Expression patterns of microRNAs in different organs and developmental stages of a superhybrid rice LYP9 and its parental lines

Heterosis is an important phenomenon, and the molecular mechanisms underlying heterosis are still enigmatic. microRNAs (miRNAs) play vital roles in many aspects of plant development. A set of miRNAs was selected to investigate the roles of miRNAs in heterosis displayed in a superhybrid rice. We analysed the expression patterns of miRNAs in different organs and developmental stages of the superhybrid rice and its parental lines. All possible modes of miRNA action were observed, including additive, high‐ and low‐parent value, above high‐ and below low‐parent value. Different organs and developmental stages exhibited different modes of miRNA expression. Overall, the non‐additive mode is the predominant expression pattern of miRNAs observed in this superhybrid. Many heterotic QTL intervals harbour miRNAs, whose expression patterns reveal their specific roles in different organs and developmental stages. miRNAs regulate the expression levels of target genes that have important functions in plant development. The predominant non‐additive mode of miRNA expression pattern in the hybrid suggests that miRNAs play critical roles in hybrid development, in particular, those miRNAs located in the heterotic QTL intervals may have important roles in heterosis. Our research sheds new light on understanding of the molecular mechanisms of heterosis.     

Differential gene expression in salt-tolerant rice mutant and its parental variety

The differential expressions of three genes rbcL, salT and rab!6 in response to ABA, NaCl, PEG and heat shock were investigated in seedlings of a salt-tolerant rice mutant 20 (mutant 20) and its parental variety Oryza sativa var. japonica 77-170(170). By Northern blot analysis it was found that ABA induced the expression of all three genes of rbcL, salT and rab16 in shoots and roots of both 170 and mutant 20 with the exceptions of rab16 in shoots of mutant 20 and rbcL in roots of 170. Lower concentrations of NaCl induced rbcL expression in shoots of mutant 20 but not 170. Higher concentrations of NaCl decreased rbcL expression but induced expressions of salT and rab16 in shoots of both 170 and mutant 20. PEG(15%) and 37℃ heat shock showed almost no effects on the expression of the three genes in mutant 20. However, they caused a decrease in rbcL expression and slight induction of the rab16 gene in 170, with salT expression unaffected. These results indicated that mutant 20 was relatively less responsiv     

Spatial and temporal expression modes of MicroRNAs in an elite rice hybrid and its parental lines

Heterosis is a commonly observed phenomenon in nature and refers to the superior performance of hybrids relative to both parents. The molecular mechanisms of heterosis are mostly unknown. Quantitative trait locus (QTL) mapping has been used to explain the genetic basis of heterosis, and large amounts of QTLs have been mapped for various agronomic traits, but the nature of QTL contributing to heterosis is still enigmatic. MicroRNAs (miRNAs) are master regulators in the processes of plant development and trait performance, and many miRNAs are predicted to reside in QTL intervals. We analyzed the expression modes of miRNAs, which were picked up from miRNA databases and chosen from those predicted from QTL intervals by bioinformatic approaches, in different organs at developmental stages of an elite rice hybrid and its parents. All possible modes of action for miRNA expression were detected, but most miRNAs showed nonadditive mode, and different stages and distinct organs displayed different patterns of miRNA expression. A large proportion of miRNAs were not detected for expression in leaves but expressed in the culms and roots of the hybrid at tillering stage. MiRNAs from grain-weight QTL intervals have multiple effects on grain development. Together, our results reveal that miRNAs, especially those from QTL intervals, play roles in heterotic performance in this elite rice hybrid, our results also shade new light on understanding the molecular mechanisms of heterosis.     

