What exactly one "library" corresponds to in RNA-seq analysis
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5.7 years ago
tunl ▴ 70

I understand that we need to do library preparation for Illumina in RNA-seq.

I am just not sure what exactly one “library” corresponds to. For example, in the fastq files I received, I have two conditions and each condition has 4 replicates. Does this mean I have 4 libraries for each condition so totally I have 8 libraries?

RNA-Seq • 1.8k views
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One sample should lead to one library (unless more than one library is constructed for that sample, which would then be a technical replicate(s)).

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Thank you so much!

So you mean, each biological replicate leads to one library, right?

So in our case (two conditions and each condition has 4 replicates), we have totally 8 libraries.

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As long as those are true biological replicates you will have 8 libraries.

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Thanks a lot!

On the metadata table I received, for each sample there are two tubes. Does this mean there are two technical replicates for each sample? If so, there would be 2 libraries for each sample, right?

Also, for each tube, there are 3 lanes. Does number of lanes have anything to do with number of libraries?

Thank you so much for the help!

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I am not sure what to make of

for each sample there are two tubes

that is a detail you would have to clarify with the submitter's. The tubes may represent one biological sample split in two tubes (for archival/other reasons) or something else.

A library, once made, can be run on multiple lanes (or flowcells) to collect as many reads as needed for the experiment. You should be able to combine all data for one library into one alignment file. You can do it post-alignment or before by cat'ing the fastq files before alignment.