Pentatricopeptide repeat(PPR)proteins represent one of the largest protein families in plants and typically localize to organelles like mitochondria and chloroplasts.By contrast,CYTOPLASMLOCALIZED PPR1(OsCPPR1)is a cy...Pentatricopeptide repeat(PPR)proteins represent one of the largest protein families in plants and typically localize to organelles like mitochondria and chloroplasts.By contrast,CYTOPLASMLOCALIZED PPR1(OsCPPR1)is a cytoplasm-localized PPR protein that can degrade OsGOLDENLIKE1(OsGLK1)mRNA in the tapetum of rice anther.However,the mechanism,by which OsCPPR1 recognizes and binds to OsGLK1 transcripts,remains unknown.Through protein structure prediction and macromolecular docking experiments,we observed that distinct PPR motif structures of OsCPPR1 exhibited varying binding efficiencies to OsGLK1 RNA.Moreover,RNA-electrophoretic mobility shift assay experiment demonstrated that the recombinant OsCPPR1 can directly recognize and bind to OsGLK1 mRNA in vitro.This further confirmed that the mutations in the conserved amino acids in each PPR motif resulted in loss of activity,while truncation of OsCPPR1 decreased its binding efficiency.These findings collectively suggest that it may require some co-factors to assist in cleavage,a facet that warrants further exploration in subsequent studies.展开更多
UDP-glucose pyrophosphorylase (UGPase) is predominantly present and plays significant role in carbohydrate metabolism in plants. Two homologous UGPase genes, OsUgp1 and OsUgp2, exist in rice genome. OsUgp1 has recentl...UDP-glucose pyrophosphorylase (UGPase) is predominantly present and plays significant role in carbohydrate metabolism in plants. Two homologous UGPase genes, OsUgp1 and OsUgp2, exist in rice genome. OsUgp1 has recently been reported to be essential for callose deposition during pollen mother cell and meiosis stages as well as for seed carbohydrate metabolism. In this study, a full-length cDNA of OsUgp2 was isolated from rice anther. Northern blot and RNA in situ hybridization indicated that the expression of OsUgp2 was preferentially in pollen and developmentally regulated. No transcripts were found in leaf, stem, lemma/palea, ripening grain and florets before the uninucleate microspore developmental stage, but a large quantity of OsUgp2 mRNA was found in pollen at the binucleate and mature stages. The immunolocalization of OsUgp2 showed a similar expression pattern to that by RNA in situ hybridization. The function of OsUgp2 was investigated by dsRNA-mediated transcriptional gene silencing. The pollen fertility of 16 independent transgenic rice plants was found between 25% and 90%, which was correlated with the amount of OsUgp2 mRNA. The results of morphological changes and starch variation during pollen development in transgenic rice showed that the abnormal feature of pollen development appeared after the uninucleate microspore stage. Starch failed to accumulate in pollen and thus led to sterile pollens. These results demonstrated that OsUgp2 is a pollen-preferential "late gene" and plays a key role during pollen maturation, especially for starch accumulation. OsUgp2 complements OsUgp1 to fulfill the UGPase’s functions necessary for the full process of pollen development.展开更多
With the cDNA suppression subtraction hybridization method, a spikelet-specific cDNA library was constructed that expressed at meiosis stage in rice. A total of 121 cDNA fragments were selected from the library and us...With the cDNA suppression subtraction hybridization method, a spikelet-specific cDNA library was constructed that expressed at meiosis stage in rice. A total of 121 cDNA fragments were selected from the library and used as EST (expressed se-quence tags) markers to detect the polymorphism between Annong N, a normal fertile Indica rice line and Annong S-1, its spontaneous mutant with thermo-sensitive genic male sterility, using the RFLP (restriction fragment length polymorphism) technique. HN57, one of the EST probes, could detect poly-morphism between them. The results of segregation analysis with the F2 population developed from An-nong S-1 and Annong N indicate that HN57 co-seg- regates with the thermo-sensitive genic male-sterility controlled by tms5, the recessive gene in Annong S-1. This marker is located on the 31.2-cM region of the chromosome 2 of RGP (rice genome research pro-gram) genetic map. To further determine the location of tms5, 80 SSR (simple sequence repeat) markers around this region were developed, and 12 of them were polymorphic. And finally, the tms5 was mapped within region of 181 kb by using these new markers.展开更多
