Bacterial wilt in ginger (Zingiber officinale Rosc.) caused by Ralstonia solanacearum is one of the most important production constraints in tropical, sub-tropical and warm temperature regions of the world. Lack of resistant genotype adds constraints to the crop management. However, mango ginger (Curcuma amada Roxb.), which is resistant to R. solanacearum, is a potential donor, if the exact mechanism of resistance is understood. To identify genes involved in resistance to R. solanacearum, we have sequenced the transcriptome from wilt-sensitive ginger and wilt-resistant mango ginger using Illumina sequencing technology. A total of 26387032 and 22268804 paired-end reads were obtained after quality filtering for C. amada and Z. officinale, respectively. A total of 36359 and 32312 assembled transcript sequences were obtained from both the species. The functions of the unigenes cover a diverse set of molecular functions and biological processes, among which we identified a large number of genes associated with resistance to stresses and response to biotic stimuli. Large scale expression profiling showed that many of the disease resistance related genes were expressed more in C. amada. Comparative analysis also identified genes belonging to different pathways of plant defense against biotic stresses that are differentially expressed in either ginger or mango ginger. The identification of many defense related genes differentially expressed provides many insights to the resistance mechanism to R. solanacearum and for studying potential pathways involved in responses to pathogen. Also, several candidate genes that may underline the difference in resistance to R. solanacearum between ginger and mango ginger were identified. Finally, we have developed a web resource, ginger transcriptome database, which provides public access to the data. Our study is among the first to demonstrate the use of Illumina short read sequencing for de novo transcriptome assembly and comparison in non-model species of Zingiberaceae.

No abstract

Understanding biology and genetics at molecular level has become very important for dissection and manipulation of genome architecture for addressing evolutionary and taxonomic questions. Knowledge of genetic variation and genetic relationship among genotypes is an important consideration for classification, utilization of germplasm resources, and breeding. Molecular markers have contributed significantly in this respect and have been widely used in plant science in a number of ways, including genetic fingerprinting, diagnostics, identification of duplicates and selecting core collections, determination of genetic distances, genome analysis, developing molecular maps, and identification of markers associated with desirable breeding traits. The application of molecular markers largely depends on the type of markers employed, distribution of markers in the genome, type of loci they amplify, level of polymorphism, and reproducibility of products. Among many DNA markers available, random amplified polymorphic DNA (RAPD) is the simplest and cost-effective and can be performed in a moderate laboratory for most of its applications. In addition RAPDs can touch much of the genome and has the advantage that no prior knowledge of the genome under research is necessary. The recent improvements in the RAPD technique like AP-PCR, SCAR, DAF, SRAP, CAPS, RAMPO, and RAHM can complement the shortcomings of RAPDs and have enhanced the utility of this simple technique for specific applications. Simple protocols for these techniques are presented.

A systematic validation of reference genes is a prerequisite for the proper normalization of gene transcripts. In the present study, the annotated sequences from black pepper (Piper nigrum L.) leaf transcriptome were used as reference genes namely actin (PnACT), glyceraldehyde phosphate dehydrogenase (PnGAPDH), b-tubulin (PnTUB), ubiquitin conjugating enzyme (PnUBCE), 18srRNA and elongation factor-1-a (PnElF) to identify the stable reference gene. We focused the selection of stable reference gene on important biotic stress (Phytophthora) with different algorithms (geNorm, NormFinder and BestKeeper) along with Reffinder which resulted in identification of PnGAPDH and PnUBCE as stable genes. Norm qPCR (R package) was also used to estimate the stability of the selected genes. We elucidated the expression patterns of a target gene PnBGLU which codes for 1,3 beta glucanase with most stable as well as least stable reference genes by which the importance of selecting the stable gene for gene expression studies in this system was emphasized. The mean expression levels of PnBGLU was significantly overestimated and misinterpreted when least stable reference gene was used as normalizer. The selected reference genes on further analysis of the expression dynamics of PnBGLU among resistant and susceptible genotypes showed PnGAPDH as the suitable reference gene for P. nigrum-P. capsici pathosystem.

No abstract