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The main aim of the work is to study the regulation of gene expression in the interaction between rice and Magnaporthe oryzae by gene chip technology. In this study, we mainly focused on changes of gene expression at 24, 48, and 72 hours post-inoculation (hpi), through which we could conduct a more comprehensive analysis of rice blast-related genes in the process of infection. The results showed that the experimental groups contained 460, 1227, and 3937 significant differentially expressed genes at 24, 48, and 72 hpi, respectively. Furthermore, 115 significantly differentially expressed genes were identified in response to rice blast infection at all three time points. By annotating these 115 genes, they were divided into three categories: metabolic pathways, proteins or enzymes, and organelle components. As expected, many of these genes were known rice blast-related genes; however, we discovered new genes with high fold changes. Most of them encoded conserved hypothetical proteins, and some were hypothetically conserved genes. Our study may contribute to finding new resistance genes and understanding the mechanism of rice blast development.

In vitroembryogenic callus is a critical factor for genetic transformation of rice, especially for indica varieties. In this study, we investigated the relationship between polyamines, including putrescine (Put), spermidine (Spd) and spermine (Spm), and callus browning, and we studied the effect of exogenous Put on callus regeneration and on the content of endogenous polyamines. In addition, the expression levels of arginine decarboxylase gene (Adc1) and S-adenosylmethionine decarboxylase gene (Samdc) in embryogenic callus were studied by quantitative Real-time PCR analysis. The results showed that the contents of endogenous Put and Spd in the browning callus were significantly lower than those in normal callus. Exogenous Put could effectively improve the growing state of callus of indica rice and enhance the development of embryogenic callus. The content of endogenous polyamines in embryogenic callus, especially Spd and Spm, was increased after addition of exogenous Put. Additionally, exogenous Put also had an obvious impact on the expression levels ofAdc1but partial effect on the expression levels ofSamdcgene. This study could increase the knowledge of both embryogenic callus induction and polyamine catabolism in callus in indica rice.

Rapid and inexpensive preparation of genomic DNA from rice seeds for marker-assisted selection and seed purity estimation is a major bottleneck for plant breeders. Here, we describe a high-throughput method that provides DNA at sufficient quantity and quality for these applications. Optimization of buffer composition and individual protocol stages allow processing of 384 samples within 2 h, yielding templates that reliably support downstream polymerase chain reaction of single copy amplicons up to 1.2 kb.

Rice blast resistance (R) genes-mediated resistance response depends on various resistance-related genes involved in incompatible interactions. In this work, the expression profiles of innate rice immunity related genes were examined in the mediated resistance response of true/field resistance genes. Three sets of rice near-isogenic lines (NILs) were used: the resistant NILs carrying true resistance genes in the genetic background of the susceptible cultivar Nipponbare (NB), NB-Pib, NB-Pizt, NB-Pik and NB-Pita2; NILs bearing field resistance genes pi21 in the susceptible cultivar Aichiasahi (AA) AA-pi21, Kahei (KHR). The marker gene OsWRKY45 of salicylic acid (SA) signalling was upregulated in all tested cultivars. And, JAmyb (marker gene of jasmonic acid signalling) showed higher upregulation in the resistance lines with nucleotide-binding sites and leucine-rich repeat (NBS-LRR) R genes Pib, Pizt, Pik, Pita2 and Pikahei than in NB and KHS. SalT of abscisic acid (ABA) signalling may be involved in the R/Avr interaction, including Pizt, Pik, pi21 and Pikahei. However, SalT was shown to negatively regulate Pib/AvrPib interaction. OsPR1b and PBZ1 were differentially expressed and strongly activated at a later stage by 48 h post-inoculation. Interestingly, there was evidence that OsPR1b and PBZ1 played an important role in the pi21-mediated response. It was shown that OsRAR1 could be upregulated in the true resistance line NB-Pita2 and the field resistance line KHR, while OsSGT1 and OsHSP90 could be upregulated in all tested lines. The involvement of these genes illustrated the complexity of the downstream signalling pathways in the mediated resistance response of true/field resistance genes.