Hybrid sterility is a major hindrance to utiliz-ing the heterosis in indica-japonica hybrids. To isolate a gene Sc conferring the hybrid sterility, the locus was mapped us-ing molecular markers and an F2 population de...Hybrid sterility is a major hindrance to utiliz-ing the heterosis in indica-japonica hybrids. To isolate a gene Sc conferring the hybrid sterility, the locus was mapped us-ing molecular markers and an F2 population derived from a cross between near isogenic lines. A primary linkage analysis showed that Sc was linked closely with 4 markers on chro-mosome 3, on which the genetic distance between a marker RG227 and Sc was 0.07 cM. Chromosome walking with a rice TAC genomic library was carried out using RG227 as a starting probe, and a contig of ca. 320 kb covering the Sc locus was constructed. Two TAC clones, M45E14 and M90J01 that might cover the Sc locus, were partially se-quenced. By searching the rice sequence databases with se-quences of the TACs and RG227 a japonica rice BAC se-quence, OSJNBb0078P24 was identified. By comparing the TAC and BAC sequences, six new PCR-based markers were developed. With these markers the Sc locus was further mapped to a region of 46 kb. The results suggest that the BAC OSJNBb0078P24 and TAC M45E14 contain the Sc gene. Six ORFs were predicted in the focused 46-kb region.展开更多
Dry rice seeds(Oryza sativa,subspecies indica)were sandwiched between nuclear track detectors,aboard the Chinese spaceship Shenzhou-3 for seven days.The seeds were recovered and the genomic polymorphism in 201 plants ...Dry rice seeds(Oryza sativa,subspecies indica)were sandwiched between nuclear track detectors,aboard the Chinese spaceship Shenzhou-3 for seven days.The seeds were recovered and the genomic polymorphism in 201 plants developed from these seeds was studied using random amplified polymorphism DNA analysis.When compared with plants from ground-based control seeds,the genomic polymorphic bands were amplified in 30.2%of the plants from the seeds exposed in space.The results for sequencing and SNP analyses of the polymorphic bands verified the single nucleotide variations in these plants.Genomic poly-morphisms in the consecutive generations of individual plants of the seeds from space were also discovered.Seven seeds receiving hits of HZE(high atomic number and high energy)particles from space were selected for further analyses and variable genomic polymorphisms were detected in all plants that developed from these seeds.Among them,the embryos of three seeds were hit at least once,and mutants with significant changes in agronomic traits were only found in later generations of these seeds.This result implies that the HZE particles of space are effective in inducing the changes of plant genome of inherited phenotypes.展开更多
基金the Open Competition Program of Top Ten Critical Priorities of Agricultural Science and Technology Innovation for the 14th Five-Year Plan of Guangdong Province,China(Grant No.2022SDZG05)the National Natural Science Foundation of China(Grants Nos.32000457,31871231,and 31921004)+1 种基金the Major Program of Guangdong Basic and Applied Research,China(Grant No.2019B030302006)the Double First-Class Discipline Promotion Project,China(Grant No.2021B10564001).
文摘Pentatricopeptide repeat(PPR)proteins represent one of the largest protein families in plants and typically localize to organelles like mitochondria and chloroplasts.By contrast,CYTOPLASMLOCALIZED PPR1(OsCPPR1)is a cytoplasm-localized PPR protein that can degrade OsGOLDENLIKE1(OsGLK1)mRNA in the tapetum of rice anther.However,the mechanism,by which OsCPPR1 recognizes and binds to OsGLK1 transcripts,remains unknown.Through protein structure prediction and macromolecular docking experiments,we observed that distinct PPR motif structures of OsCPPR1 exhibited varying binding efficiencies to OsGLK1 RNA.Moreover,RNA-electrophoretic mobility shift assay experiment demonstrated that the recombinant OsCPPR1 can directly recognize and bind to OsGLK1 mRNA in vitro.This further confirmed that the mutations in the conserved amino acids in each PPR motif resulted in loss of activity,while truncation of OsCPPR1 decreased its binding efficiency.These findings collectively suggest that it may require some co-factors to assist in cleavage,a facet that warrants further exploration in subsequent studies.
