Stamen specification and anther development in rice

Male reproductive development is a complex biological process which includes the formation of the stamen with differentiated anther tissues, in which microspores/pollens are generated, then anther dehiscence and subsequently pollination. Stamen specification and anther development involve a number of extraordinary events such as meristem transition, cell division and differentiation, cell to cell communication, etc., which need the cooperative interaction of sporophytic and gametophytic genes. The advent of various tools for rice functional gene identification, such as complete genome sequence, genome-wide microarrays, collections of mutants, has greatly facilitated our understanding of mechanisms of rice stamen specification and anther development. Male sterile lines are critical for hybrid rice breeding, therefore understanding these processes will not only contribute greatly to the basic knowledge of crop developmental biology, but also to the development of new varieties for hybrid rice breeding in the future.     

Identification and functional analysis of 23 novel box C／D snoRNAs from Oryza sativa

In a cDNA library generated from rice small nuclear RNAs,30box C/D small nucleolar RNAs (snoRNAs) were identiffied through preliminary screen.Except 7 known snoRNAs such as U14,all snoRNAs were identified in rice for the first time experimentally.Among the 23 novel snoRNAs,11 snoRNAs appear rice-specific,6 snoRNAs are unique to plants,the remaining 6 snoRNAs have their counterparts in both Arabidopsis and yeast or mammals according to the conserved antisense sequencs that guide 2‘-O-ribose methylation of rRNA,17 of the 23 novel snoRNAs were predicted to guide 24 2‘‘-O-ribose methylations at the specificsites of rice 5.8S,18S,25S rRNAs,among which 19 methylated sites were determined by primer extension at low dNTP concentrations.The remaining 6 snoRNAs devoid of rRNA antisense elements may represent novel snoRNA species in rice.The results show that constructing a cDNA library from small nuclear RNAs is an effective experimental approach for novel snoRNA is identification.The novel snoRNAs are important in elucidating the genomic organization and expression of plant snoRNA genes and the mechanism through which 2‘‘-O-ribose methylations took place in rRNAs.     

A simple and effective method for total RNA isolation of appressoria in Magnaporthe oryzae

Appressorium formation is an important event in establishing a successful interaction between the rice blast fungus, Magnaporthe oryzae, and its host plant, rice. An understanding of molecular events occurring in appressorium differentiation will give new strategies to control rice blast. A quick and reliable method to extract total RNA from appressorium is essential for studying gene expression during appressorium formation and its mechanism. We found that duplicate film is an efficient substratum for appressorium formation, even when inoculated with high density conidia. When inoculated with conidia at 1 × 106 ml^-1, the percentages of conidium germination and appressorium formation were （97.98±0.67）% and （97.88±0.45）%, respectively. We applied Trizol before appressorium collection for total RNA isolation, and as much as 113.6 lag total RNA was isolated from the mature appressoria at 24 h after inoculation. Functional analysis of two genes, MNH6 and MgATG1, isolated from the cDNA subtractive library, revealed that the quantity of RNA was good enough to construct a cDNA （complementary DNA） library or a cDNA subtractive library. This method may be also applicable for the appressorium RNA isolation of other pathogenic fungi in which conidia differentiate into appressoria in the early stages of host infection.     

Construction and characterization of a normalized whole-life-cycle cDNA library of rice

A cDNA library with genomic complete coverage is a powerful tool for functional genomic studies.For studying the functions of rice genes on a large scale,a normalized whole-life-cycle cDNA library is constructed based on the strategy of saturation hybridization with genomic DNA using rice cultivar Minghui 63,an elite restorer line for a number of rice hybrids that are widely cultivated in China,This library consists of cDNA from 15 directionally cloned cDNA libraries constructed with different tissues from 9 developmental stages.For normalization,the denatured plasmids purified from the 15 directionally cloned libraries are mixed and hybridized with saturated genomic DNA labeled with magnetic beads in two complementary systems. Well-matched plasmids are captured from the hybridized genomic DNA and electroporated into competent DH10B E. coli for construction of the normalized whole-life-cycle cDNA library.This library consists of 62000 clones with an average insert length about 1.4kb.Inverse Northern blotting shows that this cDNA library included many rarely expressed genes and tissue-specific genes.Sequencing of 10750 cDNA clones of this library reveals 6399 unique EST s(expressed sequence tags),indicating that the non-redundancy of the library is about 59.5%.This library has been used to make cDNA microarrays for functional genomic studies.     

