Introduction: MiR-3099 was reported to play a role in neuronal cell differentiation/function in the brain during late embryonic and early neonatal development. To further explore its potential regulatory effects on embryonic brain development, this study aims to construct and validate an expression vector of miR-3099 for future gain-of-function and loss-of-function studies. Methods: pCAG-eGFP vector was modified to include IRES2 and miR-3099 with 150bp upstream and downstream genomic sequences. The newly constructed vector, pCAG-miR-3099-IRES2-eGFP, consists of CAG promoter. The in vitro expression level of miR-3099 was measured using stem-loop RT-qPCR after it was transfected into 293FT cell. Later, the vector was electroporated into the embryonic brain at E15.5. Three days later, the E18.5 embryonic brain was harvested and cryopreserved. Immunohistochemistry was performed by using antibody against eGFP to validate the in utero expression of the transgene in the neocortex of the brain. Results: Our finding showed that, the expression level of miR-3099 was significantly upregulated (p
Introduction: One of the commonly used techniques for mutation screening is High Resolution Melting (HRM) analysis. HRM is a post PCR method that relies on the detection of the fluorescent signals acquired due to the release of DNA intercalated dyes upon the melting of dsDNA to ssDNA. The method is simple, inexpensive and does not require post PCR-handling, making it suitable for high throughput screening. Methods: This study aimed to develop and validate HRM technique for the screening of two disease-associated single nucleotide polymorphisms (SNPs) namely BDNF rs6265 and DAT1 rs40184 using a total of 30 gDNA samples. The obtained results were confirmed and validated by sequencing. Results: HRM analysis showed that the predicted genotypes of BDNF rs6265 and DAT1 rs40184 among all the gDNA samples were in 100% concordance with the sequencing results, making it an accurate and sensitive method for the detection of SNPs. Conclusions: The application of HRM can accurately determine the genotype of BDNF rs6265 and DAT1 rs40184 SNPs, making it a promising tool for rapid and high-throughput screening of targeted SNPs in a large population study.