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Comprehensive Map of Molecules Implicated in Obesity
Kamal Rawal, Corresponding Author/Lead, JIIT University
Obesity is a global epidemic affecting over 1.5 billion people and is one of the risk factors for several diseases such as Type 2 diabetes mellitus and hypertension. We have constructed a comprehensive map of the molecules reported to be implicated in obesity. A deep curation strategy was complemented by a novel semi-automated text mining system in order to screen 1000 full-length research articles and over 80,000 abstracts that are relevant to obesity. We obtain a scale free network of 804 nodes and 971 edges, composed of 510 proteins, 115 genes, 62 complexes, 23 RNA molecules, 83 simple molecules, 3 phenotype and 3 drugs in“bow-tie” architecture. We classify this network into 5 modules and identify new links between the recently discovered fat mass and obesity associated FTO gene with well studied examples such as insulin and leptin. We further built an automated docking pipeline to dock orlistat as well as other drugs against the 24,000 proteins in the human structural proteome to explain the therapeutics and side effects at a network level. Based upon our experiments on several disease networks and drugs, we propose that a given drug is able to produce a therapeutic effect due to binding to several target molecules in a selected part of the network.
Whole transcriptome RNA-SEQ with Ion Torrent platform: FFPE, fresh LCM and FFPE LCM samples. Increasingly difficult.
Francesca Lessi, Last author, Pisa Science Foundation
Introduction: The advent of the NGS revolutionized cancer research by making it possible to study the complexity of cancer using high throughput sequencing methodologies. The current trends are to adapt high-throughput sequencing technologies to the level of small cell populations and even individual cells [1,2]. In our laboratory, we developed some different methods that would allow us to work optimally with very low amount of material, based on the technique of the SMARTer technology and on the Ion Torrent protocols that we modified carefully, applied to Ion Proton system.
Materials and Methods: We collected 12 FFPE tissues and 20 Laser Capture Microdissection (LCM) FFPE samples derived from brain and breast cancer and 13 LCM fresh frozen samples derived from mouse brain cells.
Results: We developed high performance methods to analyze the whole transcriptoms of our FFPE and LCM samples, obtaining a very high number of reads (78,186,377 usable reads), perfectly comparable with samples with a large amount of RNA such as samples obtained from cells or fresh tissues.
We have found that the combination of SMARTer technology and Ion TargetSeq Exome Enrichment kit, in addition to some improvements to their conventional protocols, provides excellent results with challenging samples.
Study of the active bacterial community in two membrane bioreactors
Laura Moreno Mesonero, PhD student, IIAMA
Membrane bioreactor (MBR) systems allow for the separation of mixed liquor and treated water thanks to membranes. Bacterial communities in MBRs can be quite different from conventional ones due to the long retention time and, therefore, the low metabolic cellular activity. In this work, bacterial communities from two MBR systems treating leachates were analyzed, with and without a viability dye, propidium monoazide (PMA) that selectively remove DNA from membrane-compromised cells. DNA was isolated from the samples using FastPrep®-24 cell disrupter and prepared for Illumina MiSeq sequencing. PRO341F and PRO805R primers (1) were used to target V3-V4 regions of 16S rDNA for Bacteria and Archaea. Sequenced data was analysed using Qiime 1.8.0 pipeline (2), where they were normalized and assigned taxonomy using Greengenes database clustered at 97% identity. a diversity revealed highly diverse microbial communities (41 bacterial and 1 archaeal phyla). Generally, the most abundant phyla were Bacteroidetes, Proteobacteria and Chloroflexi, which represented 63.40% of the total population. PMA treatment resulted in a reduction of the observed species in the rarefaction curves, revealing the potentially active microorganisms. Firmicutes was the phylum most affected by PMA, thus revealing that many of these bacteria were present but not active.
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