Enhancing Phosphorous Recovery in a Novel Biofilter System

Developing new techniques to remove and recover phosphorous simultaneously from wastewaters is very important for sustainable utilization of phosphorous resource and prevention of eutrophication. The feasibility of phosphorus bioaccumulation in a novel continuous alternating anaerobic /aerobic biofilter and benefits of such accumulation for its further recovery as magnesium ammonia phosphate （MAP） were studied. The system was operated for seven months and employed the atomic absorption spectrophotometer （AAS） method to analyze the distribution change of magnesium and calcium with phosphate in the biofilter during that period. The benefits of utilizing the biofilter system to accumulate phosphorous ＆ magnesium and suppress calcium simultaneously to phosphorous recovery as MAP were explained with the aid of chemical equilibrium models. The improved phosphorus recovery from 42% to 82% was approved in the recovery experiments and the recovery products mainly as MAP were evidenced and characterized with scanning electron microscopy （SEM） . The study provides references for designing and operating the novel alternating anaerobic /aerobic biofilter for phosphorus bioaccumulation and recovery.     

Improve Fractal Compression Encoding Speed Using Feature Extraction and Self-organization Network

Image compression consists of two main parts: encoding and decoding. One of the important problems of the fractal theory is the long encoding implementation time, which hindered the acceptance of fractal image compression as a practical method. The long encoding time results from the need to perform a large number of domain-range matches, the total encoding time is the product of the number of matches and the time to perform each match. In order to improve encoding speed, a hybrid method combining features extraction and self-organization network has been provided, which is based on the feature extraction approach the comparison pixels by pixels between the feature of range blocks and domains blocks. The efficiency of the new method was been proved by examples.     

Cloning and expression of <Emphasis Type="Italic">AtPLC6</Emphasis>, a gene encoding a phosphatidylinositol-specific phospholipase C in <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis>

A full-length cDNA clone corresponding to a putative phosphatidylinositol-specific phospholipase C(PIPLC) was isolated from Arabidopsis thaliana by screening a cDNA library and using RT-PCR strategy.The cDNA,designated AtPLC6,encodes a putative polypeptide of 578 amino acid residues with a calculated molecular mass of 66251.84 D and a pI of 7.24. The sequence analysis indicates that the polypeptide contains X, Y, EF-hand and C2 domains.The overall structure of putative AtPLC6 protein, like other plant PI-PLCs,is most similar to that of mammalian PLCδ The recombinant AtPLC6 protein expressed in E. coil was able to hydrolyze phosphatidylinositol 4,5-biophosphate (PIP2) to generate inositol 1,4,5-trisphate (IP3) and 1,2-diacylglycerol (DAG).The protein hydrolyzes PIP2 in a Ca^2 -dependent manner and the optimum concentration of Ca^2 is 10μmol/L.These results suggested that AtPLC6 gene encodes a genuine PIPLC.Northern blot analysis showed that the AtPLC6 gene is expressed at low level in all examined tissues, such as roots,stems,leaves,flowers,siliques and seedlings under normal growth conditions.The gene is strongly induced under low temperature and weakly induced under various stresses,such as ABA, high-salt stress and heat. These results suggested that AtPLC6 might be involved in the signal-transduction pathways of cold responses of the plants.     

