SOPs

Isolation of total RNA from Bacillus Subtilis using phenol-chloroform extraction method by maintaining cryogenec conditions initailly to prevent RNA degradation. Quality of the obtained RNA is then tested with Agilent Bioanalyser before proceeding for gene expression analysis.

An overview of creating MAGE-TAB compliant spreadsheets for transcriptomics data in SysMO SEEK

The procedure describes the preparation of fluorescent DNA probes from human mRNA or total RNA.

This is a protocol for determining the activity of the T. brucei Trypanothione synthetase under in vivo-like conditions.

This method describes the preparation of the in vivo-like buffer for the measurement of bloodstream T. brucei recombinant enzymes under pseudo-physiological conditions.

Preparation of cell free extracts of the recombinant E. coli strains expressing the gluconeogenic S. solfataricus enzymes.

Cloning and heterologous expression of gluconeogenic enzymes from S. solfataricus in E. coli

Purification of gluconeogenic enzymes from S. solfataricus in recombinant E.coli extracts

This assay uses a dual-wavelength spectrophotometer to quantify cytochromes present in the E. coli respiratory chain.

This is a well-established, classical genetic method of constructing chromosomal monolysogenic fusions to a promoterless lacZ gene.

The phosphorylation level of a particular protein can be determined using a procedure based upon western immunoblotting, with Phos-tag™ reagent present in the SDS-PAGE gel. The Phos-tag™ reagent, supplied in the form of Phos-tag™ acrylamide (Wako Pure Chemical Industries, AAL-107), causes proteins to be resolved both on the basis of size and phosphorylation state. This means that phosphorylated and de-phosphorylated forms of the same protein can be distinguished.

The Gene-doctoring method of lambda-red deletion (Lee et al., 2009) was modified slightly to create chromosomal mutations of fnr.

This method describes how one can quench metabolism of Escherichia coli and extract metabolites from many kinds of metabolite classes like: nucleotides, sugar-phosphates, organic acids ....

Sample preparation procedure for metabolic analysis on T. b. brucei 427 using LC/MS

Labelling and extraction procedure for uniformly 13C-labelled E. coli (MG1655) for absolute quantification using isotope dilution technique by LC-MS

Sample preparation procedure for metabolic footprint analysis on T. b. brucei 427 using LC/MS

Assay methodologies for individual glycolytic isoenzymes from the Mendes Group, University of Manchester, UK

This is the general protocol for the glycolytic enzyme measurements. Detailed Information for each Enzyme can be found in the SOP: Assays for measuring the activities of the individual glycolytic isoenzymes of Saccharomyces cerevisiae

For the study of mRNA decay rates, transcription was inhibited with ActinomycinD, and RNA splicing with Sinefungin, at different time points, in the Matthews lab. rRNA depleted RNA was extracted from each of the samples in the Clayton lab, and sent for deep sequencing at the BioQuant facility in Heidelberg

Protocol for transfer of plasmids into Clostridium acetobutylicum ATCC 824 by electroporation

ClosTron mutants should always be subjected to Southern blot analysis to ensure that only one intron insertion has occurred.

A protocol to improve conventional, recombination-based gene knock-out methodologies thtough the provision of negative selection markers, pyrE or codA.

Protocol for transfer of plasmids into Clostridium spp. by conjugation

A refined and streamlined procedure to generate mutant in a wide range of different clostridial species, using group II intron retargeting methodologies.

This protocol describes the transcriptional profiling of E. coli cultures using microarrays. The protocol utilises RNA isolated as described in another SOP (SUMO RNA isolation from E. coli) and with hybridisation to Ocimum Ocichip E. coli K-12 microarrays.

This SOP describes the preparation of E. coli RNA

SOP for growing yeast in anaerobic conditions