I've worked with tolerance to drought in sugarcane in Brazil and recently I came to USA to perform some RNA-seq at the University of Illinois. I got 4 cDNAs libraries from two contrasting genotypes, one very tolerant and the other very sensitive, in two conditions each: irrigated control and severe drought. We had paired-end sequencing for the 4 RNAseq libraries, using Illima. The yield of these libraries was high at over 13 million reads per library. The average quality scores of all bases in the first and second read were above 20, the error rate of this run was <0.8%.
I want to find differentially expressed genes between the contrasting genotypes, so I've used novoalign to do the alignment against the sorghum genome (gene models) and now I'm trying to find a way to normalize my data and calculate the differentially expressed genes between the two genotypes and two conditions. I've found some papers but most of them tells about doing de novo assembly and after that calculating the expression, but I'm not interested in assemblying, only on the differences of gene expression between genotypes.
I'd like to know if there is any formula or way to normalize and calculate the gene expression ... Thanks