BMC Genomics 2009, 10:415 http://www.biomedcentral.com/1471-2164/10/415 FRFuiNgsaAurriubemlo5tinacnoamlypsiastiobflefeiwnt eimramctuionnity induced genes of Ca- RNA blot analysis of few immunity induced genes of Ca-Fusarium incompatible interaction. 20 g of total root RNA isolated from 25-day old chickpea seedlings were separated by 1.5% agarose gel. Lane U represents signal from the water treated control plants whereas lane 24 h repre- sents RNA from 24 hours post pathogen challenge. The blots were hybridized with the respective gene specific probes as revealed by the microarray results. intron and splice sites http://www.ncbi.nlm.nih.gov/ SNP/. The regulation of gene expression is an important mechanism, which controls cellular processes and it is conceivable that the functional transcripts resulted from a controllable transcription and posttranscriptional proc- esses may shape the specificity at genotype/haplotype level. In an attempt to understand the molecular basis of immune response in chickpea during vascular wilt, the transcriptional changes were monitored by EST sequenc- ing approach. The sequences reported in this study have enriched the collection of ESTs in this food legume which would form a basis in our understanding of plant metab- olism, development and adaptation to pathogen stress. These data are particularly important, at least in part, due to the fact that there are only 7464 chickpea EST sequences available currently in the NCBI database and we have doubled the number. This would broaden the current chickpea database for further functional studies. Although a large percentage of ESTs (46.27%) was com- mon to both the susceptible and resistant genotypes, a sig- nificant percentage of ESTs was specific to each of them. The percentage of unigenes specific to susceptible geno- type was 19.16%, while that of resistant genotype was 34.56% (Figure 1) indicating the possible divergence in immune signaling pathways. The occurrence of almost the double the number of ESTs in the resistant genotype as compared to the susceptible one may be attributed to the fact that responses in the susceptible and resistant geno- types to pathogen stress are largely quantitative and kinet- ically controlled. It is known that resistant genotype shows immediate response upon pathogen infestation which is of high amplitude, in contrarary, the susceptible genotype shows late response [92]. Further, among the entire set of ESTs, 40% represented the sequences of hypo- thetical proteins or unknown proteins or showed no sig- nificant homology to the previously identified genes or proteins in the NCBI database. These ESTs may represent unique chickpea sequences. It is of interest that the Fusar- ium infection of chickpea causes substantial changes in the expression of numerous unknown transcripts yet to assign a cellular function. Comparative abundance of ESTs in resistant and susceptible genotypes Analysis of EST frequency comprising a contig and the source of the contig provides an insight with respect to the spatial expression level and biochemical functions even without subsequent microarray analysis. In most of the cases and within statistical limitations [93], the abun- dance of a specific transcript in the EST dataset serves as a measure of gene expression. Therefore, we identified and compared relative abundance of ESTs in different contigs to unravel the transcript signatures, which might deter- mine the genotype-specific immune response. Contigs having ESTs from a single genotype were designated as specific. Of the 973 contigs, 433 were found to be specific to the resistant genotype, 261 to the susceptible genotype, and 279 contigs represented both the genotypes. While looking at the abundance of ESTs, we observed that the genes involved in the pathways that could be employed in cell defense, were represented more in resistant genotype than susceptible one. In this context, the gene encoding transaldolase that is involved in pentose phosphate path- way was found to be associated with the resistant geno- type. Expression of this gene may be explained to the possible need of NADPH required for biosynthesis of var- ious secondary metabolites like PR proteins. Similar was the case with enolase, a glycolytic enzyme, involved in the formation of phosphoenol pyruvate, which contributes to the formation of aromatic amino acids via shikimic acid pathway [94]. Aromatic amino acids act as primary sub- strates for phenylpropanoid pathway. Many genes involved in phenylpropanoid pathway, for example, chal- cone reductase, chalcone synthase, and isoflavone reduct- ase were more abundant in resistant genotype. These are canonical genes well known for their definitive role in defense pathways [66,67]. Many other stress related genes were also found to be more abundant in resistant geno- type. Some of the examples are putative imbibition pro- tein, aquaporin, and DNAJ-like protein. Although aquaporin is known to be induced in response to abiotic stresses [95] its role in patho-stress remains to be investi- Page 13 of 21 (page number not for citation purposes)