水稻OsGRF6的克隆及调控初生根发育功能的初步分析

OsGRF6隶属于GRF家族，参与调控植物的生长发育和调控植物对非生物胁迫的耐受性。为了研究OsGRF6调控水稻初生根发育的分子机制，该研究从水稻品种‘ZH11’中克隆得到转录因子基因OsGRF6，并对其进行了进化树构建，启动子分析，并对OsGRF6-OE和osgrf6进行了基因型分析和qRT-PCR鉴定及水稻表型观察，并通过酵母单杂交检测了OsGRF6的结合基序，进一步通过ChIP-seq和RNA-seq数据分析了OsGRF6下游的候选靶基因。结果显示：（1）OsGRF6含有一个QLQ和一个WRC保守的功能结构域，与拟南芥AtGRF1和AtGRF2以及水稻OsGRF7亲缘关系较近。OsGRF6启动子区域含有多个非生物胁迫和激素响应顺式作用元件。（2）OsGRF6在水稻种子、幼穗和根中表达量较高，而在叶中表达量较低。亚细胞定位显示，OsGRF6定位于细胞核，并且OsGRF6具有转录激活活性。（3）与野生型相比，过表达OsGRF6的转基因材料表现出初生根长度较长，而突变体材料则表现出初生根变短的表型。（4）OsGRF6能与CGGCA基序结合。（5）结合ChIP-seqs和RNA-seqs分析发现，OsGRF6靶基因中包含多个参与调控水稻根发育的基因，如OsARF7，OsARF4等。     

长穗颈水稻的内源GA1、IAA和ABA含量变化

分别以携有长穗颈基因eui1、eui2和野生型基因Eui的协青早不育系和保持系6个水稻品种为材料,测定它们在抽穗始期植株中内源GA1、IAA和ABA含量的变化。结果表明,携有eui1和eui2基因的水稻可以在植株体内产生大量内源GA1,携有eui1基因的GA1含量比携有eui2基因的高。携有eui1基因的ABA含量最高,携有eui2基因的其次,而携有Eui基因的最低。IAA含量也表现出同样的趋势。表明长穗颈基因主要是通过调节内源GA1含量促进水稻最上节间的剧烈伸长。     

Overexpression of Rice Sphingosine-1-Phoshpate Lyase Gene OsSPL1 in Transgenic Tobacco Reduces Salt and Oxidative Stress Tolerance

Sphingolipids, including sphingosine-1-phosphate (S1P), have been shown to function as signaling mediators to regulate diverse aspects of plant growth, development, and stress response. In this study, we performed functional analysis of a rice (Oryza sativa) S1P lyase gene OsSPL1 in transgenic tobacco plants and explored its possible involvement in abiotic stress response. Overexpression of OsSPL1 in transgenic tobacco resulted in enhanced sensitivity to exogenous abscisic acid (ABA), and decreased tolerance to salt and oxidative stress, when compared with the wild type. Furthermore, the expression levels of some selected stress-related genes in OsSPL1-overexpressing plants were reduced after application of salt or oxidative stress, indicating that the altered responsiveness of stress-related genes may be responsible for the reduced tolerance in OsSPL1-overexpressing tobacco plants under salt and oxidative stress. Our results suggest that rice OsSPL1 plays an important role in abiotic stress responses.     

Genotypes of Heading Date of Middle Indica Rice in the Mid-lower Region of the Yangtze River

Middle indica cultivars are planted in the middle to lower regions of the Yangtze River. Hybrid combinations with these cultivars have a high yield potential but the presence of late-transgressive-segregants limits the further exploitation of heterosis. To understand the genetic basis of the heading date in these middle maturing cultivars, we carried out a genetic analysis of 10 typical middle-season cultivars using a number of heading date isogenic lines under both long and short day conditions. The results showed that Teqing, 752, CDR22, Bo B, 9311 and 11-32B carry two photoperiod sensitive genes E1 or Se.1 and E3, and Teqing, 752 and CDR22 carry a dominant early-heading gene Ef-1 while Bo B, 9311 and 11-32B carry a recessive late-heading gene ef-1. Based on the findings of the present and previous works on Guichao 2, Minghui 63, Nanjing 11 and Pei＇ai64S, it was concluded that all of these middle indica cultivars carried the recessive allele hd2, which could inhibit the expression of El or Se.1, and they formed rational combinations of genotype for heading date during a long period of evolution. The effects of the different combinations of genotype for heading date on rice cultivation and extension were discussed, and genetic basis of broad adaptability of hybrid middle indica cultivars was analyzed.     

