Heterologous expression of a Glyoxalase I gene from sugarcane confers tolerance to several environmental stresses in bacteria

Glyoxalase I belongs to the glyoxalase system that detoxifies methylglyoxal (MG), a cytotoxic by-product produced mainly from triose phosphates. The concentration of MG increases rapidly under stress conditions. In this study, a novel glyoxalase I gene, designated as SoGloI was identified from sugarcane. SoGloI had a size of 1,091 bp with one open reading frame (ORF) of 885 bp encoding a protein of 294 amino acids. SoGloI was predicted as a Ni2+-dependent GLOI protein with two typical glyoxalase domains at positions 28–149 and 159–283, respectively. SoGloI was cloned into an expression plasmid vector, and the Trx-His-S-tag SoGloI protein produced in Escherichia coli was about 51 kDa. The recombinant E. coli cells expressing SoGloI compared to the control grew faster and tolerated higher concentrations of NaCl, CuCl2, CdCl2, or ZnSO4. SoGloI ubiquitously expressed in various sugarcane tissues. The expression was up-regulated under the treatments of NaCl, CuCl2, CdCl2, ZnSO4 and abscisic acid (ABA), or under simulated biotic stress conditions upon exposure to salicylic acid (SA) and methyl jasmonate (MeJA). SoGloI activity steadily increased when sugarcane was subjected to NaCl, CuCl2, CdCl2, or ZnSO4 treatments. Sub-cellular observations indicated that the SoGloI protein was located in both cytosol and nucleus. These results suggest that the SoGloI gene may play an important role in sugarcane’s response to various biotic and abiotic stresses

Effects of long-term flutamide treatment during development in zebra finches

The molecular mechanisms responsible for the sexual differentiation of the zebra finch song system remain mysterious. Androgen receptors are expressed in a sexually dimorphic fashion in the zebra finch song system: males have more cells expressing androgen receptors, and this sex difference appears very early in development (day 9 posthatch). Estrogen administration to hatchling females up-regulates androgen receptor expression in their song system and profoundly masculinizes their song system's morphology. Co-administering flutamide, an androgen receptor blocker, with estrogen impedes estrogen's masculinizing effects on the song system, suggesting that androgens are required for masculine development. Accordingly, to investigate further the role of androgens in the sexual differentiation of the zebra finch song system, we sought to block androgen activity in males by administering large, sustained doses of flutamide from just before androgen receptors are expressed in the song system (day 7) through to the day of sacrifice (days 61-63). Flutamide profoundly reduced the size of the testes, demonstrating that this drug and mode of administration could have a large impact on tissues. In contrast, flutamide had only a minor impact on the song system: the number of RA neurons was slightly reduced, and the corrected HVC volume showed a trend toward demasculinization. Other brain measures (uncorrected HVC, and corrected and uncorrected volumes of Area X, lMAN, RA, and Rotundus; neuron size in lMAN, HVC, and RA; and number of HVC and LMAN neurons) were not significantly affected. The present results do not support an important role for androgen in masculinizing the song circuit after posthatch day 7. (c) 2007 Elsevier Ireland Ltd. All rights reserved

Effects of long-term flutamide treatment during development in zebra finches

The molecular mechanisms responsible for the sexual differentiation of the zebra finch song system remain mysterious. Androgen receptors are expressed in a sexually dimorphic fashion in the zebra finch song system: males have more cells expressing androgen receptors, and this sex difference appears very early in development (day 9 posthatch). Estrogen administration to hatchling females up-regulates androgen receptor expression in their song system and profoundly masculinizes their song system's morphology. Co-administering flutamide, an androgen receptor blocker, with estrogen impedes estrogen's masculinizing effects on the song system, suggesting that androgens are required for masculine development. Accordingly, to investigate further the role of androgens in the sexual differentiation of the zebra finch song system, we sought to block androgen activity in males by administering large, sustained doses of flutamide from just before androgen receptors are expressed in the song system (day 7) through to the day of sacrifice (days 61-63). Flutamide profoundly reduced the size of the testes, demonstrating that this drug and mode of administration could have a large impact on tissues. In contrast, flutamide had only a minor impact on the song system: the number of RA neurons was slightly reduced, and the corrected HVC volume showed a trend toward demasculinization. Other brain measures (uncorrected HVC, and corrected and uncorrected volumes of Area X, lMAN, RA, and Rotundus; neuron size in lMAN, HVC, and RA; and number of HVC and LMAN neurons) were not significantly affected. The present results do not support an important role for androgen in masculinizing the song circuit after posthatch day 7. (c) 2007 Elsevier Ireland Ltd. All rights reserved

