Combined ESTs from plant-microbe interactions: using GC counting to determine the species of origin

A diversity of microorganisms establishes intimate associations with plants. Whether pathogenic or symbiotic, such interactions are the result of complex recognition events between plants and microbes, leading to signaling cascades and regulation of countless genes involved in the interaction. A key step in unraveling the mysteries of plant-microbe interactions lies in defining the transcriptional changes that occur in both the host and the microbe during their association. The sum of the transcripts, from both host and microbe, which are produced during their association, has been defined as the interaction transcriptome. One approach to analyze interaction transcriptomes is to perform large-scale sequencing of cDNAs (expressed sequence tags or ESTs) obtained from infected plant tissue and representing a mixture of host and microbe sequences. In some cases, the two organisms have markedly different GC content, allowing most ESTs to be easily distinguished on this basis. In this chapter, we describe a GC counting method to determine the species of origin of ESTs obtained from interactions between plants and oomycetes or other high GC content microbes.