Freeze-Thaw Reduces Sensitivity for Targeted Sequencing on Low Viral Load Samples

摘要

Retrieving viral sequences from direct primary specimens by next-generation sequencing has been challenging, especially when peak viremia of particular viruses is low or samples were collected at the tailed-end of the viremia. During a study of viral diversity of Zika virus (ZIKV) in challenged non-human primate model, we observed that freshness of extracted RNA is important for efficiency in targeted-sequencing after amplification by Fluidigm Access Array (AA). Our hypothesis is that the integrity of the viral RNA may be reduced or lost in the course of freezing the samples after extraction. This could result in further reduction in sequence depth or coverage across the genome due to lower efficiency in amplicon synthesis during targeted sequencing.