A Comparative Study of Three Detection Techniques for Leifsonia xyli Subsp. xyli, the Causal Pathogen of Sugarcane Ratoon Stunting Disease

The ratoon stunting disease (RSD), caused by the bacterium Leifsonia xyli subsp. xyli (Lxx), is one of the most economically devastating diseases impacting sugarcane. RSD causes significant yield losses and variety degradation. Diagnosis of RSD is challenging because it does not exhibit any discernible internal and external symptoms. Moreover, the Lxx bacteria are very small and difficult to isolate, cultivate, and detect. In this study, conventional polymerase chain reaction (PCR), real-time quantitative PCR (RT-qPCR), and Lxx-loop-mediated isothermal amplification (Lxx-LAMP) were utilized to specifically detect the presence of Lxx pathogens in the juice from Lxx-infected sugarcane stalks and an Lxx-pMD18-T recombinant plasmid. The results showed that Lxx was a highly specific causal pathogen for RSD. All three techniques provided great reproducibility, while Lxx-LAMP had the highest sensitivity. When the DNA extract from Lxx-infected sugarcane juice was used as a template, Lxx-LAMP was 10 and 100 times more sensitive than RT-qPCR and conventional PCR, respectively. When the Lxx-pMD18-T recombinant plasmid was used as a template, Lxx-LAMP was as sensitive as RT-qPCR but was 10 times more sensitive than conventional PCR. Based on the Lxx-LAMP detection system established, adding 0.4 μM loop primers (LF/LP) can accelerate the reaction and reduce the total time required. In addition, the optimal amount of Bst DNA polymerase for Lxx-LAMP reactions was determined to be 6.0 U. The results provide technical support for the detection of RSD Lxx pathogen that will help manage sugarcane RSD

WGCNA Identifies a Comprehensive and Dynamic Gene Co-Expression Network That Associates with Smut Resistance in Sugarcane

Sugarcane smut is a major fungal disease caused by Sporisorium scitamineum, which seriously reduces the yield and quality of sugarcane. In this study, 36 transcriptome data were collected from two sugarcane genotypes, YT93-159 (resistant) and ROC22 (susceptible) upon S. scitamineum infection. Data analysis revealed 20,273 (12,659 up-regulated and 7614 down-regulated) and 11,897 (7806 up-regulated and 4091 down-regulated) differentially expressed genes (DEGs) in YT93-159 and ROC22, respectively. A co-expression network was then constructed by weighted gene co-expression network analysis (WGCNA), which identified 5010 DEGs in 15 co-expressed gene modules. Four of the 15 modules, namely, Skyblue, Salmon, Darkorange, and Grey60, were significantly associated with smut resistance. The GO and KEGG enrichment analyses indicated that the DEGs involving in these four modules could be enriched in stress-related metabolic pathways, such as MAPK and hormone signal transduction, plant-pathogen interaction, amino acid metabolism, glutathione metabolism, and flavonoid, and phenylpropanoid biosynthesis. In total, 38 hub genes, including six from the Skyblue module, four from the Salmon module, 12 from the Darkorange module, and 16 from the Grey60 module, were screened as candidate hub genes by calculating gene connectivity in the corresponding network. Only 30 hub genes were amplifiable with RT-qPCR, of which 27 were up-regulated upon S. scitamineum infection. The results were consistent with the trend of gene expression in RNA-Seq, suggesting their positive roles in smut resistance. Interestingly, the expression levels of AOX, Cyb5, and LAC were higher in ROC22 than in YT93-159, indicating these three genes may act as negative regulators in response to S. scitamineum infection. This study revealed the transcriptome dynamics in sugarcane challenged by S. scitamineum infection and provided gene targets for smut resistance breeding in sugarcane