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

ABSTRACT: Molecular identification and characterization of novel or re-emerging infectious pathogens is critical for disease surveillance and outbreak investigations. Next generation sequencing (NGS) using Sequence-Independent, Single-Primer Amplification (SISPA) is being used extensively in sequencing of viral genomes but it requires an expensive library preparation step. We developed a simple, low-cost method that enriches nucleic acids followed by a ligation-free (LF) 2-step Polymerase Chain Reaction (PCR) procedure for library preparation. A pan-chimeric universal primer (JS15N14) containing 15 nucleotides with a random tetradecamer (14N) attached to the 3'-end was designed. The complimentary primer (JS15) was used for nucleic acid enrichment in a first round PCR. A second PCR was designed to cr

SUBMITTER: Jothikumar N 

PROVIDER: S-EPMC11075090 | biostudies-literature | 2022 Jan

REPOSITORIES: biostudies-literature