Identification and utilization of cleistogamy gene cl7(t) in rice (Oryza sativa L.).

Gene transformation is an important method for improvement of plants into elite varieties. However, the possibility of gene flow between genetically modified (GM) crops and similar species is a serious public issue that may potentially endanger ecological stability. Cleistogamy is expected to be an ideal genetic tool for preventing transgene propagation from GM crops. A rice mutant, cl7(t), was created by ethyl methanesulfonate mutagenesis. The mutant exhibited cleistogamy, and had closed spikelets, reduced plant height, and altered morphology of the leaves, panicle, and seeds. Anatomical investigations revealed that the cl7(t) mutant contained more vascular bundles and thicker stems than the wild type, which increased the mechanical strength of its internodes, and anti-lodging ability. Further studies demonstrated that the force required to open the lemma and palea was higher in the cl7(t) mutant, and there was weak swelling ability in the lodicules, which leads to cleistogamy. Allelic analyses and complementation tests indicated that cl7(t) was a novel allele of dep2, a mutant that was previously reported to have similar panicle morphology. Sequence analysis showed that cl7(t) had a single nucleotide substitution (C to A) in the third exon that leads to a Ser substitution with a stop codon, giving a truncated DEP2 protein. Quantitative RT-PCR and in situ hybridization tests demonstrated that there was lower CL7(t) expression level in the spikelets and weaker CL7(t) signals in the lodicules of the cl7(t) mutant compared with wild type, which implies that CL7(t) might participate in the development of lodicules. To improve the agronomic traits of cl7(t) to fit the needs of field production, the cl7(t) mutant was crossed with an intermediate-type rice variety named Guanghui102, which bears some important agronomic traits, including increased grain numbers and high rate of seed setting. Through multi-generational pedigree selection, cleistogamy lines with improved economic traits were obtained, which can be used for the selection of ecologically safe GM rice varieties.

Isolation and characterization of three cadmium-inducible promoters from Oryza sativa.

Cadmium (Cd) is an important soil pollutant. Developing genetically engineered crops might be a feasible strategy for Cd decontamination and damage prevention. Both genes and promoters are critical for the effective construction of genetically modified plants. Although many functional genes for Cd tolerance and accumulation have been identified, few reports have focused on plant Cd-inducible promoters. Here, we identified three Cd-inducible genes in the rice genome: two tau class glutathione S-transferase (GSTU) genes, OsGSTU5 and OsGSTU37, and an HSP20/alpha crystallin family protein gene, OsHSP18.6. The promoter sequences were isolated and tested in transgenic rice lines using a GUSplus reporter gene. All of the promoters exhibited low background expression under normal conditions and could be strongly induced by Cd stress. Although their strength was comparable to that of the constitutive OsACTIN promoter under Cd stress, their time-dependent expression patterns under both short- and long-term Cd exposure were markedly different. The responses of the three promoters to other heavy metals were also examined. Furthermore, heavy metal-responsive cis elements in the promoters were computationally analyzed, and regions determining the Cd stress response were analyzed using a series of truncations. Our results indicate that the three Cd-inducible rice promoters described herein could potentially be used in applications aimed at improving heavy metal tolerance in crops or for the bio-monitoring of environmental contamination.