Illumina sequencing library preparation for highly multiplexed target capture and sequencing.
Illumina sequencing library preparation for highly multiplexed target capture and sequencing.
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DOI：
10.1101/pdb.prot5448
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发表时间：
2010-06-01
影响因子：
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通讯作者：
Kircher, Martin
中科院分区：
文献类型：
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作者：
Meyer, Matthias;Kircher, Martin
通过高通量测序技术产生的大量DNA序列数据通常允许在单个测序运行中并行地对多个样品进行测序。如果研究的是基因组的子集而不是完整的基因组，这一点尤其正确。近年来，从测序文库中捕获靶标已在很大程度上取代聚合酶链反应（PCR）作为靶标富集的优选方法。使多个样品的靶捕获和测序同步化需要将样品特异性条形码并入测序文库中，这对于追溯每个序列的样品来源是必要的。该方案描述了一种快速可靠的方法，用于为Illumina的基因组分析仪平台制备条形码（“索引”）测序文库。该方案避免了昂贵的商业文库制备试剂盒，并且可以使用多通道移液器在96孔板设置中进行，需要不超过两天或三天的实验室工作。文库可以从任何类型的双链DNA制备，即使以亚纳克的量存在。
The large amount of DNA sequence data generated by high-throughput sequencing technologies often allows multiple samples to be sequenced in parallel on a single sequencing run. This is particularly true if subsets of the genome are studied rather than complete genomes. In recent years, target capture from sequencing libraries has largely replaced polymerase chain reaction (PCR) as the preferred method of target enrichment. Parallelizing target capture and sequencing for multiple samples requires the incorporation of sample-specific barcodes into sequencing libraries, which is necessary to trace back the sample source of each sequence. This protocol describes a fast and reliable method for the preparation of barcoded ("indexed") sequencing libraries for Illumina's Genome Analyzer platform. The protocol avoids expensive commercial library preparation kits and can be performed in a 96-well plate setup using multi-channel pipettes, requiring not more than two or three days of lab work. Libraries can be prepared from any type of double-stranded DNA, even if present in subnanogram quantity.