稻瘟病菌(Magnaporthe grisea)诱导的水稻早期反应基因ER1的5' 端cDNA片段克隆及序列分析

研究高等生物基因表达与调控的一个重要方面是分离基因的编码区及其上游的调控序列（ＤｅＶｅｅｒ等１９９７）,这需要获得一个基因的ｃＤＮＡ全长及从植物基因组获取全基因。在前文（周建明等１９９９）中曾经分离了稻瘟病菌侵染诱导的水稻早期反应基因ＥＲ１的ｃＤＮＡ片段,但是运用ｍＲＮＡ差异显示技术分离的ｃＤＮＡ片段往往只有近ｍＲＮＡ３’端的一部分,难以反映基因的结构及功能特点,因此,必须进一步分离其５’端的部分才有可能比较全面地了解此基因的特点。ＲＡＣＥ（ｒａｐｉｄａｍｐｌｉｆｉｃａｔｉｏｎｏｆｃＤＮＡｅｎ…     

从水稻细胞质型果糖-1,6-二磷酸酶启动子上游中去除93 bp可以彻底改变其表达模式

细胞质型果糖-1,6-二磷酸基因ATG上游1 195bp侧翼序列可调控GUS基因在水稻(Oryza sativa L.)中特异性表达,因此该片段包含有使报告基因在叶肉细胞中特异性表达的所有顺式元件.为了研究其调控特异表达的顺式元件,对启动子5′端进行了一系列的缺失,得到4种与GUS基因融合的植物表达载体,通过基因枪法转入水稻.结果表明,自启动子5′端-1 195 bp缺失至-1 102 bp时,GUS基因由叶肉细胞特异性表达变为组成型表达,且表达活性有所提高,推测在该区段中存在调控叶肉细胞特异性表达的顺式元件.进一步缺失仍然保持组成性表达的模式,即在转化株的根、茎和叶中的所有细胞中均有表达,同时启动子活性有所提高.这一结果暗示该启动子具有很大的应用潜力.     

水稻P0491E01基因调控花药发育的功能分析(简报)

高等植物的花药发育是包含基因不同程度相互作用的复杂发育过程,一般可分为两个阶段。第一个是花药的形态建成阶段,在这一阶段中细胞与组织发生分化,小孢子母细胞进行减数分裂形成四分     

Roles of OsCKI1, a rice casein kinase I, in root development and plant hormone sensitivity

Casein kinases are critical in cell division and differentiation across species. A rice cDNA fragment encoding a putative casein kinase I (CKI) was identified via cDNA macroarray under brassinosteroid (BR) treatment, and a 1939-bp full-length cDNA, OsCKI1, was isolated and found to encode a putative 463-aa protein. RT-PCR and Northern blot analysis indicated that OsCKI1 was constitutively expressed in various rice tissues and upregulated by treatments with BR and abscisic acid (ABA). Enzymatic assay of recombinant OsCKI1 proteins expressed in Escherichia coli showed that the protein was capable of phosphorylating casein. The physiological roles of OsCKI1 were studied through antisense transgenic approaches, and homozygous transgenic plants showed abnormal root development, including fewer lateral and adventitious roots, and shortened primary roots as a result of reduced cell elongation. Treatment of wild-type plants with CKI-7, a specific inhibitor of CKI, also confirmed these functions of OsCKI1. Interestingly, in transgenic and CKI-7-treated plants, exogenously supplied IAA could restore normal root development, and measurement of free IAA content in CKI-deficient primary and adventitious roots revealed altered auxin content, indicating that OsCKI1 is involved in auxin metabolism or that it may affect auxin levels. Transgenic plants were less sensitive than control plants to ABA or BR treatment during germination, suggesting that OsCKI1 may be involved in various hormone-signaling pathways. OsCKI1-GFP fusion studies revealed the localization of OsCKI1 to the nucleus, suggesting a possible involvement in regulation of gene expression. In OsCKI1-deficient plants, differential gene expression was investigated using cDNA chip technology, and results indicated that genes related to signal transduction and hormone metabolism were indeed with altered expression.     

