P.U. KrishnarajSachin A. More2016-11-112016-11-112011http://krishikosh.egranth.ac.in/handle/1/85110Metagenomics is an emerging field of science mainly aims at studying the activity of uncultured microorganisms through the nucleotide sequences. Standard culture techniques allow only 1% of microorganisms to grow in defined laboratory conditions. Thus, culture dependent study of microbial diversity unravels only part of the microbial populations present in the environment. The present study was done to analyze the soil microbial diversity between forest and farm soils through the metagenomic approach. The success of any metagenomic study depends on the method of DNA extraction. So considering the significance and need, method was optimized for extraction of microbial community DNA from forest and farm soils. Extracted DNA was purified with the aid of chemical flocculation using FeCl3 as chemical flocculent. Purity of DNA was confirmed by effective PCR amplification using 16S rDNA primers. The purified DNA from respective soil was PCR amplified using DGGE specific 16S rDNA primers. The amplified PCR products were then separated on DGGE gel to analyze fingerprinting pattern in each soil types. Soil microbial diversity analyses of fingerprints were done by using NTSYS software. DGGE analysis was shown that forest soil microbial community shares more similarity with organic farming soil microbial communities. The sequence based metagenomics approach was used to know the microbial community composition. This was done by constructing PCR based 16S rDNA library of metagenome of each soil using T/A cloning method. About 17 clones of each soil were randomly selected from respective 16S rDNA library and sequenced. Blast analyses of each clone form respective soil were done in RDP database to know the taxonomic affiliation of each clone. Further the processed 51 sequences were then submitted to NCBI database. Blast results obtained shown that majority of the microbial communities in forest and farm soil contained uncultured microorganisms.Plant BiotechnologyThesis