Qingyao Shu

Gene actions of QTLs affecting several agronomic traits resolved in a recombinant inbred rice population and two backcross populations

To understand the types of gene action controlling seven quantitative traits in rice, we carried out quantitative trait locus (QTL) mapping in order to distinguish between the main-effect QTLs (M-QTLs) and digenic epistatic QTLs (E-QTLs) responsible for the trait performance of 254 recombinant inbred lines (RILs) from rice varieties Lemont/Teqing and two backcross hybrid (BCF1) populations derived from these RILs. We identified 44 M-QTL and 95 E-QTL pairs in the RI and BCF1 populations as having significant effects on the mean values and mid-parental heterosis of heading date, plant height, flag leaf length, flag leaf width, panicle length, spikelet number and spikelet fertility. The E-QTLs detected collectively explained a larger portion of the total phenotypic variation than the M-QTLs in both the RI and BCF1 populations. In both BCF1 populations, over-dominant (or under-dominant) loci were more important than additive and complete or partially dominant loci for M-QTLs and E-QTL pairs, thereby supporting prior findings that overdominance resulting from epistatic loci are the primary genetic basis of inbreeding depression and heterosis in rice.

Transgenic rice plants with a synthetic cry1Ab gene from Bacillus thuringiensis were highly resistant to eight lepidopteran rice pest species.

To fully explore the resistance potential of transgenic rice produced by Agrobacterium-mediated transformation, an elite line KMD1 was assessed for its resistance to eight lepidopteran rice pest species. KMD1 contained a synthetic cry1Ab gene from Bacillus thuringiensis under the control of a maize ubiquitin promoter. It was derived from a commercial japonica Chinese rice variety Xiushui 11, and bred true for both agronomic traits and a cry1Ab gene when the bioassays were done in 1998 in the R5 generation. The eight lepidopteran pest species were: four Pyralidae species: Chilo suppressalis (striped stem borer, SSB), Scirpophaga incertulas (yellow stem borer, YSB), Cnaphalocrocis medinalis (leaf folder), Herpitogramma licarisalis; two Noctuidae: Sesamia inferens (pink stem borer, PSB) and Naranga anescens; one Stayridae: Mycalesis gotama; and one Hesperiidae, Parnara guttata. In laboratory bioassays, 100% mortality was observed in all insect species when their newly hatched or third-instar larvae were fed KMD1 leaf tissues, whereas only 9.65% of the neonates and none of the third-instar larvae died when fed the leaf tissues of non-transgenic control. Moreover, the leaf area of control tissues consumed in four days by stem borers was 20 to 40 times higher than that of KMD1 tissues, and the area of control tissues eaten by leaf-feeding species was 120 to 180 times greater than that of the transgenic tissues. Under natural infestation, no KMD1 plant was visibly damaged by the SSB, YSB and leaf folder in field evaluation. On the other hand, 80, 9.3 and 88.7% of control plants were injured by SSB, YSB, and leaf folder, respectively. These data disclosed that the transgenic line was highly resistant to a broad spectrum of lepidopteran insect species and could be useful in insect resistance breeding programs.

Field Evaluation of Resistance of Transgenic Rice Containing a Synthetic cry1Ab Gene from Bacillus thuringiensis Berliner to Two Stem Borers

Abstract Two transgenic rice (Oryza sativa L.) lines, KMD1 and KMD2 at the R4 generation, transformed with a synthetic cry1Ab gene from Bacillus thuringiensis Berliner, were first evaluated for stem borer resistance in the field during the rice growing season of 1998 in two areas of Zhejiang Province, China. Both KMD1 and KMD2 were highly resistant to the stem borers Chilo suppressalis (Walker) and Scirpophaga incertulas (Walker), and were completely undamaged during the whole rice growing season. In contrast, damage to the plants of the untransformed parental control (Xiushui 11) was in the form of deadhearts or whiteheads. Under natural infestation by the C. suppressalis, the damage to control plants reached a peak of 88.7% of plants and 20.1% of tillers encountered with deadhearts. Under artificial and natural infestation of neonate striped stem borers at the vegetative stage and booting stage, 100% of plants and 25.6% of tillers, 78.9% of plants and 15.6% of productive tillers among artificially infested control plants were observed with the symptom of deadhearts and whiteheads, respectively. Damage to the control plants from artificial infestation by the S. incertulas reached a peak of 97.0% of plants and 22.9% of tillers damaged. The field research indicated that both KMD1 and KMD2 show great potential for protecting rice from attack by these two stem borers.

