Journal ID : CJOHNS-22-04-2024-11439
[This article belongs to Volume - 55, Issue - 04]

Title : Evaluation of minimally invasive oropharyngeal specimens and extraction of nucleic acids for molecular diagnostic analysis

Abstract :

The current study aimed to minimally invasive oropharyngeal specimens and extraction of nucleic acids for molecular diagnostic analysis and find out the benefits, drawbacks, and challenges of developing a non-invasive material sampling technique for molecular downstream analysis that is both consistent and dependable. A more and more crucial clinical tool is molecular diagnostics, particularly for routine sampling. We assessed two non-invasive techniques for extracting nucleic acids from the oral/pharyngeal region: mouthwashes and oral swabs. We developed a procedure from sample collection to investigation. The specimens were subsequently examined for their cellular makeup and the resultant DNA/RNA's purity, deterioration, and microbial concentration. We identified the best housekeeping genes that work with these kinds of samples. Depending on the cellular composition, Mouthwashes had a higher concentration of bacteria and immune cells. Despite the need for more particular optimization in the methodology for bacterial RNA extraction, microbial RNA could be obtained from both sample techniques. We could produce consistent amounts of DNA and RNA by streamlining the method. There was no discernible difference in the RNA or DNA purity metrics between the two sampling techniques. While integrity analysis revealed significant RNA degradation, matching criteria validated their viability for sequencing. TATA-Box Binding Protein was the best housekeeping gene by RT-qPCR analysis. In conclusion, we have created a reliable approach that works with various downstream diagnostic methods.

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