DNA sequencing is vital for many aspects of biological research and diagnostics. Despite the development of second and third generation sequencing technologies, Sanger sequencing has long been the only choice when required to precisely track each sequenced plasmids or DNA fragments. Here, we report a complete set of novel barcoding and assembling system, Highly-parallel Indexed Tagmentation-reads Assembled Consensus sequencing (HITAC-seq), that could massively sequence and track the identities of each individual sequencing sample. With the cost of much less than that of single read of Sanger sequencing, HITAC-seq can generate high-quality contiguous sequences of up to 10 kilobases or longer. The capability of HITAC-seq was confirmed through large-scale sequencing of thousands of plasmid clones and hundreds of amplicon fragments using approximately 100 pg of input DNAs. Due to its long synthetic length, HITAC-seq was effective in detecting relatively large structural variations, as demonstrated by the identification of a ∼1.3 kb Copia retrotransposon insertion in the upstream of a likely maize domestication gene. Besides being a practical alternative to traditional Sanger sequencing, HITAC-seq is suitable for many high-throughput sequencing and genotyping applications.

DNA 测序对于生物研究和诊断的许多方面都至关重要。尽管第二代和第三代测序技术不断发展，但桑格测序长期以来一直是需要精确追踪每个已测序质粒或DNA片段时的唯一选择。在这里，我们报告了一整套新颖的条形码和组装系统，高度并行索引标签读取组装一致性测序（HITAC-seq），它可以大规模测序并跟踪每个单独测序样本的身份。 HITAC-seq 的成本远低于桑格测序单次读取的成本，可以生成长达 10 KB 或更长的高质量连续序列。 HITAC-seq 的功能通过使用约 100 pg 的输入 DNA 对数千个质粒克隆和数百个扩增子片段进行大规模测序得到了证实。由于其合成长度较长，HITAC-seq 可有效检测相对较大的结构变异，这一点通过在可能的玉米驯化基因上游插入约 1.3 kb Copia逆转录转座子的鉴定得到证明。除了作为传统桑格测序的实用替代方案之外，HITAC-seq 还适用于许多高通量测序和基因分型应用。