Our sequencing center has informed us that our RNA quality does not pass their guidelines, and we are not certain how to proceed. The center recommends using RNA with a RIN value of 8 or greater, however I have read that some people suggest never using samples with a RIN below 9.7, while others state that different species are fine down to RIN values of 3 or 4. All eight of our samples fall in a range of values from 5.5 to 6.3, and the profiles are quite similar. I have attached a representative Bioanalyzer profile from a sample with an RIN of 6.3.
Can anyone give me further insight on the quality of these RNA? Does this look like typical bacterial RNA, (Enterobacteriaceae) or is it too far gone? We had gone to great lengths to ensure there were no DNA contamination in the samples, involving multiple rounds of extended DNase treatment.
Additional info:
RNA purification with Qiagen RNeasy Protect Bacteria mini kit
DNase treatment with DNase I amplification grade, cleaned up with Qiagen MinElute RNA kit
Second DNase treatment followed by cleanup and elution into water
RNA destined for Illumina HiSeq 2000 100bp PE sequencing
Can anyone give me further insight on the quality of these RNA? Does this look like typical bacterial RNA, (Enterobacteriaceae) or is it too far gone? We had gone to great lengths to ensure there were no DNA contamination in the samples, involving multiple rounds of extended DNase treatment.
Additional info:
RNA purification with Qiagen RNeasy Protect Bacteria mini kit
DNase treatment with DNase I amplification grade, cleaned up with Qiagen MinElute RNA kit
Second DNase treatment followed by cleanup and elution into water
RNA destined for Illumina HiSeq 2000 100bp PE sequencing
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