TruSeq DNA PCR Free library preparation input

Why are different input amounts used to generate 350 bp and 550 bp PCR-Free libraries? The input recommendations were tested and validated to yield appropriate final yields for sequencing. Illumina recommends 1 µg input for the 350 bp target insert size and 2 µg for the 550 bp target insert size. In order to generate sufficient nanomolar concentration of libraries for sequencing with the larger 550 bp insert length, 2 μg of input material is required, due to the longer insert size compared to the 350 bp workflow.

Can the input DNA quantity be reduced? Illumina does not support or recommend input amounts lower than 1 μg for a 350 bp insert size or 2 μg for a 550 bp library. It is possible that reducing the amount of input material may fail to generate enough library to successfully proceed with sequencing.

For any feedback or questions regarding this article (Illumina Knowledge Article #1182), contact Illumina Technical Support [email protected].