unable to carry oxygen and the patients manifest as cyanosis clinically. A 9-year-old Malay girl was incidentally found to be cyanotic when she presented to a health clinic. Laboratory investigations revealed raised methaemoglobin levels and Hb analysis findings were consistent with Hb-M Hyde Park. β gene sequencing confirmed a point mutation of CAC>TAC on codon 92 in one of the β genes. The family study done on the individuals with cyanosis showed similar findings. A diagnosis of heterozygous Hb-M Hyde Park was made. Patients with this variant Hb usually presented with cyanosis with mild haemolysis and maybe misdiagnosed as congenital heart disease. No further treatment is needed as patients are relatively asymptomatic. Although the disease is harmless in the heterozygous carriers but the offspring of the carriers may suffer severe haemolytic anaemia when the offspring also inherit other β haemoglobinopathies/thalassemia. This can happen due to high prevalence of β thalassemia carrier (3.5-4 %) found in Malaysia. At the time of writing, this is the first case of hereditary Hb-M Hyde Park diagnosed in a Malay family living in Malaysia

Body composition and risk factors for cardiovascular disease in global multi-ethnic populations

Background: No large-scale studies have compared associations between body composition and cardiovascular risk factors across multi-ethnic populations. Methods: Population-based surveys included 30,721 Malay, 10,865 Indian and 25,296 Chinese adults from The Malaysian Cohort, and 413,737 White adults from UK Biobank. Sex-specific linear regression models estimated associations of anthropometry and body composition (body mass index [BMI], waist circumference [WC], fat mass, appendicular lean mass) with systolic blood pressure (SBP), low-density lipoprotein cholesterol (LDL-C), triglycerides and HbA1c. Results: Compared to Malay and Indian participants, Chinese adults had lower BMI and fat mass while White participants were taller with more appendicular lean mass. For BMI and fat mass, positive associations with SBP and HbA1c were strongest among the Chinese and Malay and weaker in White participants. Associations with triglycerides were considerably weaker in those of Indian ethnicity (eg 0.09 [0.02] mmol/L per 5 kg/m2 BMI in men, vs 0.38 [0.02] in Chinese). For appendicular lean mass, there were weak associations among men; but stronger positive associations with SBP, triglycerides, and HbA1c, and inverse associations with LDL-C, among Malay and Indian women. Associations between WC and risk factors were generally strongest in Chinese and weakest in Indian ethnicities, although this pattern was reversed for HbA1c. Conclusion: There were distinct patterns of adiposity and body composition and cardiovascular risk factors across ethnic groups. We need to better understand the mechanisms relating body composition with cardiovascular risk to attenuate the increasing global burden of obesity-related disease

mIR-99a-5p and mIR-148a-3p as candidate molecular biomarkers for the survival of lung cancer patients

MicroRNA (miRNA) has emerged as a promising biomarker for improving the current state of an early lung cancer diagnosis. Multiple studies have reported that circulating miRNAs are usually combined in a single panel to determine lung cancer risk. In this study, we sought to assess the prognostic predictive values of the potential miRNAs for lung cancer survival among Malaysian patients. The microarray analysis was performed on the isolated miRNA samples of formalin-fixed lung cancer tissues from Malaysian populations. The correlation between miRNA expression and lung adenocarcinoma (LUAD) patient survival was predicted using TGGA data, followed by extensive in silico analyses, including miRNA target gene identification, protein-protein interaction (PPI) network construction, subnetwork (SN) detection, functional enrichment analysis, gene-disease associations, and survival analysis in advanced-stage LUAD. Overall, two promising miR-99a-5p and miR-148a-3p were upregulated in the patients with good survival. We found that 64 miR-99a-5p and 95 miR-148a-3p target genes were associated with poor prognosis and highly participated in cancer-associated processes, such as apoptosis, mRNA transport and cell-cell adhesion. The density score of 4.667, 3.333, and 3.000 in respective SN1, SN2, and SN3 showed the significant subnetworks of constructed PPI leading to the identification of 17 targets, of which ~79% of them involved in neoplastic diseases. Four high-confidence target genes (SUDS3, TOMM22, KPNA4, and HMGB1) were associated with worse overall survival in LUAD patients, implying their critical roles in LUAD pathogenesis. These findings shed additional light on the roles of miR-99a-5p and miR-148a-3p as potential biomarkers for LUAD survival

