Tuyển tập các báo cáo nghiên cứu về y học được đăng trên tạp chí y học Wertheim cung cấp cho các bạn kiến thức về ngành y đề tài: Rapid, low-input, low-bias construction of shotgun fragment libraries by high-density in vitro transposition. | Adey et al. Genome Biology 2010 11 R119 http content 11 12 R119 w Genome Biology METHOD Open Access Rapid low-input low-bias construction of shotgun fragment libraries by high-density in vitro transposition 1Ỷ 2Ỷ 3Ỷ z 3Ỷ 1 1 1 Andrew Adey Hilary G Morrison Asan Xu Xun Jacob O Kitzman Emily H Turner Bethany Stackhouse Alexandra P MacKenzie1 Nicholas C Caruccio4 Xiuqing Zhang3 Jay Shendure1 Abstract We characterize and extend a highly efficient method for constructing shotgun fragment libraries in which transposase catalyzes in vitro DNA fragmentation and adaptor incorporation simultaneously. We apply this method to sequencing a human genome and find that coverage biases are comparable to those of conventional protocols. We also extend its capabilities by developing protocols for sub-nanogram library construction exome capture from 50 ng of input DNA PCR-free and colony PCR library construction and 96-plex sample indexing. Background Massively parallel DNA sequencing methods are rapidly achieving broad adoption by the life sciences research community 1 2 . As the productivity of these platforms continues to grow with hardware and software optimizations the bottleneck experienced by researchers is increasingly at the front end the construction of sequencing libraries and at the back end data analysis and interpretation rather than in the sequencing itself. The input material for commonly used platforms such as the Illumina Genome Analyzer 3 the Roche 454 Genome Sequencer 4 the Life Technologies SOLiD platform 5 as well as for real-time third-generation sequencers such as Pacific Biosciences 6 consists of complex libraries of genome- or transcriptome-derived DNA fragments flanked by platform-specific adaptors. The standard method for constructing such libraries is entirely in vitro and typically includes fragmentation of DNA mechanical or enzymatic end-polishing ligation of adaptor sequences gel-based size-selection and PCR amplification Figure 1a . .
