Project description:<p>RNA sequencing (RNAseq) of peripheral blood lymphocytes was used to develop a means to assess immune function in a way that can be used in discovery science and applied to patients individually in clinical settings. The premise is that profiles of RNA present in immune cells is reflective of the combined influence of genetic and environmental variation on immune potential of individuals and that this potential can be tapped to understand human immunity in a variety of biological contexts. CD4+ cells were isolated from fresh whole blood via positive magnetic bead selection and cell lysates were prepared using Qiazol (QIAGEN) and stored at -80ºC for 3 to 8 months. RNA was extracted in batches for cDNA library preparation and RNA-Seq. For this study, we developed standard operating procedures for handling human blood samples and determined: a) the best way to enrich for CD4+ T cells from whole blood and yield high quality RNA, b) the sensitivity of this RNA profiling strategy, and c) the reproducibility of generated immune profiles from healthy subjects. We then developed bioinformatics processes to establish immune response signatures and immune response phenotypes within cohorts of individuals.</p>
Project description:In the present study, we used a top-down approach using microarray analysis to evaluate the novel molecular signatures that differentiate between subjects with stable coronary artery disease and normal healthy controls and which remain unresponsive to the standard therapies currently in clinical practice . Micro-array analysis revealed that inspite of concerted treatment efforts 513 genes were differentially expressed in our patient group vs healthy controls.
Project description:In the present study, we used a top-down approach using microarray analysis to evaluate the novel molecular signatures that differentiate between subjects with stable coronary artery disease and normal healthy controls and which remain unresponsive to the standard therapies currently in clinical practice . Micro-array analysis revealed that inspite of concerted treatment efforts 513 genes were differentially expressed in our patient group vs healthy controls. To access the effect of ongoing therapies on stable CAD patients and to obtain gene signatures from these patients. We selected 4 angiographically proven stable CAD patients who were on medications for more than 3 months or more from Group-1 (n=100) and compared the data with 2 normal age and sex-matched healthy Controls (n=50) belonging to the same ethnic origin and socio-economic status. RNA extraction and hybridization was done using Affymetrix microarrays.
Project description:RNA was extracted from neutrophils from healthy subjects whose neutrophil function has been extensively characterized for RNA sequencing.
Project description:We developed a method to convert gene expression signatures across the Illumina and Affymetrix platforms. We generated an E2F1 signature. We processed RNA for E2F1, MYC, and Ras signatures on the Illumina platform. 16 Melanoma samples are also included for validation.