Kaedah diagnostik semasa dan penggunaan ujian Titik Penjagaan Pantas (POC) bagi mendiagnos Hiperkolesterolemia Famili (FH)

Hiperkolesterolemia Famili (FH) ialah penyakit genetik yang diwarisi secara autosomal dominan dan dicirikan melalui peningkatan kepekatan plasma kolesterol lipoprotein berketumpatan rendah (LDL-C) di dalam darah. Pesakit FH yang tidak didiagnosis di peringkat awal dan tidak dirawat boleh meningkatkan risiko penyakit jantung koronari pramatang. Dengan kepesatan teknologi dalam bidang biologi molekul, terdapat pelbagai strategi telah diambil untuk membolehkan diagnosis awal FH dilakukan. Teknik-teknik ini dapat meningkatkan keberkesanan kos dan tempoh masa pengesanan adalah lebih cepat. Kaedah diagnostik semasa yang sedia ada untuk mendiagnosis FH yang melibatkan kriteria skor berasaskan algoritma dan pelbagai kaedah diagnosis molekul seperti kaedah penjujukan generasi kedua (NGS), penjujukan Sanger, multiplex ligation-dependent probe amplification (MLPA) dan mikroatur melalui hibridisasi DNA akan dibincangkan di dalam ulasan ini. Walau bagaimanapun, ujian genetik molekul ini tidak tersedia secara meluas atas sebab seperti prosedur yang memakan masa, kos yang tinggi dan keperluan kepada kakitangan terlatih. Oleh itu, ulasan ini memberi penekanan kepada penggunaan ujian titik penjagaan pantas (point of care, POC) sebagai pendekatan untuk mendiagnosis FH kerana ketiadaan ujian genetik bagi pemeriksaan rutin di negara yang kekurangan infrastruktur dan kepakaran. Ujian aliran lateral (LFA) telah mendapat perhatian sebagai kaedah diagnostik POC kerana ianya mudah, memerlukan kos yang rendah dan proses yang lebih cepat. Kelebihan ini menjadikan LFA sebagai teknik yang berpotensi dalam menangani beberapa cabaran bagi diagnosis FH khususnya bagi diagnosis awal terhadap keluarga pesakit

Mutations in KIF27, GNAS and IFT140 genes in a patient with VACTERL association: a case report

VACTERL association is a rare genetic disorder involving at least three of the following congenital malformations: vertebral defects (V), anal atresia (A), cardiac defects (C), trachea-oesophageal fistula with or without oesophageal atresia (TE), renal anomalies (R) and limb abnormalities (L). Until now, the aetiology of VACTERL association is unknown, particularly at the molecular level. Here, we performed whole exome sequencing (WES) of an infant with VACTERL association. The patient was delivered prematurely at 30 weeks and had 4/6 of the VACTERL malformations. Trio-WES analysis was performed using Torrent Suite and ANNOVAR. Polymorphisms with an allele frequency of >0.01 were excluded, and the remaining variants were filtered based on de novo mutations, autosomal recessive, X-linked and di-genic inheritance traits. In this patient, no homozygous, compound heterozygous or X-linked mutations was associated with VACTERL. However, we identified two heterozygous mutations; KIF27 (ENST00000297814: c.3004A> C:p.N1002H) and GNAS (ENST00000371098: c.205C>A:p.H69N) genes that were inherited from her father and mother respectively. A de novo, IFT140 gene mutation (ENST00000426508: c.683C>G:p.S228C) was also identified in this patient. The VACTERL phenotype in this patient may due to heterozygous mutations affecting KIF27 and GNAS genes, inherited via autosomal recessive trait. In addition, the IFT140 gene mutation may also be involved. These genes are known to be directly or non-directly involved in the sonic hedgehog signalling that is known to be implicated in VACTERL. This is the first report of these genetic mutations in association with VACTERL