Journal: Journal of virological methods

Article Title: Development and evaluation of a ligation-free sequence-independent, single-primer amplification (LF-SISPA) assay for whole genome characterization of viruses

doi: 10.1016/j.jviromet.2021.114346

Figure Lengend Snippet: Stepwise overview of single-indexed library preparation workflow containing Illumina adapters and barcode without ligation for Illumina sequencing (MiSeq). The JS15 primer with 14 random nucleotides at the 3′ end (JS15N14) generates single-stranded and double-stranded DNA, respectively, by reverse transcription and a Klenow reaction. In the first-step PCR primer JS15 randomly amplifies and enriches the sample. In the second step-PCR using a pair of forward and reverse primers encoding Illumina adaptor (MiSeq) and JS15 primer sequences are incorporated into the amplicon from the first round PCR product. The reverse primer carries a unique barcoding index sequence embedded between the Illumina adaptor and JS15 primer sequences. The entire protocol is completed in approximately 1 day.

Article Snippet: Illumina NGS Shotgun metagenomic sequencing was performed on the Illumina MiSeq platform (Illumina, USA).

Techniques: Ligation, Sequencing, Amplification