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Sample preparation for Illumina-based mRNA sequencing was conducted based upon the manufacturer's protocols "mRNA Sequencing Sample Preparation Guide" and "Preparing Samples for Multiplexed Paired-End Sequencing" (Part# 1005361Rev B, and Part# 1004898Rev D, Illumina, Inc., San Diego, CA).
The Illumina TruSeq™ RNA Sample Preparation Guide was followed to prepare the samples for sequencing.
Libraries were prepared following the TruSeq RNA Sample Preparation Guide (Illumina).
Paired-end libraries were prepared according to the TruSeq™ RNA Sample Preparation Guide (Illumina).
Illumina libraries (ab, bc, and ac) were prepared according to the Illumina paired-end sample preparation guide with the following modifications.
The manufacturer's mRNA sequencing sample preparation guide (Illumina, Inc., San Diego, CA) was used to prepare two pools of total RNA for paired-end sequencing (101 bp read length).
The workflow in the Nextera Enrichment Sample Preparation Guide (Revision B) by Illumina was followed to prepare the libraries for whole-exome sequencing.
The library was prepared as recommended by Illumina (MiSeq Reagent Kit v2 – Reagent Preparation Guide) and was loaded at 7.5 pM on one MiSeq flowcell with 10% phiX spiked in.
Total RNA samples were prepared for smRNA sequencing using Illumina's Small RNA v1.5 Sample Preparation Guide.
Illumina RNA-seq libraries from three biological replicates of each tissue were prepared using the TruSeq mRNA sample kit (Illumina TruSeq RNA sample preparation guide Part 15008136 Rev A, Nov 2010) using 4 μg total RNA.
250 and 500 bp paired-end libraries were prepared using enzymes from NEBNext Modules and following Illumina TruSeq DNA sample preparation guide.
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