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Methods Mol Biol. 2011;733:257-66. doi: 10.1007/978-1-61779-089-8_18.

Amplification-free library preparation for paired-end Illumina sequencing.

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The Wellcome Trust Sanger Institute, Hinxton, Cambridge, UK.


The library preparation step is of critical importance for the quality of next-generation sequencing data. The use of the polymerase chain reaction (PCR) as a part of the standard Illumina library preparation protocol causes an appreciable proportion of the obtained sequences to be duplicates, making the sequencing run less efficient. Also, amplification introduces biases, particularly for genomes with high or low GC content, which reduces the complexity of the resulting library. To overcome these difficulties, we developed an amplification-free library preparation. By the use of custom adapters, unamplified, ligated samples can hybridize directly to the oligonucleotides on the flowcell surface.

[Indexed for MEDLINE]

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