Probably you assigned sizes first and then did the estimate, which may just scale your assigned sizes back to 1. I think the right way is to estimate the factor directly from the 'cds' object without assigning any libsizes.

The author of DESeq wrote a post on SeqAnswers a while back about how the size factors thing work in DESeq. I'll try to find the post, but basically it normalizes the datasets by:

-Take the geometric mean of each condition for a gene and use that as the reference expression data set.

-For each condition, get a list of quotients of each gene expression value to its reference expression.

-The median of each condition quotient list is the normalization factor for that data set.

Probably you assigned sizes first and then did the estimate, which may just scale your assigned sizes back to 1. I think the right way is to estimate the factor directly from the 'cds' object without assigning any libsizes.

Hi Jeremy thanks for your comment could u please tell me what did this sizeFactors function? is it for normalization of data (scale normalization)?