| This dissertation addresses three main questions in molecular biology and evolution with the application of transcriptomics. Chapter I and V give the overview of introduction and conclusions to the dissertation respectively.;Chapter II titled "CADBURE: Generic tool to evaluate spliced aligner performance on RNA-Seq dataset and an evaluation of methods used for RNA-Seq based differential expression analysis"; addresses the problem of choosing an optimal spliced aligner and an optimal statistical method for quantifying differential gene expression across biological samples. Here, I introduce a novel approach for comparing alignment results of user data based on the relative reliability of uniquely aligned reads (CADBURE). In addition, I provide an evaluation of five popular (two versions of DESeq, Cuffdiff2 and two different versions of CLC) methods for discriminating differential gene expression.;Chapter III titled "Insilico analysis of the sequence features responsible for alternatively spliced introns in protein coding genes of Chlamydomonas reinhardtii"; addresses the reasons for the most frequently used alternative splice form in this alga. Using over 7 million cDNA sequences from public databases, we found that a much higher percentage of genes ( ∼20% of multi-exon genes) undergo alternative splicing than previously reported (3--5%). On comparing the shorter constitutive introns with that of retained introns, I found the majority of retained introns differ in terms of their strength of splice sites, regulatory motifs, and ability to form secondary structure.;Chapter IV titled "Genome-wide estimation of molecular evolutionary rate among five species of conifers and a comparison with Angiosperms "; addresses the estimation of substitution rate among the five species of conifers. Here, using the pairwise orthologous genes of spruce-to-spruce comparison, 3 pine-to-pine comparisons and 3 pine-to-spruce comparisons, I estimated the different substitution rate among the different lineages of conifers and compare it to Angiosperms. I find conifers, on average, have slower annual substitution rates when compared to Angiosperms. |