水稻AtpH基因的表达受冷抑制

采用RT-PCR差异显示法,从水稻(Oryza sativa L.)幼苗克隆了1个受冷抑制表达的cDNA片段.该片段序列与水稻叶绿体基因组编码ATP合酶CF0Ⅲ亚基的atpH基因完全同源,且覆盖了atpH基因编码区.以Northern杂交分析了水稻幼苗在冷处理不同时间后的atpH基因转录水平,结果表明,atpH基因的转录受冷抑制,在冷处理第1天就明显下降,第2天以后完全受抑制.     

水稻AtpH基因的表达受冷抑制

采用RT-PCR差异显示法,从水稻（Oryza sativaL.)幼苗克隆了1个受冷抑制表达的cDNA片段。该片段序列与水稻叶绿体基因组编码ATP合酶CF0Ⅲ亚基的atpH基因完全同源,且覆盖了atpH基因编码区。以Northern 杂交分析了水稻幼苗在冷处理不同时间后的atpH基因转录水平,结果表明,atpH基因的转录受冷抑制,在冷处理第1天就明显下降,第2天以后完全受抑制。     

Inducible Expression of Translation Elongation Factor 1A Gene in Rice Seedlings in Response to Environmental Stresses

Differences of gene expression between salinity-stressed and control rice ( Oryza sativa L. ssp. indica) cultivar "Zhaiyeqing 8" were compared using differential display PCR (DD-PCR) technique. Sequence analysis of one salt-inducible cDNA clone revealed that this clone represented a new member of rice translation elongation factor lA (eEF1A) gene family and was tentatively named REF1A. Northern blot hybridization using REF1A fragment as a probe was performed to investigate the expression of rice translation elongation factor lA gene in response to various environmental factors. It was observed that expression of the eEF1A gene in rice shoots was dramatically induced by salinity stress or exogenous application of abscisic acid (ABA). The induction of this gene by ABA stress occurred more quickly than that by salinity stress. In addition, expression of rice translation elongation factor lA gene was also induced by drought (15% PEG6000), cold (4 ℃ ) or heat-shock (37 ℃ ) stresses. The results suggested that the induction of translation elongation factor lA gene expression by environmental stresses might reflect the general adaptive response of rice plants to the adverse circumstances.     

Isolation of a 1 195 bp 5′-Flanking Region of Rice Cytosolic Fructose-1,6-bisphosphatase and Analysis of Its Expression in Transgenic Rice

A genomic DNA fragment containing the 5′-upstream sequence and part of the open reading frame corresponding to the cytosolic fructose-1,6-bisphosphatase (cyFBPase) cDNA was isolated by Genome Walking. The 1 195 bp 5′-flanking region which started from the translation initiation ATG codon was fused to reporter gene encoding β-glucuronidase (GUS) and stably transferred to rice via particle bombardment. Strong GUS activity was detected in leaves and leaf sheaths of transgenic rice, but not in culms and roots. Histochemical localization revealed that GUS expression was exclusively restricted to mesophyll cells in transgenic rice. Our results indicate that the 1 195 bp fragment contains all the cis-elements required for directing mesophyll-specific expression pattern in rice. Key words: rice (Oryza sativa); promoter;　cytosolic fructose-1,6-bisphosphatase gene; mesophyll-specific expression