Genetics for Rice Improvement, 2017

 

Project Leader

Tom Tai, research geneticist, USDA-ARS Crops Pathology and Genetics Research Unit, Dept. of Plant Sciences, UC Davis

The overall goal of this project is to employ traditional and advanced genetic screens to identify and characterize novel rice germplasm to advance the understanding of agronomic performance and grain quality and for incorporation into breeding programs for the California rice industry.

Primary emphasis is on screening rice populations generated by traditional mutagenesis for new traits that improve grain quality and reduce production costs. This is achieved by directly screening plant materials for traits of interest and by identifying changes in the DNA sequence of genes that may result in the expression of these traits. Specific targets currently include reduced uptake and/or localization of arsenic in milled rice grains and resistance or tolerance of rice plants to selected herbicides.

The traditional approach to mutant populations is to conduct screens that identify mutant phenotypes of interest. Examples of this “forward” genetics approach in rice include the identification of the semidwarf trait, conditional male sterility, and herbicide tolerance.

Reverse genetics is a complementary approach for exploiting mutant populations. This strategy requires prior knowledge of the genes involved in traits of interest. These genes are used to screen populations to identify mutated versions that may result in the expression of novel traits.

One method of reverse genetics is the Targeting of Induced Local Lesions in Genomes (TILLING) strategy. It is based on the detection of mutations in target gene sequences by screening DNA isolated and pooled from hundreds of mutant lines (typically 2,000 lines total) generated by traditional chemical mutagenesis. A service to identify mutations of interest with this technique is operated by the TILLING Core Lab at the UC Davis Genome Center. Using this service, researchers have successfully identified mutations in genes involved in arsenic uptake and accumulation and genes that encode protein targets of various herbicides.

The major objectives of research in 2017 were:

  • Characterization of rice mutants identified by forward and reverse genetic screens.
  • Confirmation and evaluation of additional mutants identified by reverse genetic screens of genes encoding proteins targeted by selected herbicides and genes that control arsenic uptake and accumulation.

Work on this project was conducted in the USDA-ARS rice genetics lab, greenhouses, and other research facilities at UC Davis. Research efforts were disrupted by the required relocation of the rice genetics laboratory. Consequently, some work was not completed and some funding was rolled over to 2018. Research highlights from 2017 work is summarized below.

Arsenic is known to enter rice plants through the same pathway used by silicon. Genetic screening can identify mutants exhibiting altered uptake and accumulation of silicon based on resistance to the element germanium. Because of similar chemical properties, germanium is also taken up the same way as silicon, but it is toxic to rice plants and results in the formation of lesions and eventually death in seedlings.

Previously, several lines carrying mutations in genes involved in silicon and arsenic uptake or accumulation were identified by TILLING and confirmed using DNA sequencing. Seeds were harvested from mutants harboring fixed mutations (important for trait expression). Forward genetic screening using germanium also resulted in the identification of several mutant lines that are potentially altered in the ability to take up silicon and arsenic.

In 2017, nine mutant lines carrying fixed mutations in the genes targeted in this study were grown in the field to provide tissue samples for analysis of silicon and total arsenic (organic and inorganic arsenic) content. Evaluation of these mutants using a germanium assay revealed that two of three mutations detected in the Lsi1 gene appeared to confer an increased tolerance to germanium, suggesting these lines may have reduced silicon and arsenic in their tissues. Nine lines carrying mutations in Lsi2 and one carrying an OsABCC1 mutation did not appear to be qualitatively different in response to germanium when compared to wild-type Nipponbare rice.

Preliminary elemental analysis of mutant lines grown in the UC Davis rice field facility in 2017 will be conducted in 2018 to determine the silicon and total arsenic content in the aboveground tissues. Screening of another set of 2,048 mutants using the TILLING approach identified a number of new mutations in several of the arsenic uptake/accumulation genes. These mutations are in various stages of confirmation and identification of fixed mutant lines.

Microtubules are proteins that play an important role in cell growth and development. The chemical pendimethalin (active ingredient of the herbicide Prowl®) inhibits microtubules by interacting with a component called alpha-2 tubulin, which then disrupts cell division and normal growth. Seven mutant lines harboring different mutations in the gene encoding alpha-2 tubulin were evaluated for their response to pendimethalin. All the lines exhibited more sensitivity than the wild type. This unexpected result is being investigated by cooperators at UC Davis, who are examining the growth and development of these mutants in more detail. Mutants carrying mutations in other genes encoding herbicide targets will be evaluated for their response to various herbicides in 2018.

The TILLING screening work has concluded and future efforts will focus on mutant trait evaluation and generating specific mutations in genes of interest through targeted mutagenesis by gene editing.