水稻突变体与功能基因组学

突变体是功能基因组学研究的重要材料。本文综述了水稻突变体的创制方法、变异的机制、水稻突变体,基因分类和数量、克隆的水稻重要基因的研究进展,包括1698个水稻突变体,基因的分类和43个克隆的水稻突变体基因,并提出了利用水稻突变体的展望。     

Genetic resources offer efficient tools for rice functional genomics research

Rice is an important crop and major model plant for monocot functional genomics studies. With the establishment of various genetic resources for rice genomics, the next challenge is to systematically assign functions to predicted genes in the rice genome. Compared with the robustness of genome sequencing and bioinformatics techniques, progress in understanding the function of rice genes has lagged, hampering the utilization of rice genes for cereal crop improvement. The use of transfer DNA (T‐DNA) insertional mutagenesis offers the advantage of uniform distribution throughout the rice genome, but preferentially in gene‐rich regions, resulting in direct gene knockout or activation of genes within 20–30 kb up‐ and downstream of the T‐DNA insertion site and high gene tagging efficiency. Here, we summarize the recent progress in functional genomics using the T‐DNA‐tagged rice mutant population. We also discuss important features of T‐DNA activation‐ and knockout‐tagging and promoter‐trapping of the rice genome in relation to mutant and candidate gene characterizations and how to more efficiently utilize rice mutant populations and datasets for high‐throughput functional genomics and phenomics studies by forward and reverse genetics approaches. These studies may facilitate the translation of rice functional genomics research to improvements of rice and other cereal crops.     

Contribution of the Tos17 retrotransposon to rice functional genomics

The ongoing international efforts of the Rice Genomic Sequencing Project have already generated a large amount of sequence data. The next important challenge will be to construct saturation mutant lines for the functional analysis of all of the genes revealed by this effort in the context of the rice plant as a whole. Recently, the endogenous retrotransposon Tos17 has been shown to be an efficient insertional mutagen. Considering the ease of mutagenesis with Tos17 and its multiple-copy nature, saturation mutagenesis with this retrotransposon should be feasible in rice. Ongoing reverse-genetics studies, such as the PCR-screening of mutants and cataloguing of mutants by sequencing Tos17-insertion sites, as well as traditional forward-genetics studies, have clearly demonstrated that the Tos17 system can significantly contribute to the functional genomics of rice.     

Identification and functional analysis of light-responsive unique genes and gene family members in rice

Functional redundancy limits detailed analysis of genes in many organisms. Here, we report a method to efficiently overcome this obstacle by combining gene expression data with analysis of gene-indexed mutants. Using a rice NSF45K oligo-microarray to compare 2-week-old light- and dark-grown rice leaf tissue, we identified 365 genes that showed significant 8-fold or greater induction in the light relative to dark conditions. We then screened collections of rice T-DNA insertional mutants to identify rice lines with mutations in the strongly light-induced genes. From this analysis, we identified 74 different lines comprising two independent mutant lines for each of 37 light-induced genes. This list was further refined by mining gene expression data to exclude genes that had potential functional redundancy due to co-expressed family members (12 genes) and genes that had inconsistent light responses across other publicly available microarray datasets (five genes). We next characterized the phenotypes of rice lines carrying mutations in ten of the remaining candidate genes and then carried out co-expression analysis associated with these genes. This analysis effectively provided candidate functions for two genes of previously unknown function and for one gene not directly linked to the tested biochemical pathways. These data demonstrate the efficiency of combining gene family-based expression profiles with analyses of insertional mutants to identify novel genes and their functions, even among members of multi-gene families.     

Rice functional genomics research in China

Rice functional genomics is a scientific approach that seeks to identify and define the function of rice genes, and uncover when and how genes work together to produce phenotypic traits. Rapid progress in rice genome sequencing has facilitated research in rice functional genomics in China. The Ministry of Science and Technology of China has funded two major rice functional genomics research programmes for building up the infrastructures of the functional genomics study such as developing rice functional genomics tools and resources. The programmes were also aimed at cloning and functional analyses of a number of genes controlling important agronomic traits from rice. National and international collaborations on rice functional genomics study are accelerating rice gene discovery and application.     

Splicing and alternative splicing in rice and humans

Rice is a monocot gramineous crop, and one of the most important staple foods. Rice is considered a model species for most gramineous crops. Extensive research on rice has provided critical guidance for other crops, such as maize and wheat. In recent years, climate change and exacerbated soil degradation have resulted in a variety of abiotic stresses, such as greenhouse effects, lower temperatures, drought, floods, soil salinization and heavy metal pollution. As such, there is an extremely high demand for additional research, in order to address these negative factors. Studies have shown that the alternative splicing of many genes in rice is affected by stress conditions, suggesting that manipulation of the alternative splicing of specific genes may be an effective approach for rice to adapt to abiotic stress. With the advancement of microarrays, and more recently, next generation sequencing technology, several studies have shown that more than half of the genes in the rice genome undergo alternative splicing. This mini-review summarizes the latest progress in the research of splicing and alternative splicing in rice, compared to splicing in humans. Furthermore, we discuss how additional studies may change the landscape of investigation of rice functional genomics and genetically improved rice. [BMB Reports 2013; 46(9): 439-447]     

Simple RNAi vectors for stable and transient suppression of gene function in rice

Since the recent sequencing of the rice genome, the functional identification of rice genes has become increasingly important. Various tagged lines have been generated; however, the number of tagged genes available is not sufficient for extensive study of gene function. To help identify the functions of genes in rice, we developed a Gateway vector, pANDA, for RNA interference of rice genes. This vector can be used for Agrobacterium transformation of rice and allows easy and fast construction of efficient RNAi vectors. In the construct, hairpin RNA derived from a given gene is transcribed from a strong maize ubiquitin promoter, and an intron is placed 5' upstream of inverted repeats to enhance RNA expression. Analysis of rice genes using this vector showed that suppression of mRNA expression was observed in more than 90% of transgenic plants examined, and short interfering RNA indicative of RNA silencing was detected in each silenced plant. A similar vector, pANDA-mini, was also developed for direct transfer into leaf cells or protoplasts. This vector can be used for transient suppression of gene function in rice. These vectors should help identify the functions of rice genes whose tagged mutants are not available at present and complement existing methods for functional genomics of rice.