Effect of gamma irradiation on starch viscosity and physicochemical properties of different rice

Abstract Three types of rice cultivars (indica, japonica and hybrid rice) with similar intermediate apparent amylose content (AAC) as well as early indica rice cultivars with different amounts of AAC were selected for studying the effects of gamma irradiation on starch viscosity, physicochemical properties and starch granule structure. Four major parameters of RVA profile, that was determined by a rapid visco analyser (RVA, Model-3D), peak viscosity, hot pasting viscosity, cool pasting viscosity, and setback viscosity, were considerably decreased with increasing dose levels. Gamma irradiation reduced the amylose contents in the cultivars with low AAC, intermediate AAC, and glutinous rice, but had no effects on the high AAC cultivar. No visible changes in gelatinization temperature were detected after irradiation, but the peak time was reduced with the dose levels. Gel consistency was significantly increased in the tested cultivars, especially in the high AAC indica rice. The starch granules were somewhat deformed by gamma irradiation. These results suggested that it is promising to use gamma irradiation to improve rice eating or cooking quality.

High levels of stable resistance in transgenic rice with a cry1Ab gene from Bacillus thuringiensis Berliner to rice leaffolder, Cnaphalocrocis medinalis (Guenée) under field conditions

Abstract Two transgenic rice (Oryza satica L.) lines, KMD1 and KMD2, containing a synthetic cry1Ab gene from Bacillus thuringiensis Berliner, were evaluated for resistance to the rice leaffolder (RLF), Cnaphalocrocis medinalis (Guenee) under field conditions for 3 years in Zhejiang Province, China. Both KMD1 and KMD2 exhibited high and stable resistance against natural infestations by the RLF, and showed no symptoms of damaged leaves throughout the rice-growing season. In contrast, the untransformed parental control line (Xiushui 11) showed serious RLF damage symptoms not only in untreated plots, but also in plots treated once with chemical insecticides. These results demonstrate that both KMD1 and KMD2 have potential for protecting rice from the RLF damage.

QTL mapping for the paste viscosity characteristics in rice (Oryza sativa L.)

Abstract In order to understand the genetic basis of the paste viscosity characteristics (RVA profile, which is tested on the Rapid Visco Analyser) of the rice grain, we mapped QTLs for RVA profile parameters using a DH population derived from a cross between an indica variety, Zai-Ye-Qing 8 (ZYQ8), and a japonica variety, Jing-Xi 17 (JX17). Evidence of genotype-by-environment interaction was found by comparing the mapped QTLs between two locations, Hainan (HN) and Hangzhou (HZ). A total of 20 QTLs for six parameters of the RVA profiles were identified at least one location. Only the waxy locus (wx) located on chromosome 6 was detected significantly at both environments for five traits, i.e. hot paste viscosity (HPV), cool paste viscosity (CPV), breakdown viscosity (BDV), consistency viscosity (CSV) and setback viscosity (SBV). This locus explained 19.5%–63.7% of the total variations at both environments, suggesting that the RVA profiles were mainly controlled by the wx gene. HPV, CPV, BDV, CSV and SBV were also controlled by other QTLs whose effects on the respective parameter were detected only in one environment, while for the peak viscosity (PKV), only 2 QTLs, 1 at HN,the other at HZ, were identified. These results indicate that RVA profiles are obviously affected by environment.

A Rice cis-Natural antisense RNA acts as a translational enhancer for its cognate mRNA and contributes to phosphate homeostasis and plant fitness

cis-natural antisense transcripts (cis-NAT) are widespread in plants and typically associated with transcriptional downregulation of their cognate sense gene. This works shows that a noncoding cis-NAT associated with the rice phosphate exporter gene PHO1;2 enhances translation of the sense PHO1;2 mRNA. Reduction of cis-NATPHO1;2 expression negatively impacts phosphate homeostasis and seed yield. cis-natural antisense transcripts (cis-NATs) are widespread in plants and are often associated with downregulation of their associated sense genes. We found that a cis-NAT positively regulates the level of a protein critical for phosphate homeostasis in rice (Oryza sativa). PHOSPHATE1;2 (PHO1;2), a gene involved in phosphate loading into the xylem in rice, and its associated cis-NATPHO1;2 are both controlled by promoters active in the vascular cylinder of roots and leaves. While the PHO1;2 promoter is unresponsive to the plant phosphate status, the cis-NATPHO1;2 promoter is strongly upregulated under phosphate deficiency. Expression of both cis-NATPHO1;2 and the PHO1;2 protein increased in phosphate-deficient plants, while the PHO1;2 mRNA level remained stable. Downregulation of cis-NATPHO1;2 expression by RNA interference resulted in a decrease in PHO1;2 protein, impaired the transfer of phosphate from root to shoot, and decreased seed yield. Constitutive overexpression of NATPHO1;2 in trans led to a strong increase of PHO1;2, even under phosphate-sufficient conditions. Under all conditions, no changes occurred in the level of expression, sequence, or nuclear export of PHO1;2 mRNA. However, expression of cis-NATPHO1;2 was associated with a shift of both PHO1;2 and cis-NATPHO1;2 toward the polysomes. These findings reveal an unexpected role for cis-NATPHO1;2 in promoting PHO1;2 translation and affecting phosphate homeostasis and plant fitness.