草菇耐低温突变菌株Vtlt-1和原始菌株V23转录组差异的研究

为了培育草菇耐低温菌株与解析其耐低温的分子机制,采用紫外诱变的方法,选育出耐低温草菇菌株Vtlt-1,并利用表达谱芯片技术,比较突变菌株Vtlt-1与原始菌株V23的表达差异基因,筛选差异显著基因后利用GO（gene ontology）功能注释和KEGG（kyoto encyclopedia of genes and genomes）通路富集等方法分析,结果发现与V23相比Vtlt-1表达显著差异基因共有1 600个,其中704个基因上调表达,896个基因下调表达。针对差异基因的GO功能分类结果发现：生物学过程方面,差异基因主要分布在金属离子结合、氧化还原过程、碳水化合物的代谢与核酸的结合。细胞组分方面主要与细胞核相关;分子功能中,氧化还原活性以及解旋酶活性,依赖于ATP 的解旋酶活性、DNA指导的RNA聚合酶活性。KEGG注释结果发现差异表达基因主要富集在氨基酸与氮类物质的代谢、脂肪酸与生物碱的合成这两方面,此外还富集到核糖体的生物合成、细胞色素P450、RNA聚合酶、硫和氨基酸的代谢、核酸的修复等通路。这些结果为解析草菇耐低温机制提供分子依据和理论基础。     

Serial analysis of gene expression (SAGE): unraveling the bioinformatics tools

Serial analysis of gene expression (SAGE) is a powerful technique that can be used for global analysis of gene expression. Its chief advantage over other methods is that it does not require prior knowledge of the genes of interest and provides qualitative and quantitative data of potentially every transcribed sequence in a particular cell or tissue type. This is a technique of expression profiling, which permits simultaneous, comparative and quantitative analysis of gene-specific, 9- to 13-basepair sequences. These short sequences, called SAGE tags, are linked together for efficient sequencing. The sequencing data are then analyzed to identify each gene expressed in the cell and the levels at which each gene is expressed. The main benefit of SAGE includes the digital output and the identification of novel genes. In this review, we present an outline of the method, various bioinformatics methods for data analysis and general applications of this important technology.     

Characterization of a rice pollen-specific gene and its expression

Pollen development requires a large number of genes expressed in both sporophytic and gametophytic tissues. We have isolated a pollen-specific gene, PS1, from rice. PS1 is a unique gene in the rice genome and encodes a 164 amino acid long protein. RNA blot analysis shows that PS1 mRNAs accumulate specifically in rice anthers. When introduced into rice tissues by microprojectile bombardment, the PS1 promoter drives expression of a marker gene, β-glucuronidase, specifically in rice pollen. The PS1 gene and the deduced amino acid sequence of the PS1 protein share significant levels of homology with another monocot pollen-specific gene—the maize Zm-13 gene and its deduced protein, respectively. PS1 also shows some homology with the dicot tomato anther-specific gene LAT-52. Interestingly, the structure of the PS1 gene is more similar to that of the LAT-52 gene than to Zm-13. The coding regions of both PS1 and LAT-52 are interrupted by a single intron, and the positions of the introns are conserved in these genes. Moreover, there is considerable sequence homology in the introns of the PS1 and LAT-52 genes in regions immediately upstream of the 3' splice sites. The upstream regulatory sequences of the PS1 gene show several regions of homology with other pollen- or anther-specific genes from a number of plant species. The conservation of coding sequences of PS1 from rice, Zm-I3 from maize, and LAT-52 from tomato suggests a functional conservation of their gene products. Similarities in the regulatory regions of PS1 and other anther- or pollen-specific genes among monocotyledonous and dicotyledonous species indicate that at least some regulatory features controlling gene expression in male reproductive tissues are conserved. This is supported by the preservation of pollen-specific expression from the rice PS1 promoter when it is introduced into tobacco plants by Agrobacterium Ti plasmid-mediated transformation.     