基金Supported by National Natural Science Foundation of China (Grant No. 30370800)Changjiang Scholars and Innovative Research Team from Ministry of Education in China (Grant No. IRT0448)
文摘UDP-glucose pyrophosphorylase (UGPase) is predominantly present and plays significant role in carbohydrate metabolism in plants. Two homologous UGPase genes, OsUgp1 and OsUgp2, exist in rice genome. OsUgp1 has recently been reported to be essential for callose deposition during pollen mother cell and meiosis stages as well as for seed carbohydrate metabolism. In this study, a full-length cDNA of OsUgp2 was isolated from rice anther. Northern blot and RNA in situ hybridization indicated that the expression of OsUgp2 was preferentially in pollen and developmentally regulated. No transcripts were found in leaf, stem, lemma/palea, ripening grain and florets before the uninucleate microspore developmental stage, but a large quantity of OsUgp2 mRNA was found in pollen at the binucleate and mature stages. The immunolocalization of OsUgp2 showed a similar expression pattern to that by RNA in situ hybridization. The function of OsUgp2 was investigated by dsRNA-mediated transcriptional gene silencing. The pollen fertility of 16 independent transgenic rice plants was found between 25% and 90%, which was correlated with the amount of OsUgp2 mRNA. The results of morphological changes and starch variation during pollen development in transgenic rice showed that the abnormal feature of pollen development appeared after the uninucleate microspore stage. Starch failed to accumulate in pollen and thus led to sterile pollens. These results demonstrated that OsUgp2 is a pollen-preferential "late gene" and plays a key role during pollen maturation, especially for starch accumulation. OsUgp2 complements OsUgp1 to fulfill the UGPase’s functions necessary for the full process of pollen development.
文摘With the cDNA suppression subtraction hybridization method, a spikelet-specific cDNA library was constructed that expressed at meiosis stage in rice. A total of 121 cDNA fragments were selected from the library and used as EST (expressed se-quence tags) markers to detect the polymorphism between Annong N, a normal fertile Indica rice line and Annong S-1, its spontaneous mutant with thermo-sensitive genic male sterility, using the RFLP (restriction fragment length polymorphism) technique. HN57, one of the EST probes, could detect poly-morphism between them. The results of segregation analysis with the F2 population developed from An-nong S-1 and Annong N indicate that HN57 co-seg- regates with the thermo-sensitive genic male-sterility controlled by tms5, the recessive gene in Annong S-1. This marker is located on the 31.2-cM region of the chromosome 2 of RGP (rice genome research pro-gram) genetic map. To further determine the location of tms5, 80 SSR (simple sequence repeat) markers around this region were developed, and 12 of them were polymorphic. And finally, the tms5 was mapped within region of 181 kb by using these new markers.
文摘Hybrid sterility is a major hindrance to utiliz-ing the heterosis in indica-japonica hybrids. To isolate a gene Sc conferring the hybrid sterility, the locus was mapped us-ing molecular markers and an F2 population derived from a cross between near isogenic lines. A primary linkage analysis showed that Sc was linked closely with 4 markers on chro-mosome 3, on which the genetic distance between a marker RG227 and Sc was 0.07 cM. Chromosome walking with a rice TAC genomic library was carried out using RG227 as a starting probe, and a contig of ca. 320 kb covering the Sc locus was constructed. Two TAC clones, M45E14 and M90J01 that might cover the Sc locus, were partially se-quenced. By searching the rice sequence databases with se-quences of the TACs and RG227 a japonica rice BAC se-quence, OSJNBb0078P24 was identified. By comparing the TAC and BAC sequences, six new PCR-based markers were developed. With these markers the Sc locus was further mapped to a region of 46 kb. The results suggest that the BAC OSJNBb0078P24 and TAC M45E14 contain the Sc gene. Six ORFs were predicted in the focused 46-kb region.
基金This study was supported by the grants from The National High-Tech R&D Program of China in Aerospace(No.2002AA744061)the National Natural Sciences Foundation of China(Grant No.30170534).
文摘Dry rice seeds(Oryza sativa,subspecies indica)were sandwiched between nuclear track detectors,aboard the Chinese spaceship Shenzhou-3 for seven days.The seeds were recovered and the genomic polymorphism in 201 plants developed from these seeds was studied using random amplified polymorphism DNA analysis.When compared with plants from ground-based control seeds,the genomic polymorphic bands were amplified in 30.2%of the plants from the seeds exposed in space.The results for sequencing and SNP analyses of the polymorphic bands verified the single nucleotide variations in these plants.Genomic poly-morphisms in the consecutive generations of individual plants of the seeds from space were also discovered.Seven seeds receiving hits of HZE(high atomic number and high energy)particles from space were selected for further analyses and variable genomic polymorphisms were detected in all plants that developed from these seeds.Among them,the embryos of three seeds were hit at least once,and mutants with significant changes in agronomic traits were only found in later generations of these seeds.This result implies that the HZE particles of space are effective in inducing the changes of plant genome of inherited phenotypes.
