重金属镉处理的水稻幼根cDNA抑制差减杂交文库的构建

重金属污染是影响水稻生长和稻米品质的重要因素,其中又以镉(Cadmium,Cd)污染最为严重,然而目前对其分子机制以及解决措施却了解甚少。本研究以水稻品种日本晴为材料,采用0.5 mmol/L镉处理水稻幼根48 h,通过运用结合分子镜像选择(mirror orientation selection,MOS)技术的抑制差减杂交(suppression subtractive hybridization,SSH)方法,构建cDNA抑制差减杂交文库,并进行microarray筛选,用于分离应答镉胁迫的差异表达基因。揭示水稻应对镉胁迫处理的分子机理,为水稻耐受镉胁迫基因的分离奠定了基础。     

海南普通野生稻ISSR-PCR反应体系的优化

通过不同浓度的Mg2+、模板DNA、dNTPs、Taq酶、引物、退火温度和循环次数的单因素试验,筛选海南普通野生稻最佳ISSR-PCR反应体系为:15μl反应体系中含2.0 mmol/L MgCl2,50 ng模板DNA,200μmol/L dNTPs,1.20 U Taq酶,1.5μmol/L ISSR引物,50 mmol/L KCl 10 mmol/L Tris-HCl(pH8.3).反应程序:94℃预变性5 min;94℃,1 min,52℃,1 min,72℃,2 min,35个循环;72℃延伸10min,4℃保存.     

水稻ⅡB型拓扑异构酶OsSpo11基因的cDNA克隆、表达谱分析及其可变剪接的检测

Spo11基因是减数分裂双链断裂（DSBs）与分裂重组的初始化基因,在基因组进化方面具有重要作用。通过改进的Nest—PCR方法,从水稻愈伤组织中克隆到了水稻DNA拓扑异构酶 OsSpo11基因的cDNA片段。克隆测序与表达研究表明,OsSpo11基因是水稻ⅡB型拓扑异构酶A亚基基因,在水稻不同组织中具有低丰度表达、组织特异性、可诱导性以及前体RNA剪接的可变性等特点。可变剪接异构体的发现,使OsPo11基因在DNA代谢、发育、遗传重组等过程中的生物学功能表现出一定的复杂性,其功能和生物学意义有待进一步研究。     

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.     

用抑制差减杂交技术分离烯丙异噻唑诱导水稻特异表达的基因

烯丙异噻唑(PBZ)处理水稻根能使其产生对稻瘟病的系统获得性抗性,因此在东南亚稻区被广泛用于防治稻瘟病,然而关于其作用的分子机理还知之甚少．运用抑制差减杂交技术,试图通过分离鉴定受PBZ诱导调控的关键基因,探索其作用的分子机理．以PBZ处理后的水稻叶片cDNA为目标群体(tester),以未处理水稻叶片cDNA为对照群体(driver),用经过对照cDNA差减的、烯丙异噻唑处理的cDNA群体构建了一个含260个重组子的差减文库．通过差示筛选鉴定出了26个。PBZ诱导水稻特异表达和增强表达的候选克隆．对26个cDNA克隆进行了双向测序和同源性比较,发现其中3个克隆：rJAB1,rTAB2和蛋白磷酸酯酶2Aδ调节亚基同型物基因,位于抗病相关信号转导途径上,它们与哺乳动物和人类免疫途径上的信号因子有明显相似之处,因此推断可能与诱导抗性有关．另外8个克隆与已知基因同源性为70％～99％．经Northern杂交分析,其中rJAB1(编码c-jun激活区结合蛋白1)受烯丙异噻唑和稻瘟菌诱导表达;膜糖蛋白同源基因及肌动蛋白(actin)α1受烯丙异噻唑诱导表达,部分克隆为低丰度转录本．     

Cu2+对水稻萌发及生理指标的影响

以常见农作物水稻为材料,在实验室条件下采用水培的方法,研究Cu2 单一污染对水稻种子的萌发及生理特性的影响.结果表明:低浓度的Cu2 对水稻生长没有明显的毒害,但随着浓度的增加,水稻受害症状比较明显。当Cu2 浓度由0增加到400 mg/L时,萌发率由对照组的43.3%降到22.2%,须根数由34个降到13个,主根长也由6.08 cm降到5.34 cm,严重影响根系的生理功能;chla和chlb的含量也显著下降,当Cu2 浓度为400 mg/L时,仅为对照组的45.1%和58.8%,影响了水稻的光合作用,而叶片的电导率显著上升,说明细胞膜的完整性和组成遭到破坏.     

光敏核不育水稻中光敏色素Ⅰ的低温荧光分析

光敏核不育水稻中光敏色素Ⅰ的低温荧光分析王伟童哲（厦门大学生物学系厦门３６１００５）（中国科学院植物研究所北京１０００４４）光敏核不育水稻（农垦５８Ｓ）是从晚粳水稻农垦５８中发现的雄性不育自然突变体，是我国特有的水稻种质资源．农垦５８Ｓ的雄性不育由光...     

Conditional and unconditional mapping of quantitative trait loci underlying plant height and tiller number in rice (Oryza sativa L.) grown at two nitrogen levels

In this study,we used 127 double haploid (DH) lines to analyze agricultural traits of rice.The DH lines,derived from a ZYQ8 (indica)/JX17 (japonica) cross by anther culture,contained 160 RFLP and 83 SSR markers.Unconditional and conditional quantitative trait loci (QTL) mapping was conducted to analyze plant height (PH) and tillers per plant (TP) at five growth stages that were grown at two nitrogen levels.Fourteen PH and 13 TP unconditional QTL were identified in the different growth stages,including 19 QTL from high-nitrogen (HN) and 14 QTL from low-nitrogen (LN) conditions.The conditional QTL for 14 genomic regions under LN/HN con-ditions showed that there was a significant effect on PH and TP across the different stages.Only one conditional QTL,ph2-3,was unable to be detected in unconditional mapping.More QTL were detected in the first four rice growth stages than in the final stage.Further-more,a line from the DH mapping population,DH78,was identified in extreme phenotypes of PH and TP that exhibited dwarfism and less-tiller (dft) characters.The gene dftl was mapped to chromosome 2 using a backcrossed population of DH78/JX17 through a map-based cloning strategy.The location of dftl coincided with the mapping region of the small-LOD peak,QTL ph2 and tp2,which were identified in plants grown in low-nitrogen conditions.Further backcrossing and fine-mapping successfully delimited the dftl locus to a 91 kb region.     

干旱对水稻根系活力与结实性状的影响

在水稻抽穗灌浆期进行人控土壤干旱,每穗实粒数,结实率,千粒重衣产量均下降,但根系活跃吸收面积与α－萘胺氧化活力均强,抗旱性强的品种,ＴＴＣ还原活力增强,而抗旱性弱的品种,ＴＴＣ还原活力降低,因此,水稻抗旱性强弱与ＴＴＣ还原活力密切相关。     

用基因枪法转bar基因以培育抗除草剂的直播稻

用基因枪法将 p CB1 质粒 (含 bar基因和 cecropin B基因 )导入到三种有应用前景的直播稻品种中 ,受体材料为来源于水稻未成熟胚的胚性愈伤组织和来源于成熟胚的悬浮细胞系 .当代转化植株显示出很强的对除草剂 Basta的抗性 ,Southern-blot分析显示 :两个外源基因共整合到转化植株基因组中 ;子一代植株中仍含有外源基因 ;来源于同一块愈伤组织的转化植株的整合模式可能是相似的 ,也可能完全不同 .