[Under construction]

A selection of Trypanosoma brucei-specific protocols, updated on an ad hoc basis. For details of standard molecular biology protocols see, for example, Current Protocols in Molecular Biology or Short Protocols in Molecular Biology, both edited by Frederick Ausubel and published by Wiley.

Trypanosoma brucei culture & manipulation

• Bloodstream-form (BSF) culture
• BSF electroporation and selection of transformants
• BSF to procyclic differentiation
• Procyclic-form culture

Trypanosoma brucei growth analysis

• Basic growth assay (& calculating a doubling time)
• EC₅₀ analysis (calculating a ‘half maximal effective growth-inhibitory concentration’)
• Isobologram analysis (synergistic, additive & antagonistic inhibitor interactions)

Molecular Biology

• ‘Sticky end’ cloning (getting a PCR amplified insert into a plasmid)
• Phenol:chloroform:isoamyl alcohol extraction and ethanol precipitation of DNA
• Electroporating E. coli and identifying recombinant clones

Cell biology

• Immunofluorescence
• Mitotracker staining (localising the mitochondrion)
• DAPI cell cycle analysis
• FACS analysis

Protein analysis

• Protein preparation
• Hypotonic lysis (soluble and ‘membrane-associated’ fractions)

RNA analysis

• PAGE separation of small RNAs
• Transcriptional run-on analysis (detecting nascent RNAs in Trypanosoma brucei)