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Protocol for agarose-gel typing set-up

MLVA data can be produced by a number of methods, so it can be run in any laboratory setting. For large-scale projects (hundreds of strains) it is more comfortable and less time consuming to use to use capillary electrophoresis equipment to measure PCR DNA fragment length. Lista et al., BMC microbiology 2006 multiplex 8 loci in a single PCR in their Bacillus anthracis MLVA assay. It is important to understand that all methods, i.e. agarose or more sophisticated methods, will produce the same MLVA data. So just use what is most convenient for you ! This agarose-oriented protocol was written in the context of the Paris meeting (organised in Orsay, March 2006, as part of the COST845 project). It is provided as general guidelines, with no garanty of any kind, and must be adapted to local context. The key aspects, which should be strictly followed when running agarose gels, is the importance of loading a sufficient number of marker lanes, and a sufficient number of reference strain lanes, as described. The reference strain can be any strain with well-characterized allele size. Also, it is strongly recommended to use multi-chanel pipets, in order to limit loading errors. It is advised to evaluate the data quality of a data set by blind-typing in duplicate 10 samples picked at random.