Plant basic leucine zipper (bZIP) proteins play an essential role in the genes expression and regulation in higher plants. They have been shown to regulate diverse plant specific phenomena, including germination, floral induction and development, seed maturation, photomorphogenesis, biotic and abiotic stresses. Resistance response mediated by the rice blast resistance gene Pi36 is a typical signal transduction, in which 12 OsbZIP genes were differentially expressed by microarray analyses. To understand the potential function of OsbZIP genes during the defense responses against rice blast, the expression analysis of these OsbZIP genes, in response to the blast fungus inoculation and the related defense signal molecules induction, were further conducted using real-time fluorescent quantitative polymerase chain reaction (PCR) technique. Our data indicates that among the 12 OsbZIP genes, the expression level eight tested genes were differentially regulated and maintained to 96 h points post inoculation in rice resistant and susceptible cultivars during Magnaporthe oryzae infection, and all of them were also significantly up-regulated by one or several kinds of exogenous plant hormones stresses. Although, these genes were induced only at early time points (1 to 24 h); it is evident that the OsbZIP genes may be involved in different signaling pathway, and play potential important functions in the defense response to rice blast. Keywords: OsbZIP transcription factors, rice blast, resistance response, plant hormones stresses. African Journal of Biotechnology Vol. 12(34), pp. 5294-5302

Rice blast is one of the most serious diseases of cultivated rice. We characterize here the nucleotide variation present at nine blast resistance (R) genes within a collection of 13 indica and 13 japonica rice cultivars, along with 12 accessions of wild rice (Oryza rufipogon). Compared with other functional genes, high genetic diversity has been observed in blast R genes (mean nucleotide diversity of 3.2%), which might be the most important characteristics of blast R genes. Even if the overall diversity in R genes is high, the genes can also be divided into three types according to different degrees of conservation by principal component analysis: relative conservation (Pita, Pit, Pi5-1, and Pid-2), intermediate diversity (Pi37, Pib, and Pi9), and high diversity (Pi36 and Pikm-2), suggesting that these types of R genes might recognize different categories of molecules released by rice blast pathogens. Our data also revealed that a significant regression was observed between the wild and the cultivated acc...

The single‐locus rice blast resistance gene Pi36 is of the coiled‐coil NBS‐LRR (nucleotide binding site–leucine‐rich repeat) type. We have investigated its evolution by resequencing a set of haplotypes (six ssp. indica and four ssp. japonica cultivars and three wild rice accessions) both within and outside the gene’s coding region. It was found that the average nucleotide diversity was ∼4.5%, characteristic of an intermediately diversified pattern. The Ka/Ks ratio and Tajima’s D test were used to show that the gene’s evolution reflects a mixture of purifying and balancing selection. In all, 186 significant gene conversion events were identified across the full coding region, suggesting that gene conversion has also been an important driver of sequence diversification.

Blast, caused by the ascomycete fungus Magnaporthe oryzae, is one of the most devastating diseases of rice worldwide. The Chinese native cultivar (cv.) Q15 expresses the broad-spectrum resistance to most of the isolates collected from China. To effectively utilize the resistance, three rounds of linkage analysis were performed in an F2 population derived from a cross of Q15 and a susceptible cv. Tsuyuake, which segregated into 3:1 (resistant/susceptible) ratio. The first round of linkage analysis employing simple sequence repeat (SSR) markers was carried out in the F2 population through bulked-segregant assay. A total of 180 SSR markers selected from each chromosome equally were surveyed. The results revealed that only two polymorphic markers, RM247 and RM463, located on chromosome 12, were linked to the resistance (R) gene. To further define the chromosomal location of the R gene locus, the second round of linkage analysis was performed using additional five SSR markers, which located in the region anchored by markers RM247 and RM463. The locus was further mapped to a 0.27 cM region bounded by markers RM27933 and RM27940 in the pericentromeric region towards the short arm. For fine mapping of the R locus, seven new markers were developed in the smaller region for the third round of linkage analysis, based on the reference sequences. The R locus was further mapped to a 0.18 cM region flanked by marker clusters 39M11 and 39M22, which is closest to, but away from the Pita/Pita 2 locus by 0.09 cM. To physically map the locus, all the linked markers were landed on the respective bacterial artificial chromosome clones of the reference cv. Nipponbare. Sequence information of these clones was used to construct a physical map of the locus, in silico, by bioinformatics analysis. The locus was physically defined to an interval of ≈37 kb. To further characterize the R gene, five R genes mapped near the locus, as well as 10 main R genes those might be exploited in the resistance breeding programs, were selected for differential tests with 475 Chinese isolates. The R gene carrier Q15 conveys resistances distinct from those conditioned by the carriers of the 15 R genes. Together, this valuable R gene was, therefore, designated as Pi39(t). The sequence information of the R gene locus could be used for further marker-based selection and cloning.