M.ruminantium B.proteoclasticus coculture vs monocultures

This study was undertaken with two major goals in mind: i) To investigate the potential for interspecies hydrogen transfer between a hemicellulytic rumen bacterium (B. proteoclasticus) and a methanogenic achaea (M. ruminantium) Microscopic examination had previously shown B. proteoclasticus in co-culture with M. ruminantium rapidly formed cell to cell co-aggregates. ii) To examine the expression of genes involved in methanogenesis under more rumen-like conditions. Cells of M.ruminantium and B. proteoclasticus were grown in triplicate as mono- or co-cultures until a mid-exponential growth phase was reached. RNAs were extracted and purified. The relative quantities of RNAs contributed by each organism to the co-culture samples were determined by rt qPCR using genes previously shown to be constitutively expressed. The mono-culture RNAs were then combined in the determined proportions to normalise mRNA abundance with their co-culture replicates. cDNAs were synthesized and labelled and hybridised to the arrays. 3 biological replicates were used and the cDNA from each was divided in two to allow each to undergo a dyeswap (total of six arrays) giving technical replicates. Each gene was probed in triplicate adding further technical replication. Blanks and negative controls were also included. The log ratios for each probes replicates (biological and technical) were averaged prior to further analysis.