Gene expression profiling of rice embryogenic calli after Agrobacterium infection

Agrobacterium tumefaciens-mediated genetic transformation has been routinely used in rice for more than a decade. However, the transformation efficiency of the indica rice variety is still unsatisfactory and much lower than that of japonica cultivars. Further improvement on the transformation efficiency lies in the genetic manipulation of the plant itself, which requires a better understanding of the underlying process accounting for the susceptibility of plant cells to Agrobacterium infection as well as the identification of plant genes involved in the transformation process. In order to investigate the related genes affecting the transformation efficiency of embryogenic calli of different rice cultivars, we used Affymetrix GeneChip® Rice Genome Array to measure the global gene expression profiling just before transformation and at four different time points after transformation (1 h, 6 h, 12 h, 24 h) in both japonica rice cultivar Nipponbare and indica rice cultivar Zhenshan 97. The mature embryo-derived embryogenic calli of Nipponbare (Nip) and Zhenshan 97 (ZS) were infected by Agrobacterium. Calli of Nip and ZS were sampled just before infection (0 h) and 1h, 6h, 12 h and 24h after infection, respectively. Three independent biological replications for each time point of the two varieties were used. To avoid the influence of polymorphisms between the probe sequence on the array and the genomes of the varieties used, we used a genomic DNA (gDNA)-based probe-selection strategy based on the hybridization efficiency of gDNA from Nip and ZS with the PM oligonucleotide probes on the rice array. The genomic DNA of Nip and ZS were extracted and hybridized to the Affymetrix Rice Genome Arrays. Three biological replications per cultivar were performed.