Hi All, This is my first time posting and have more experience with the wet-lab part of things, so please let me know how I can improve this post. I have checked the forum archives for an answer, but have not found it.
We are trying to sequence cell-specific RNA from mouse spleen and compare to whole spleen. Because these samples are low amount (50 ng total) and quality (RIN 3.5), we sent one of each (cell-specific and whole spleen) for a pilot library construction and a 150 bp, paired end MiSeq run . We do not expect the quality of the data to be great, but we are trying to determine if it is good enough for deeper sequencing. FastQC showed decent read quality, but a really high GC content throughout (60-80%), which is exacerbated at the beginning of the read. There is particular depletion of T throughout. There are also several overrepresented sequences and only 45% overall read mapping rate on Tophat. My questions are: 1. What is the likely cause of the high GC content? 2. Is it worth sequencing deeper?