Comparative proteomic study reveals dynamic proteome changes between superhybrid rice LYP9 and its parents at different developmental stages

Heterosis is a common phenomenon in which the hybrids exhibit superior agronomic performance than either inbred parental lines. Although hybrid rice is one of the most successful apotheoses in crops utilizing heterosis, the molecular mechanisms underlying rice heterosis remain elusive. To gain a better understanding of the molecular mechanisms of rice heterosis, comparative leaf proteomic analysis between a superhybrid rice LYP9 and its parental cultivars 9311 and PA64s at tillering, flowering and grain-filling stages were carried out. A total of 384 differentially expressed proteins (DP) were detected and 297 DP were identified, corresponding to 222 unique proteins. As DP were divided into those between the parents (DP_PP) and between the hybrid and its parents (DP_HP), the comparative results demonstrate that proteins in the categories of photosynthesis, glycolysis, and disease/defense were mainly enriched in DP. Moreover, the number of identified DP_HP involved in photosynthesis, glycolysis, and disease/defense increased at flowering and grain-filling stages as compared to that at the tillering stage. Most of the up-regulated DP_HP involved in the three categories showed greater expression in LYP9 at flowering and grain-filling stages than at the tillering stage. In addition, CO₂ assimilation rate and apparent quantum yield of photosynthesis also showed a greater increase in LYP9 at flowering and grain-filling stages than at the tillering stage. These results suggest that the proteins involved in photosynthesis, glycolysis, and disease/defense as well as their dynamic regulation at different developmental stages may be responsible for heterosis in rice.     

Characteristics of chlorophyll fluorescence and antioxidative system in super-hybrid rice and its parental cultivars under chilling stress

Characteristics of chlorophyll fluorescence and antioxidative system were investigated in rice (Oryza sativa L.) super-hybrid Liangyoupeijiu (LYPJ), maternal cultivar Peiai64s, and paternal cultivar indica rice 9311 under chilling stress. During 6-d chilling treatment, chlorophyll content of all three genotypes was gradually declined. However, the decrease in photosystem 2 (PS 2) maximum photochemical efficiency (F_v/F_m) and quantum yield of PS 2 (Φ_PS2) was less expressive in LYPJ than in parental cultivars The electrolyte leakage and malondialdehyde content in all cultivars increased after chilling treatment, but LYPJ exhibited the least increasing tendency. Activities of superoxide dismutase (SOD), ascorbate peroxidase (APX) and glutathione reductase (GR) were higher in LYPJ than in parental cultivars. The results demonstrated that tolerance to chilling stress in LYPJ might be adopted mostly from its maternal cultivar.     

Identification and expression analysis of cobia (Rachycentron canadum) Toll-like receptor 9 gene

Cobia culture is hindered by bacterial infection (Photobacterium damselae subsp. piscicida) and in order to study the effect of P. damselae subsp. piscicida challenge and CpG ODN stimulation on cobia Toll like receptor 9 (RCTLR9), we used PCR to clone RCTLR9 gene and qRT-PCR to quantify gene expression. The results indicated that RCTLR9 cDNA contains 3141 bp. It encodes 1047 amino acids containing 16 typical structures of leucine-rich repeats (LRRs) including an LRRTYP, LRRCT and a motif involved in PAMP binding was identified at position 240–253 amino acid. Broad expression of RCTLR9 was found in larval, juvenile and adult stages irrespective of the tissues. In larval stage, RCTLR9 mRNA expression decreased at 5 d and then increased at 10 dph. At juvenile stage cobia, the expression was significantly high (p < 0.05) in spleen and intestine compared to gill, kidney, liver and skin. However, at adult stage, the significant high expression was found in gill and intestine. Cobia challenged with P. damselae subsp. piscicida showed significant increase in RCTLR9 expression at 24 h post challenge in intestine, spleen and liver, while in kidney the expression was peak at 12 h and later it decreased at 24 h. The highest expression was 40 fold increase in spleen and the lowest expression was ∼3.6 fold increase in liver. Cobia stimulated with CpG oligonucleotides showed that the induction of these genes was CpG ODN type and time dependent. In spleen and liver, CpG ODNs 1668 and 2006 injected group showed high expression of RCTLR9, IL-1β, chemokine CC compared to other groups. Meanwhile, CpG ODN 2006 has induced high expression of IgM. The CpG ODNs 2395 have induced significant high expression of Mx in spleen and liver. These results demonstrates the potential of using CpG ODN to enhance cobia resistance to P. damselae subsp. piscicida infection and use as an adjuvant in vaccine development.