Inheritance and expression of the cry1Ab gene in Bt (Bacillus thuringiensis) transgenic rice

Abstract The inheritance and expression patterns of the cry1Ab gene were studied in the progenies derived from different Bt (Bacillus thuringiensis) transgenic japonica rice lines under field conditions. Both Mendelian and distorted segregation ratios were observed in some selfed and crossed F2 populations. Crosses between japonica intra-subspecies had no significant effect on the segregation ratios of the cry1Ab gene, but crossing between japonica and indica inter-subspecies led to distorted segregation of the cry1Ab gene in the F2 population. Field-release experiments indicated that the cry1Ab gene was stably transmitted in an intact manner via successive sexual generations, and the concentration of the Cry1Ab protein was kept quantitatively stable up to the R6 generation. The cry1Ab gene, driven by the maize ubiquitin promoter, displayed certain kinds of spatial and temporal expression patterns under field conditions. The content of the Cry1Ab protein varied in different tissues of the main stems, the primary tillers and the secondary tillers. Higher levels of the Cry1Ab protein were found in the stems, leaves and leaf sheaths than in the roots, while the lowest level was detected in grains at the maturation stage. The content of the Cry1Ab protein in the leaves peaked at the booting stage and was lowest at the heading stage. Furthermore, the Cry1Ab content of cry1Ab expression in different tissues of transgenic rice varied individually with temperature.

Metabolite Profiling of Germinating Rice Seeds

A metabolite profiling approach based on gas chromatography-mass spectrometry (GC-MS) was used to investigate time-dependent metabolic changes in the course of the germination of rice. Brown rice kernels were soaked and incubated for a total of 96 h under ambient conditions. Samples taken during the germination process were subjected to an extraction and fractionation procedure covering a broad spectrum of lipophilic (e.g., fatty acid methyl esters, hydrocarbons, fatty alcohols, sterols) and hydrophilic (e.g., sugars, acids, amino acids, amines) low molecular weight rice constituents. Investigation of the obtained fractions by GC resulted in the detection of 615 distinct peaks, of which 174 were identified by means of MS. Statistical assessment of the data via principal component analysis demonstrated that the metabolic changes during the germination process are reflected by time-dependent shifts of the scores, which were similar for the three rice materials investigated. Analysis of the corresponding loadings showed that polar metabolites were major contributors to the separation along the first principal component. Relative quantifications based on standardized peak heights revealed dynamic changes of the metabolites in the course of the germination.

QTL for rice grain quality based on a DH population derived from parents with similar apparent amylose content

A doubled haploid (DH)population consisting of 135 lines, derived from an indica (IR64) and a japonica (Azucena) rice with a similar apparent amylose content (AAC), was used to investigate the genetic factors affecting cooking and eating quality of rice. AAC,gelatinization temperature (GT), gel consistency (GC) and six starch pasting viscosity parameters were measured for quantitative trait loci (QTL) analysis using 193 molecular markers mapped on the DH population. A total of 17 QTLs were detected for the 9 traits, with at least one QTL and as many as 3 QTLs for each individual trait. No QTL for the measured parameters was found at the wx locus,possibly because of the similar AAC between the parents. Several QTLs with important effects on the variations in the measured parameters were detected in the present study which have not been found in earlier reports based on populations derived from parents with different AAC and wxgene alleles. Two interesting loci could be deduced from the present study according to the marker order compared with other genetic linkage maps. A QTL flanked by Amy2A and RG433 on the end of the long arm of chromosome 6, identified for GT, set back and consistency viscosity, might cover the gene encoding starch branching enzyme I. Similarly, a QTL flanked by RG139 and RZ58on chromosome 2, detected for hot paste viscosity and breakdown viscosity, might cover the gene encoding starch branching enzyme III. Generally, traits significantly correlated with each other shared identical QTL, but it was not true in some cases. The fine molecular mechanisms underlying these traits await further elucidation for the improvement of eating and cooking quality of rice.