SOP for cDNA labelling and Microarray hybridizaton Version 1
The reverse transcriptase synthesizes DNA, which complements the mRNA template (complementary DNA, cDNA). Cy3/Cy5-dCTP are incorporated into cDNA during Reverse transcription. The obtained Cy3/Cy5 cDNA are then competitively hybridised onto Agilent microarray slide and subsequently scanned.
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Created: 26th Oct 2009 at 14:48
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Version 1 (earliest) Created 26th Oct 2009 at 14:48 by Praveen kumar Sappa
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Projects: BaCell-SysMO
Institutions: University of Greifswald
I am PhD student at Prof.Uwe Voelker lab in Department of Functional Genomics. My area of research is microbial functional genomics in particular analysing the whole transcriptome(by microarray and other molecular biolology methods) of B.subtilis under various stress conditions. I use QconCAT strategy for absolute quantification of carbon metabolic enzymes via MRM(multiple reaction monitoring) by LC-MS/MS. I also perofrm experiments for understanding of dynamics of SigmaB network for modelling.
SysMO is a European transnational funding and research initiative on "Systems Biology of Microorganisms".
The goal pursued by SysMO was to record and describe the dynamic molecular processes going on in unicellular microorganisms in a comprehensive way and to present these processes in the form of computerized mathematical models.
Systems biology will raise biomedical and biotechnological research to a new quality level and contribute markedly to progress in understanding. Pooling European research ...
Projects: BaCell-SysMO, COSMIC, SUMO, KOSMOBAC, SysMO-LAB, PSYSMO, SCaRAB, MOSES, TRANSLUCENT, STREAM, SulfoSys, SysMO DB, SysMO Funders, SilicoTryp, Noisy-Strep
Web page: http://sysmo.net/
BaCell-SysMO 2 Modelling carbon core metabolism in Bacillus subtilis – Exploring the contribution of protein complexes in core carbon and nitrogen metabolism.
Bacillus subtilis is a prime model organism for systems biology approaches because it is one of the most advanced models for functional genomics. Furthermore, comprehensive information on cell and molecular biology, physiology and genetics is available and the European Bacillus community (BACELL) has a well-established reputation for applying ...
Programme: SysMO
Public web page: http://www.sysmo.net/index.php?index=53
The aim of the study is to assess the global function of RNase Y in RNA processing and degradation in Bacillus subtilis. To this end we constructed a strain allowing controlled depletion of RNase Y and used microarrays to analyze the transcriptome in response to the expression level of RNase Y.
The objective of this project is an integrated understanding the metabolic, proteomic and genetic network that controls the transition from growth to glucose starvation. This transition is a fundamental ecophysiological response that serves as a scientific model for environmental signal integration and is pivotal for industrial fermentations of Bacillus that occur predominantly under nutrient starvation.
Keywords: Glucose starvation, Transcriptomics, Proteomics, Metabolomics,Bacillus subtilis,
Submitter: Praveen kumar Sappa
Studies: B. subtilis Transcription Factor Competition, Batchfermentation exp-starv01_090204, Biphase Batch Fermentation(2009/02/04), Controlled sigmaB induction in shake flask, Transition to starvation in shake flask
Assays: 2D-gelbased analysis of intracellular proteins, Absolute quantification of proteins by the AQUA-technology, B. subtilis Transcription Factor Competition - theoretical interpretation, B. subtilis Transcription Factor Competition - theoretical interpretation, Fermentation-BM5_SysMo, Gene expression(Transcriptome), IPTG induction of sigmaB in BSA115, IPTG induction of sigmaB in BSA115, Relative quantification of proteins by metabolic labeling, Stressosome activation dynamics, metabolome-LCMS
High salinity chemostat cultivation, multiomics sampling (proteome, transcriptome, metabolome, fluxome) and modelling of carbon core metabolism of Bacillus subtilis 168.
Submitter: Sandra Maass
Studies: B. subtilis_SysMo2_Chemostat_growthrate-salt, Fluxome analysis of Bacillus subtilis 168 under osmotic stress
Assays: 13C Metabolic Flux Analysis of Bacillus subtilis 168 in continuous high-..., Absolute quantification of proteins by the AQUA-technology, Absolute quantification of proteins using QconCAT technology, Relative quantification of proteins by metabolic labeling, Transcriptome data for chemostat cultivated samples, extracellular metabolites, intracellular metabolites
The aim of the study is to assess the global function of RNase Y in RNA processing and degradation in Bacillus subtilis. To this end we constructed a strain allowing controlled depletion of RNase Y and used microarrays to analyze the transcriptome in response to the expression level of RNase Y.
Submitter: Praveen kumar Sappa
Investigation: Identification of targets of the essential RNas...
Assays: Transcritome data_Identification of targets of the essential RNase Y of ...
B. subtilis was grown in minimal media in a chemostat at different growth rates (µ= max, µ=0.1, µ=0.4) and in the presence of 1.2M NaCl (µ=0.1) with or without glycinebetaine. Transcriptome, proteome and metabolome were investigated.
Submitter: Sandra Maass
Investigation: Multiomics study of Bacillus subtilis under osm...
Assays: Absolute quantification of proteins by the AQUA-technology, Absolute quantification of proteins using QconCAT technology, Relative quantification of proteins by metabolic labeling, Transcriptome data for chemostat cultivated samples, extracellular metabolites, intracellular metabolites
The main aim of this experiment is to actively grow B.subtilis in presense of glucose until high optical density in an aerobic fermentor and then, at a definite point of the growth, the glucose supply is shut down which leads to complete glucose exhaustion in the media. Simultaneusly samples for transcriptomics, intra and extracellular metabolomics, intra and extracellular proteomics are harvested through out the experiment.
Submitter: Praveen kumar Sappa
Investigation: The transition from growing to non-growing Baci...
Assays: 2D-gelbased analysis of intracellular proteins, Absolute quantification of proteins by the AQUA-technology, Fermentation-BM5_SysMo, Gene expression(Transcriptome), Relative quantification of proteins by metabolic labeling, metabolome-LCMS
Submitter: Praveen kumar Sappa
Assay type: Transcriptomics
Technology type: Custom Array
Investigation: The transition from growing to non-growing Baci...
Transcriptome analysis for the samples harvested from Chemostat cultivated samples.
Submitter: Praveen kumar Sappa
Assay type: Transcriptomics
Technology type: Microarray
Investigation: Multiomics study of Bacillus subtilis under osm...
Submitter: Praveen kumar Sappa
Assay type: Transcriptomics
Technology type: Custom Array
Investigation: Identification of targets of the essential RNas...
