Aims: The application of mouthwash is one of the oral hygiene treatments that commonly use after tooth brushing to control the bacterial colonization from overgrowth. This research is focused on investigating the effect of mouthwash on oral microbiome by analyzing the quality and yield of DNA obtained before and after using mouthwash and also to compare the bacterial abundance via 16S rRNA PCR detection. Methodology and results: The DNA was extracted from the saliva samples before and after using mouthwash using Phenol-Chloroform extraction method. The DNA extract was then evaluated using Nano Drop ND-1000 UV/VIS Spectrophotometer to determine the DNA quality and DNA yield. After that, the 16S rRNA gene was amplified via PCR for bacterial detection in the saliva using 27 F and 1492 R primers set, and the PCR products were observed on 1.5% gel electrophoresis. Statistical analysis was performed by using Graphpad Prism 7.03 software. For DNA yield, there was significantly higher yield observed after mouthwash usage with 80% of the samples was found to yield more DNA. To assess DNA quality, absorbance ratio of A260/A280 and A260/A230 were used. The DNA quality was seen to be similar for both A260/A280 and A260/A230 absorbance ratio even after the usage of mouthwash. The amplification of 16S rRNA gene was successful and 1500 bp expected band size was observed. Conclusion, significance and impact of study This study demonstrated the usage of mouthwash is useful to increase the DNA yield as compared to without using mouthwash. However in terms of quality, no difference is seen. This result can be used to provide insight on mouthwash usage for saliva sampling in a non-invasive manner.