PLANT METHODS METHODOLOGY Open Access (original) (raw)
Related papers
Background: TILLING (Targeting Induced Local Lesions IN Genomes) is a reverse genetic method that combines chemical mutagenesis with high-throughput genome-wide screening for point mutation detection in genes of interest. However, this mutation discovery approach faces a particular problem which is how to obtain a mutant population with a sufficiently high mutation density. Furthermore, plant mutagenesis protocols require two successive generations (M1, M2) for mutation fixation to occur before the analysis of the genotype can begin. Results: Here, we describe a new TILLING approach for rice based on ethyl methanesulfonate (EMS) mutagenesis of mature seed-derived calli and direct screening of in vitro regenerated plants. A high mutagenesis rate was obtained (i.e. one mutation in every 451 Kb) when plants were screened for two senescence-related genes. Screening was carried out in 2400 individuals from a mutant population of 6912. Seven sense change mutations out of 15 point mutations were identified.
Discovery of chemically induced mutations in rice by TILLING
BMC Plant Biology, 2007
Background: Rice is both a food source for a majority of the world's population and an important model system. Available functional genomics resources include targeted insertion mutagenesis and transgenic tools. While these can be powerful, a non-transgenic, unbiased targeted mutagenesis method that can generate a range of allele types would add considerably to the analysis of the rice genome. TILLING (Targeting Induced Local Lesions in Genomes), a general reverse genetic technique that combines traditional mutagenesis with high throughput methods for mutation discovery, is such a method.
EMS Induced Mutants of Upland Rice Variety Nagina22: Generation and Characterization
Proceedings of the Indian National Science Academy, 2014
One of the approaches for understanding the functions of the genes predicted in the rice genome requires use of mutants. Mutations induced in a given genetic background provide opportunities to assign function to a given gene with a minimum of background genetic noise. This paper describes generation and initial characterization of a large set of Ethyl Methane Sulphonate (EMS) induced mutants in the upland rice variety Nagina22, through a National Initiative involving six
Ds insertion mutagenesis as an efficient tool to produce diverse variations for rice breeding
Plant Molecular Biology, 2007
The availability of diversified germplasm resources is the most important for developing improved rice varieties with higher seed yield or tolerance to various biotic or abiotic stresses. Here we report an efficient tool to create increased variations in rice by maize Ac/Ds transposon (a gene trap system) insertion mutagenesis. We have generated around 20,000 Ds insertion rice lines of which majority are homozygous for Ds element. We subjected these lines to phenotypic and abiotic stress screens and evaluated these lines with respect to their seed yields and other agronomic traits as well as their tolerance to drought, salinity and cold. Based on this evaluation, we observed that random Ds insertions into rice genome have led to diverse variations including a range of morphological and conditional phenotypes. Such differences in phenotype among these lines were accompanied by differential gene expression revealed by GUS histochemical staining of gene trapped lines. Among the various phenotypes identified, some Ds lines showed significantly higher grain yield compared to wild-type plants under normal growth conditions indicating that rice could be improved in grain yield by disrupting certain endogenous genes. In addition, several 1,000s of Ds lines were subjected to abiotic stresses to identify conditional mutants. Subsequent to these screens, over 800 lines responsive to drought, salinity or cold stress were obtained, suggesting that rice has the genetic potential to survive under abiotic stresses when appropriate endogenous genes were suppressed. The mutant lines that have higher seed yielding potential or display higher tolerance to abiotic stresses may be used for rice breeding by conventional backcrossing combining with molecular marker-assisted selection. In addition, by exploiting the behavior of Ds to leave footprints upon remobilization, we have shown an alternative strategy to develop new rice varieties without foreign DNA sequences in their genome.
Protocol for ethyl methanesulphonate (EMS) mutagenesis application in rice
Open Research Europe, 2021
Background: Non-transgenic chemical mutagen application, particularly ethyl methanesulfonate (EMS), is an important tool to create mutations and gain a new genetic makeup for plants. It is useful to obtain a sufficient number of mutant plants instead of working with a severe mutation in a few plants. EMS dose and exposure period have been previously studied in several crops; however, EMS used to create point mutations in presoaked rice seeds has not been sufficiently studied and there is no standard protocol for such treatment. The aim of this study is to establish a standard protocol for EMS mutagenesis application in rice. Methods: Two studies were conducted to evaluate the effect of four durations of rice seed presoaking (0, 6, 12, and 24 hours), four EMS concentration doses (0.0%, 0.5%, 1.0%, and 2.0%), and four EMS exposure periods (6, 12, 24, and 48 hours). Germination rate, plumula and radicle length, seedling survival, LD50 (Lethal Dose) determination, shoot length, root len...