Molecular cloning of rice cDNAs related to pollen development using the subtraction hybridization technique

The meiotic stage of pollen mother cell is a very important stage in controlling the development and formation of pollen. In order to clone the rice cDNA(s) of this stage, a normal rice, Annong N and its thermosensitive mutant, Annong S-1 were used as the plant material. The mRNA has been extracted from the young panicle at the meiotic stage. By using the cDNA subtraction hybridization technique, three cDNA fragments, RP-1, RP-2 and RP-3 have been successfully cloned from Annong N. Northern blot analysis reveals that the mRNA of these three clones are expressed only in anthers, and not leaves. The mRNA levels of these clones are lower in anthers of Annong S-1 than in Annong N. Furthermore, the amount of mRNA extracted from anthers of Annong S-1 growing under high temperature (28℃) is lower than plants growing at lower temperature (25℃). Sequence analysis and homology search indicate that these three clones display no similarity to the current database. It is concluded that the three novel cDNA cloned are related to pollen development in rice.     

Salt tolerance of transgenic rice (Oryza sativa L.) with mtlD gene and gutD gene

Southern blot analysis indicated that mtlD gene (encoding mannitol-1-phosphate dehydrogenase) and gutD gene (encoding glucitol-6-phosphate dehydrogenase) had been integrated into the rice genome mediated by Agrobacterium tumefaciens LBA4404(pBIGM). The expression of the above two genes in transgenic rice plants was demonstrated by Northern blot analysis and enzymatic activity assay. Analysis of sugar alcohol showed that transgenic rice plants could produce and accumulate mannitol and sorbitol. The salt tolerance of transgenic plants was much higher than that of their controls.     

Salt tolerance of transgenic rice (Oryza sativa L.) withmtlD gene andgutD gene

Southern blot analysis indicated thatmtlD gene (encoding mannitol-1-phosphate dehydrogenase) andgutD gene (encoding glucitol-6-phosphate dehydrogenase) had been integrated into the rice genome mediated byAgrobacterium tumefaciens LBA4404(pBIGM). The expression of the above two genes in transgenic rice plants was demonstrated by Northern blot analysis and enzymatic activity assay. Analysis of sugar alcohol showed that transgenic rice plants could produce and accumulate mannitol and sorbitol. The salt tolerance of transgenic plants was much higher than that of their controls.     

Isolation ofosRACD gene encoding a small GTP-binding protein from rice

Using an improved version of mRNA differential display technology, we have obtained a differentially displayed fragment RDP-8. Homologous comparison indicated that the fragment RDP-8 has high homology with the gene encoding maize small GTP-binding protein. By screening cDNA library of the rice Nongken 58N pan icle using the newly obtained fragment RDP-8 as probe, we further found the full-length cDNA of osRACD gene that encodes a rice small GTP-binding protein. Asco mpared with maize RACD gene, the osRACD of rice shows remarkable homology in both nucleotide sequence and amino acid sequence, 88% and 97% respectively. Evidence from RT-PCR study indicates that osRACD gene is related to photoperiod fertility conversion of photoperiod sensitive genic male sterility (PSGMS) rice.     

水稻高亲和力磷酸盐转运蛋白基因的克隆、表达及检测

以水稻（粳稻）cDNA为模板,设计一对特异性引物,获得编码水稻高亲和力磷酸盐转运蛋白基因的全序列,ORF区编码含535个氨基酸的蛋白,具有磷酸盐转运蛋白保守的蛋白激酶C作用位点、N端糖基化位点与酪蛋白激酶Ⅱ作用位点Southern blot分析显示此基因在水稻基因组中具有多个拷贝,与水稻基因组序列分析结果一致Westernblot预测目的基因表达的蛋白大小约为57．7ku．该基因的真核表达研究正在进行中．     

Cloning and expression of a cDNA encoding ribosomal protein S4 from Rice (Oryza sativa)

A cDNA clone, pS4, has been isolated from a cDNA library prepared from rice anthers of about 1.0 mm in length. DNA sequence analysis and database search show that the cDNA encodes a protein which is highly homologous to eukaryotic 80S ribosomal protein subunit 4 (S4). Northern hybridization indicates that this gene expresses in all tissues analyzed although the expression level varies and it cannot be induced by mechanical wounding in leaves. Southern blot analysis demonstrates that this rice S4 gene is from a multigene family.     

白鱀豚某些组织中谷丙、谷草转氨酶的初步研究

本文对白鱀豚几种组织中的GPT和GOT活性及肝脏中此两种酶的一些性质进行了探讨。