Project description:Serum miRNAs are considered useful as non-invasive biomarkers for various diseases, but the optimal method for extracting RNA from serum is currently unknown. In this study, several RNA extraction kits were used to determine which kit is the optimal method. RNA was extracted from the serum of 8-week-old C57BL/6NJcl male mice according to the protocol of each RNA extraction kit. The yield of extracted RNA samples was calculated and electrophoretic patterns were evaluated by Agilent bioanalyzer. Expression patterns of the extracted RNA samples were confirmed by Agilent mouse miRNA microarray. The results showed significant differences in RNA yields in the miRNeasy serum/plasma advanced kit, and mirVana™ PARIS™ RNA and Native Protein Purification Kit compared to almost all other samples. Furthermore, two peaks were identified in the miRNeasy serum/plasma advanced kit using small RNA kit of Agilent bioanalyzer, one at 20-40 nucleotides (nt) and the other around 40-100 nt whereas the other reagents had a single peak. In addition, a high correlation was observed between the two RNA extraction kits in microarray. These results suggest that the above two kits are suitable for miRNA extraction from mouse serum.
Project description:This study systematically evaluated the AllPrep co-extraction strategy in multi-omics analysis of mouse brain tissues. At the transcriptomic level, AllPrep produced higher RNA yields and an 88.14% gene detection rate, ensuring broader coverage. In proteomics, it preserved identification depth while improving detection of membrane proteins, ubiquitin ligases, and transcription factors. Phosphoproteomics further revealed 4,347 high-confidence phosphorylation sites, including CAMK family regulatory patterns. Integrated analysis showed stronger RNA–protein correlations and enriched neuroscience-related pathways. Overall, AllPrep provides a standardized, scalable workflow that enhances data quality and supports deeper exploration of neurobiological mechanisms.
Project description:Entamoeba histolytica membrane proteins are important players in the parasite’s pathogenicity. However, most of the proteins have not been identified. This study reports the membrane proteins extracted using three extractions methods: two commercial kits (ProteoExtract® from Calbiochem and ProteoPrep® from Sigma), and a conventional laboratory method. The resulting membrane fractions (MF) and cytosolic fractions (CF)were analysed using LC-ESI-MS/MS. The proteins identified in at least two out of three biological replicates revealed a total of 490, 492, and 587 MF proteins extracted using the ProteoExtract® kit, ProteoPrep® kit and conventional method, respectively. Meanwhile, 487, 611 and 343 proteins were identified in the CF extracted using the ProteoExtract® kit, ProteoPrep® kit and conventional method, respectively. Analysis of the identified MF and CF proteins extracted by the respective extraction kits suggests that the ProteoPrep® extraction kit was the most selective in separating MF and CF among the three extraction methods.
Project description:Human stool samples were collected, microbial cells were harvested and subjected to protein extraction using different protocols for metaproteomics analyses. The protein extraction methods differ from the usage of different lysis buffer (SDS, urea and commercially available B-Per reagent), bead beating, ultrasonication and heating. The impacts of different protein extraction methods on metaproteomics results were evaluated with multiple criteria including protein yields, peptide identifications, taxonomic compositions and functional compositions.
Project description:We present three different methods for protein extraction from fresh frozen mouse heart tissue: 1) extraction using SDS buffer followed by FASP purification, 2) extraction using SDS buffer followed by S-Trap purification, and 3) extraction using a guanidine hydrochloride buffer followed by in-solution digestion. Based on bicinchoninic acid assay, all three methods display similar recovery of total protein content. However, proteomics-based analysis identified far fewer proteins from the extraction guanidine hydrochloride. SDS-FASP identifies as many proteins as the SDS-STrap method with good coverage of proteins from different cellular compartments.
Project description:Effect of DNA extraction methods on the determination of the structure of microbial communities in the phosphogypsum waste heap soil
Project description:Recent advances in (meta)genomic methods have provided new opportunities to examine host-microbe-environment interactions in the human gut. While opportunities exist to extract DNA from freshly sourced colonic tissue there are potentially valuable sources of DNA from historical studies that might also be examined. We examined how four different tissue DNA extraction methods employed in past clinical trials might impact the recovery of microbial DNA from a colonic tissue sample as assessed using a custom designed phylogenetic microarray for human gut bacteria and archaebacteria. While all methods of DNA extraction produced similar phylogenetic profiles some extraction specific biases were also observed. Real time PCR analysis targeting several bacterial groups substantiated this observation. These data suggest that while the efficacy of different DNA extraction methods differs somewhat all the methods tested produce an accurate representation of microbial diversity. This suggests that DNA samples archived in biobanks should be suitable for retrospective analyses.