Gneration of rice EMS mutants in Nagina22
One of the approaches for understanding the functions of the genes predicted in the rice genome requires use of mutants. Mutations induced in a given genetic background provide opportunities to assign function to a given gene with a minimum of background genetic noise. This paper describes generation and initial characterization of a large set of Ethyl Methane Sulphonate (EMS) induced mutants in the upland rice variety Nagina22, through , funded by the Department of Biotechnology (DBT), Government of India. The uniqueness of this collaborative effort is phenotyping for a range of traits that has led to identification of mutants for plant growth and architecture, flowering, maturity, grain number, shape and size, yield, phosphorus use efficiency, resistance to blast and bacterial leaf blight diseases, and tolerance to drought, salinity and herbicide. A set of 22, 292 mutagenised lines generated under this initiative and phenotyped for the traits enlisted above has resulted in the isolation of a few promising mutants which are being characterized. Shortly, these mutants will be registered and made available to the researchers in the country for use in studies in rice genetics, breeding and functional genomics. The mutant stock is expected to serve as a national resource for understanding rice biology as well as for use in genetic improvement of the crop.
Rice agronomic traits and variability induced by mutagenesis
2021
Understanding agronomic traits at a genetic level enables the leveraging of this knowledge to produce crops that are more productive and resilient, have better quality and are adjusted for consumer preferences. In the last decade, rice has become a model to validate the function of specific genes, resulting in valuable but scattered information. Here, we aimed to identify particular genes in rice related to traits that can be targeted by different mutation techniques in the breeding of crops. We selected gain of function, misfunction, and specific mutations associated with phenotypes of agronomic interest. The review includes specific trait-related genes involved in domestication, stress, herbicide tolerance, pathogen resistance, grain number, quality, weight, plant structure, nitrogen use, and others. The information presented can be used for rice and crops with similar or homologous genes to breed crops that require improvement to achieve more sustainable production in challenging...
DBT propelled national effort in creating EMS mutant resource in rice forfunctional genomics
In 2007, with the help of DBT, a research project to create mutant resources for functional genomics in rice was launched through a national initiative involving ICAR Genetically well-defined material is a prerequisite for functional genomics. Thus, the project aimed to generate EMS mutants in the background of an upland and short duration aus genotype, Nagina22, characterize the mutants and use them in crop improvement. As of now, nearly 85,000 rice M2 mutant populations have been created under the project. Based on field phenotyping, gain and or loss of function mutants for tolerance to herbicide spray, drought, salinity and resistance to rice leaf and panicle blast, sheath blight and high phosphorus (P) use efficiency under low P field have been identified. Notably, the herbicide-tolerant mutant identified is under the process of registration for distribution to public and private rice breeders under appropriate material transfer agreement. Besides this, the project also aims to serve as a 'National Repository of rice EMS mutant resource' for the researchers involved in rice biology and improvement in the country.
International Consortium of Rice Mutagenesis: resources and beyond
Rice, 2013
Rice is one of the most important crops in the world. The rice community needs to cooperate and share efforts and resources so that we can understand the functions of rice genes, especially those with a role in important agronomical traits, for application in agricultural production. Mutation is a major source of genetic variation that can be used for studying gene function. We will present here the status of mutant collections affected in a random manner by physical/chemical and insertion mutageneses. As of early September 2013, a total of 447, 919 flanking sequence tags from rice mutant libraries with T-DNA, Ac/Ds, En/Spm, Tos17, nDART/aDART insertions have been collected and publicly available. From these, 336,262 sequences are precisely positioned on the japonica rice chromosomes, and 67.5% are in gene interval. We discuss the genome coverage and preference of the insertion, issues limiting the exchange and use of the current collections, as well as new and improved resources. We propose a call to renew all mutant populations as soon as possible. We also suggest that a common web portal should be established for ordering seeds.
Insertional Mutagenesis Approaches and Their Use in Rice for Functional Genomics
Plants
Insertional mutagenesis is an indispensable tool for engendering a mutant population using exogenous DNA as the mutagen. The advancement in the next-generation sequencing platform has allowed for faster screening and analysis of generated mutated populations. Rice is a major staple crop for more than half of the world’s population; however, the functions of most of the genes in its genome are yet to be analyzed. Various mutant populations represent extremely valuable resources in order to achieve this goal. Here, we have reviewed different insertional mutagenesis approaches that have been used in rice, and have discussed their principles, strengths, and limitations. Comparisons between transfer DNA (T-DNA), transposons, and entrapment tagging approaches have highlighted their utilization in functional genomics studies in rice. We have also summarised different forward and reverse genetics approaches used for screening of insertional mutant populations. Furthermore, we have compiled ...