Sugarcane is a C4 perennial grass able to efficiently convert light energy into biomass. Bagasse, the fibrous waste product of sugar production, and biomass from high-fiber sugarcane varieties are important feedstocks for replacing fossil carbon in the production of fuels and other chemicals. Enzymatic hydrolysis can be used to convert the cellulose and hemicellulose components of the biomass into fermentable sugars. These sugars can then be fermented into ethanol and other chemicals using microbial organisms and catalysts. However, enzymatic hydrolysis is inefficient due to the recalcitrant nature of the secondary cell wall structure in which the cellulose and hemicellulose are located. Costly pretreatments are required to break down this structure prior to enzymatic hydrolysis in order to achieve high sugar yields. To reduce costs, genetic modification of biomass composition is a promising strategy for increasing the degradability of the biomass. The aims of this thesis were to identify the biomass components that can be manipulated to increase enzymatic hydrolysis efficiency and to discover genes and markers associated with one of these components that can be targeted for rapid improvement of this trait.Enzymatic hydrolysis efficiency and biomass composition were measured in leaf and culm tissues of sugarcane genotypes that varied in their fiber makeup. Tissues were left untreated or were pretreated using one of three methods (dilute acid, ionic liquid and hydrothermal) in order to understand how composition affects hydrolysis yields after various pretreatments. Interestingly, leaf tissues had sugar yields that were 1.5-fold those of the culm indicating that the composition of leaves is more susceptible to hydrolysis than that of the culm. Thus, leaf trash could be a useful resource for making biofuels and bioproducts. Additionally, genotypes performed differently depending on the pretreatment, indicating that pretreatment is an important consideration when identifying traits that impact enzymatic hydrolysis. A path analysis revealed that acid-insoluble lignin (AIL) content, syringyl to guaiacyl (S/G) ratio and xylan content had the greatest negative effects on enzymatic hydrolysis efficiency while acid-soluble lignin (ASL) and glucan content had strong positive influences. Therefore, these traits should be the main focus for genetic engineering and breeding sugarcane for utilization of the lignocellulosic fraction for a wide range of end-use applications.S/G ratio was further investigated using a differential expression analysis to determine genes that may be targeted for genetic modification of the trait. Over 2000 transcripts were differentially expressed (DE) between two groups, each consisting of six sugarcane genotypes, that contrasted for S/G ratio. Several DE transcripts corresponded to enzymes in the lignin biosynthesis pathway including caffeic acid O-methyltransferase (COMT), cinnamyl alcohol dehydrogenase (CAD), 4- coumarate:CoA ligase (4CL) and cinnamoyl-CoA reductase (CCR). Other DE transcripts encoded transporters, dirigent proteins and regulatory proteins, such as, transcription factors, F-box proteins ii and mediator complex subunits. The frequencies of single nucleotide polymorphisms (SNPs) were also compared between the two groups of genotypes to discover alleles that were associated with lignin composition. Over 2000 SNP loci across 787 unique transcripts encoding phenylpropanoid pathway enzymes, dirigent proteins, cell wall-related transcription factors and F-box proteins had group-specific expression. Overall, the DE transcripts and alleles with group-specific expression represent promising targets and genetic markers for altering the lignin composition in sugarcane.