I have some questions about rlog transformation using DESeq2:

1) This transformation normalizes for library size, which is sequencing depth or total number of mapped reads, correct?

2) This transformation does not normalize for gene or transcript length, correct? If this is true, is there a program you would recommend to normalize transcript length using the output from rlog transformation?

Thanks!

By default, it corrects for library size using not total number of reads but instead a robust estimate of library size (see original DESeq paper which is listed as a citation from ?estimateSizeFactors).

If you use the tximport pipeline, the transformations will also normalize for differences in average transcript length across samples. Note that this is correcting for differences in transcript length across samples but not normalizing wrt transcript length across rows (genes). You could do this yourself by then dividing each row by the average transcript length.