Development of SNP-based dCAPS markers for identifying male sterile gene tms5 in two-line hybrid rice.

Molecular markers can increase both the efficiency and speed of breeding programs. Functional markers that detect the functional mutations causing phenotypic changes offer a precise method for genetic identification. In this study, we used newly derived cleaved amplified polymorphic sequence markers to detect the functional mutations of tms5, which is a male sterile gene that is widely used in rice production in China. In addition, restriction cutting sites were designed to specifically digest amplicons of tms5 but not wild type (TMS5), in order to avoid the risk of false positive results. By optimizing the condition of the polymerase chain reaction amplifications and restriction enzyme digestions, the newly designed markers could accurately distinguish between tms5 and TMS5. These markers can be applied in marker-assisted selection for breeding novel thermo-sensitive genic male sterile (TGMS) lines, as well as to rapidly identify the TGMS hybrid seed purity.

A novel synthetic Cry1Ab gene resists rice insect pests.

A few insect control genes of Bacillus thuringiensis have been modified successfully to increase the expression in plants by replacing rare codons, increasing GC content, and avoiding the DNA elements that could cause premature transcription termination, mRNA instability, and potential methylation. However, the modification process was intricate and often confused researchers. In this study, we adopted a simple method to modify Cry1Ab only by individually replacing its amino acid sequence with corresponding rice-preferred codons based on analysis of 92,188 coding DNA sequences. Unexpectedly, all elements of A+T richness, which terminate or destabilize transcription in plants, were avoided in the newly designed mCry1Ab. However, mCry1Ab had 2 notable features: less synonymous codons and high GC content. mCry1Ab only employed 22 of the 61 codons to encode protein and had an enhanced GC content of 65%. The increase in GC content caused abundant potential methylation signals to emerge in mCry1Ab. To test whether mCry1Ab could be expressed in rice, we transferred it into Oryza japonica variety Wanjing97. Insect bioassays revealed that transgenic plants harboring this gene driven by 2 promoters, CaMV35S and OsTSP I, were highly resistant to rice leaffolder (Cnaphalocrocis medinalis). Analysis of R0 to R2 generation plants indicated that the mCry1Ab was inherited stably by the progeny. Our study provided a simple modified method for expressing exogenous genes in rice and confirmed that less synonymous codons and high GC content do not affect transgene expression in rice.

Quantitative detection of the rice false smut pathogen Ustilaginoidea virens by real-time PCR.

Rice false smut (RFS) is an important rice disease that is caused by the pathogen Ustilaginoidea virens. In this study, we developed a real-time polymerase chain reaction (PCR) assay to detect U. virens and to estimate the level of disease. The genomic DNAs of U. virens and rice were extracted together from the rice samples. Real-time PCR assays were performed and compared to conventional nested-PCR assays. The real-time PCR assay presented a consistent linearity of the standard curve (R(2) = 0.9999). The detection limit could be as low as 40 fg U. virens DNA with a rice genomic DNA background on using the real-time PCR assay, which showed significantly higher sensitivity than the conventional nested-PCR assay. We conclude that the real-time PCR quantitative assay is a useful tool for detecting U. virens and for early defense and control of RFS.

[Bone metabolic disorders due to antiepileptic drugs and the therapeutic effect of vitamin D2].

Bone mineral content(BMC), serum calcium, phosphorus and AKP were measured in 64 epileptic patients on long-term treatment with antiepileptics and in 14 epileptic patients not taking antiepileptics. All these indices were also measured in a group of healthy controls. In the epileptic patients taking antiepileptics BMC was significantly lower than that in the control group. Serum calcium and phosphorus were also lower than normal, while AKP was elevated. In the epileptic patients not taking antiepileptics BMC was not significantly different from that in the control group. Serum calcium, phosphorus and AKP were all normal. After 3 months of treatment with vitamin D2, BMC and serum calcium level returned to normal, but AKP was still significantly lowered.