Hi Ege,

The only real difference between these three sequencing platforms is the scale or depth of sequencing data that they can generate (MiSeq < NextSeq < HiSeq). I'd recommend checking out Illumina's datasheets on the different platforms to give you an idea. https://emea.illumina.com/systems/sequencing-platforms/comparison-tool.html

I don't have any experience with miRNA datasets, so can't give you any advice on depth - this is completely dependent upon the application. So if you can figure out how many reads you might need across all of your samples, this will tell you whether you can use the MiSeq or whether you need a higher throughput platform.

Cheers,

Hamish

Hi Ege,

I used MiSeq for mRNA-Seq exploratory applications only since the depth limit is 30 RPKM per sample, reason why MiSeq is as Hamish mentioned not capable to reach the required depth. Although, for miRNA my understanding is that ~6 RPKM per sample are enough for publishing and that MiSeq is especially recommended for such purposes. My advice would be for you to check a publication performing a similar kind of sequencing experiment and base your design on it.

This one might be a relevant:

https://doi.org/10.1371/journal.pone.0034150

If I'm not mistaken they aimed for 5RPKM.

Best,

Pablo

Hi I agree with Hamish and Pablo, MiSeq for smallRNA sequencing is not the best choice in terms of cost/benefit. Consider another thing too: one of the main advantages of MiSeq is its long-read output (2x300), but this will be a waste in your case, given the dimension of the RNA you're looking for. Clearly you could choose shorter read length output, but then it would be wiser and more economically convenient, especially if you are planning to do a comprehensive experiment, to go on bigger machines that allow you to sequence multiple libraries in one run.

Thank you everyone for answering, from what I have read and found out was that people have done and are still doing miRNA screening with the MiSeq platform. What I understand is that by doing the sequencing experiment with MiSeq you are decreasing the number of miRNAs you have due to the reads per sample.

For instance in this study:

Article Biomarker discovery: Quantification of microRNAs and other s...

They show with even with HiSeq 2500 they get 11.6 M reads per library while the MiSeq platform generates 890 thousand reads per library. Which is not ideal. The same paper also indicates that by cutting the RPM in half from 12M to 6M the identified miRNA's decrease by 20%. Therefore maybe the real quesitons should be;

1) As Pablo suggested 6M reads might be sufficient to get a good result however the more reads the better according to this paper, therefore is it possible to get 6M reads per sample in MiSeq? If so how many can samples can I pool in one lane?

2) I do not know what "long read ouput (2x300)" means that Marco mentioned could anyone elaborate?

Thank you for your assitance everyone I have literally no experience in this field so any help is much appreciated.