RNA sequencing Rat - Hippocampal tissue

RNA-Seq is a method to sequence RNA by applying Next Generation Sequencing (NGS). The quality of RNA is critical for the success of RNA-Seq. The integrity of RNA is measured by the RNA integrity number (RIN). RIN is computed from RNA electrophoresis and electropherogram profiles (the peak area of the 28S rRNA should be approximately twice the peak area of the 18S rRNA). If you get the RIN value lower than 7, the possibility of getting the low quality of RNA-seq data is high. To get a high quality RNA, it is better to work with fresh samples or snap-freeze the tissues in liquid nitrogen as quickly as possible and store them at -80°C until further use. Make sure designated areas and all your filter tips, microfuge tubes, plastic, and glassware are RNase-free.

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Protocol tips
Libraries were created from high quality RNA (RNA integrity number of 7 or greater) using the SENSE mRNA-Seq Library Prep Kit from Lexogen (Lexogen, Vienna, Austria) according to the manufacturer’s protocol. This protocol includes an integrated poly(A) mRNA selection process, and the constructed library maintains the strand specificity. One single RNA-Seq library was constructed for each RNA sample. Library production is initiated by the random hybridization of Starter/Stopper heterodimers to the RNA template, followed by first and second strand cDNA synthesis.
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