The present study is restricted to the isolation, morphological and molecular characterization of Sordaria fimicola by applying PCR based molecular markers coupled with high resolution melt analysis to obtain adequate data for measuring genetic variations. All the collected samples isolated from different herbivore dung samples were plated; incubated and S. fimicola colonies were identified on solid PDA media. A total of 61 fungal isolates of S. fimicola were isolated by moist chamber and single plate isolation techniques. The effect of different physical parameters such as pH (4.5-8), temperature (20-45 ºC), carbon (cellulose, glucose and maltose), nitrogen sources (sodium nitrate, ammonium sulphate and casein hydrolysate) and UV light on the mycelial and perithecial growth was also determined on different strains of S. fimicola. Present attempt was made to gauge molecular differences on Sordaria fimicola‟s genome present on site of Evolution Canyon. All strains and isolates were authenticated by targeting internal transcribed spacer and hypervariable regions (ITS1, ITS2, full ITS, V3, V4 and V9). No sequence variations were observed in case of ITS and V9 regions while two polymorphic sites on position 212 A (C); 213 G (A) were found in V3 and V4 region in strains isolated from the south slope of Evolution Canyon. The PCR-based technique of randomly amplified polymorphic DNA (RAPD) was used to fingerprint and assess the genetic relatedness among eight isolates of the fungus S. fimicola isolated from various areas of Lahore, Pakistan. Each DNA sample was amplified with each of eight RAPD primers during the initial screening and the products were resolved on 1.5 % agarose gel, stained with ethidium bromide and snapped under gel documentation system. ii Four primers failed to show any amplification but the remaining four (50 %) primers generated a total of 22 bands (5.5 bands per primer) among the eight isolates of S. fimicola. Of these 22 resolved bands, 13 (3.5 per primer) were polymorphic and the remaining 9, bands were common (monomorphic) among these isolates. The least efficient primer was R8 (1.79 %), while the most efficient one was R3 (8.93 %). Primer R3 had the highest PIC value (0.5) and identified all eight isolates through unique patterns of banding. Jaccard similarity coefficient ranged between 0.2-0.6. Sequence products of 485 bp and 811 bp long nucleotides were obtained from two RAPD loci (RL-7 and RL4) and converted to sequence characterized amplified region (SCAR400 and SCAR500) markers for S. fimicola. Another aim of the experiment was to look for natural genetic variation in different natural strains of Sordaria fimicola isolated from natural and stressed ecological conditions of Evolution Canyon. It was challenging because there was no sequence information at the time, and I was required to design a set of PCR primers that would reliably amplify in S. fimicola and it was hoped that the amplified products were polymorphic. Hence the decision was made to target randomly selected genes and regions containing short sequence repeats, putting primers in the exonic parts to amplify the intron(s) they flank and used S. macrospora sequence information to strategically place the "exon-primed, intron-crossing" primers to determine nucleotide variations in fifty strains of Sordaria fimicola by sequencing PCR amplicons. As compared to the strains isolated from the north slope, strains isolated from the south slope exhibited point mutations on various positions and enrichment of short sequence repeats was observed more in strains isolated from the south slope of EC. Frequency clock proteins are significant factor of circadian clocks which regulate gene expression that react to various environmental conditions in many organisms and are iii principally involved in rhythmic movements displayed by numerous filamentous fungi. Mating type a-1 proteins, encoded by mat-a-1 genes, control the sexual compatibility and vegetative incompatibility with A mating types in many ascomycetes. In this study, I have also explored the phosphorylation and glycosylation status of frequency clock and mating type a1 proteins on serine/threonine/tyrosine residues using NetPhos3.1 and YinOYang 2.1 server.