In more than several papers studing small-RNA/RNA seq I can see for each treatment just one library has been prepared that consists of e.g. 12 pooled biological RNA samples, and no replication applied for the libraries.
However, I wonder if separate replications are not necessary for seq libraries specially where differential expression analysis is the goal of study.
Is one seq library accepted for each treatment???
Hello Zahra,
There are 2 types of replicates that we generally use in transcriptome sequencing libraries, One is called biological replicate where we harvest multiple samples to remove the biological biases another is the technical replicates to remove the technical biases. Although sequencing is an expensive process hence generally researchers performed pooling of samples to overcome biological biases. Although if replicated samples are used then while performing differential Gene Expression, dispersion value is calculated automatically from replicated samples while on performing DGE of samples without replicates you need to assign the dispersion value. Although both type of sequencing libraries are accepted.
Hope this could help you,
Romit
To add somenthing to @Romit, the dispersion assignment for perform DGE without replicates after all ends up being arbitrary, because for sure not all the genes have the same biological dispersion. Therefore, it is even better to pool several samples in each biological replicate to have a better measurement of the dispersion of the gene expression and making better estimations of DGE. Nevertheless, if there are limitations for having more replicates, better to trust in the differential expression of genes with high levels of expression than in the high fold change differences of genes with low counts.
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