“Deep” Sequencing Accuracy and Reproducibility Using Roche/454 Technology for Inferring Co-Receptor Usage in HIV-1

Next generation, “deep”, sequencing has increasing applications both clinically and in disparate fields of research. This study investigates the accuracy and reproducibility of “deep” sequencing as applied to co-receptor prediction using the V3 loop of Human Immunodeficiency Virus-1. Despite increasing use in HIV co-receptor prediction, the accuracy and reproducibility of deep sequencing technology, and the factors which can affect it, have received only a limited level of investigation. To accomplish this, repeated deep sequencing results were generated using the Roche GS-FLX (454) from a number of sources including a non-homogeneous clinical sample (N = 47 replicates over 18 deep sequencing runs), and a large clinical cohort from the MOTIVATE and A400129 studies (N = 1521). For repeated measurements of a non-homogeneous clinical sample, increasing input copy number both decreased variance in the measured proportion of non-R5 using virus (p新一代“深度”测序（deep sequencing）技术在临床及各研究领域中的应用持续拓展。本研究针对基于人类免疫缺陷病毒1型（Human Immunodeficiency Virus-1, HIV-1）V3环（V3 loop）的HIV共受体预测（co-receptor prediction）场景，探究深度测序技术的准确性与可重复性。尽管深度测序在HIV共受体预测中的应用日益广泛，但当前学界对该技术的准确性、可重复性及其影响因素的研究仍较为有限。为达成研究目标，我们采用罗氏GS-FLX（454）测序平台生成了多来源的重复深度测序数据：其一为一份非均相临床样本（在18次深度测序运行中完成47次重复检测），其二为来自MOTIVATE与A400129研究的大型临床队列（样本量N=1521）。针对非均相临床样本的重复检测结果显示，输入拷贝数的增加可降低非R5型病毒检测占比的方差（p