Assays

intracellular metabolite measured by GC/MS and LC/MS

Some examples of transcriptomics templates for Affymetrix data that conform to the MAGE-TAB specification. These templates were taken from the GEO website (http://www.ncbi.nlm.nih.gov/geo/info/spreadsheet.html) and modified to conform to the SysMO-JERM (Just enough Results Model) for transcriptomics. Using these templates will mean easier submission to GEO/ArrayExpress and greater consistency of data in SEEK.

A example of an RT-PCR Excel template. RT-PCR is Reverse Transcriptase PCR (NOT to be confused with Real Time PCR, which is normally referred to as qPCR)

This template was taken from the GEO website (http://www.ncbi.nlm.nih.gov/geo/info/spreadsheet.html) and modified to conform to the SysMO-JERM (Just enough Results Model) for transcriptomics. Using templates will mean easier submission to public databases on publication and greater consistency of data in SEEK.

This Excel template is an example taken from the GEO web site (http://www.ncbi.nlm.nih.gov/geo/info/spreadsheet.html#GAtemplates) which has been modified to conform to the SysMO JERM (Just Enough Results Model). Using templates helps with searching and comparing data as well as making it easier to submit data to public repositories for publications.

Some examples of transcriptomics templates for NimbleGen data that conform to the MAGE-TAB specification. These templates were taken from the GEO website (http://www.ncbi.nlm.nih.gov/geo/info/spreadsheet.html) and modified to conform to the SysMO-JERM (Just enough Results Model) for transcriptomics. Using templates will mean easier submission to GEO/ArrayExpress upon publication and greater consistency of data in SEEK for easier searching and comparing.

Examples of proteomics templates for gele electrophoresis data that conform to the MIAPE-GE specification

This assay is designed to measure the decay kinetics of mRNA in T. brucei blood forms. T. brucei lacks the canonical transcriptional regulation employed by other eukaryotes through transcription factors, and relies almost entirely on regulation of mRNA decay and further downstream steps in order to control gene expression. 3 replicates of 8 time points were taken to measure mRNA abundance in the cell using RNA-seq. The first time point was fot WT, untreated cells; the second was 5 min after the ...

1. Preparation of B. subtilis cultures

Inoculate cells from -80°C stocks in 10 ml time-lapse microscopy (TLM) medium (62 mM K2HPO4 , 44mM KH2PO4, 15 mM (NH4)2SO4, 6.5 mM sodium citrate, 0.8 mM MgSO4, 0.02 % casamino acids, 27.8 mM glucose, 0.1 mM L-tryptophan, the pH was set to 7 using a KOH solution) supplemented with antibiotics, if necessary. Grow the cells overnight in a shake flask (30°C, 225 rpm). The following morning, dilute the cells 1:10 in pre-warmed chemically defined medium (CDM) (62 ...

S. pyogenes wildetype, an arcA- and a glnA-deletion mutants were grown in CDM-LAB cultures at pH 6.5 and 7.5 and at a growth rate of 0.05

In order to construct an in vivo-like buffer for S. pyogenes, the intracellular concentrations of Fe, K, Mg, Mn Na, P and S elements were determined via ICP-AES (inductively coupled plasma atomic emissionspectroscopy) method at the Institute of Land Use, University of Rostock. The samples for the analysis were obtained from a steady state culture grown on CDM-LAB with glucose.

We developed a new metabolomics protocol, which involved a comparison of different harvesting techniques, quenching solutions and extraction methods. Cell leakage and metabolite recovery was determined by ATP measurements

This assay is designed to obtain the in vitro kinetic data of T. brucei recombinant trypanothione synthetase. The enzyme catalyzes the ATP-dependent ligation of spermidine (Spd) and GSH to generate glutathionylspermidine (Gsp) and also of Gsp and GSH to finally produce trypanothione (T(SH)2). The data was obtained in an spectrophotometric assay that links ADP production with NADH consumption through the piruvte kinase and lactate dehydrogenase.

This assay is for method development to quantify intra- and extra-cellular metabolites on T. brucei 427 bloodstream form using isotope ratio based MS technique with 13C-labelled E. coli extract

Intracellular metabolites in T. brucei at different stage of cell growth have been quantified absolutely by isotope ratio based MS technique using uniformly 13C-labelled E. coli extract. This is the case study for method development of absolute quantification for metabolic flux analysis.

Metabolite profiling on T. brucei exposed to methylene blue has been carried out using LC-MS to investigate metabolic changes caused by oxidative stress

26 intracellular metabolites (amino acids, polyamines, TCA intermediates) in T. brucei exposed to methylene blue have been absolutely quantified using isotope ratio based MS technique.

26 intracellular metabolites (amino acids, polyamines, TCA intermediates) in T. brucei under pH stress (pH8.7) have been absolutely quantified using isotope ratio based MS technique.

Extracellular metabolites in T. brucei at different stage of cell growth have been quantified absolutely by isotope ratio based MS technique using uniformly 13C-labelled E. coli extract. This is the case study for method development of absolute quantification for metabolic flux analysis.

Glucose transporter mutants were analyzed under aerobic and aerobic conditions in batch cultures with glucose as substrate. Acetate formation rates and glucose consumption rates were measured, as well as extracellular cAMP concentrations.

Mutant strains which lack one or more of the glucose transport systems were analyzed in aerobic chemostat cultures and compared to batch cultures.

MG1655 and mutant strains with defects in glucose transport systems were analyzed in aerobic and anaerobic batch cultures.