Fig. 1

Methodology for SARS-CoV-2 Genome Sequencing Using Oxford Nanopore Technologies: This figure illustrates the detailed methodology employed in this study for SARS-CoV-2 genome sequencing, encompassing cohort selection, sample preparation, PCR amplification, barcoding, and sequencing¹³. (A) Cohort Selection: The study cohort was divided into two groups: 24 healthy individuals and 24 COVID-19 patients, ensuring equal representation for comparative analysis. (B) Sample Preparation: Nasopharyngeal swabs were collected from participants, followed by RNA extraction. The extracted RNA was then converted to complementary DNA (cDNA) via reverse transcription, preparing the samples for further amplification and sequencing. (C) PCR and Barcoding: The SARS-CoV-2 genome was amplified using 98 pairs of primers in a multiplex PCR, yielding a 29.8 kb amplified product. Subsequent library preparation involved the use of a rapid barcoding kit and the transposome complex to ensure efficient barcoding of the amplified products. (D) Sequencing: The prepared libraries were pooled and loaded onto the MinION sequencer. Sequencing was conducted using nanopore technology, followed by base calling, and the assembly and analysis of the sequenced genome, which provided detailed phylodynamic insights.