Molecular genetic characterization of rice seed lipoxygenase 3 and assessment of its effects on seed longevity

Lipoxygenases (LOXs) are enzymes involved in lipid peroxidation. Here we reported the identification, molecular and functional characterization of the gene encoding rice (Oryza sativa L.) seed LOX3 (sLOX3). Via a map-based cloning strategy we identified Os03g0700400 as the candidate gene encoding sLOX3. Further functional complementary test and biochemical characterization of the recombinant Os03g0700400 protein verified the identification. The sLOX3 gene was highly expressed in roots, moderately in embryos and very weakly in leaves, leaf sheaths and stems. Transient expression experiment (in rice protoplasts) and subsequent laser confocal microscopic analysis demonstrated that the sLOX3 protein was localized into the cytosol. We next showed that overexpression of sLOX3 in a japonica sLOX3-normal rice cultivar, Wuyunjing 7 accelerated the decrease of seed germination ability when the seeds were routinely stored, which demonstrated that sLOX3 had a negative effect on seed longevity (storability). Meanwhile, an increased occurrence of embryo decay was observed in the same transgenic seeds, suggesting that sLOX3 might negatively affect seed longevity by facilitating colonization of particular seed pathogens. Our result forwarded the understanding of the effects of 9-LOX on rice seed longevity.     

害虫微生物防治的目标体系与效益评价方法

害虫微生物防治系统是复杂的多目标复合生态系统．对系统进行多目标效益评价有助于了解系统现状,遴选最佳防治技术,根据害虫防治的一般要求,结合微生物防治的有关特点,建立了害虫微生物防治的目标体系．对基于估计相对位置的方案排队法和混合优序图法分别进行改进,用改进算法对系统进行效益评价和方案择优,给出了计算软件．同时指出,在生态系统的评价管理中,多目标决策方法是科学合理、行之有效的定量工具．     

Characterization of glycerol dehydratase expressed by fusing its α- and β-subunits

The gdh and gdhr genes, encoding B₁₂-dependent glycerol dehydratase (GDH) and glycerol dehydratase reactivase (GDHR), respectively, in Klebsiella pneumoniae, were cloned and expressed in E. coli. Part of the β-subunit was lost during GDH purification when co-expressing α, β and γ subunit. This was overcome by fusing the β-subunit to α- or γ-subunit with/without the insertion of a linker peptide between the fusion moieties. The kinetic properties of the fusion enzymes were characterized and compared with wild type enzyme. The results demonstrated that the fusion protein GDHALB/C, constructed by linking the N-terminal of β-subunit to the C-terminal of α subunit through a (Gly₄Ser)₄ linker peptide, had the greatest catalytic activity. Similar to the wild-type enzyme, GDHALB/C underwent mechanism-based inactivation by glycerol during catalysis and could be reactivated by GDHR. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

牛病毒性腹泻病毒的成熟和释放

试验中用电镜观察了牛病毒性腹泻病毒ＯｒｅｇｏｎＣ２４Ｖ株在感染新生牛睾丸细胞中的形态发生。成熟的病毒颗粒是直径约为５０ｎｍ的球形颗粒,内含直径约为３０ｎｍ的核心。病毒在宿主细胞的胞质内复制,通过糙面内质网膜出芽成熟。病毒可以通过外排或在细胞死亡后含有病毒颗粒的空泡崩溃而释放到胞外。     

A bifunctional xylanase encoded by the xynA gene of the rumen cellulolytic bacterium Ruminococcus flavefaciens 17 comprises two dissimilar domains linked by an asparagine/glutamine-rich sequence

The nucleotide sequence of the xynA gene of Ruminococcus flavefaciens 17 was determined and found to consist of a 2862bp open reading frame beginning with a TTG start codon. The predicted product, XYLA, consisted of distinct amino-terminal (A) and carboxy terminal (C) domains (248 amino acids, including a putative signal sequence, and 332 amino acids, respectively) linked by a repetitive sequence (B, 374 amino acids) extraordinarily rich in asparagine (45%) and glutamine (26%) residues. Domains A and C were shown to be capable of expressing xylanase activity independently of each other when suitably truncated derivatives of the xynA coding region were expressed as lacZ fusions. The activities associated with the two domains were shown to differ with respect to the average size of hydrolysis products formed from oat-spelt xylan, with domain C releasing relatively more xylose and domain A more xylo-oligosaccharides. The amino acid sequence of domain A of XYLA closely resembled that of the Bacillus pumilus xynA enzyme (45% identical residues). On the other hand domain C showed significant similarity (33% to 40% identical residues) to a different group of bacterial xylanases and exoglucanases exemplified by the Caldocellum saccharolyticum xynA and celB products. The xynA product is, therefore, a bifunctional enzyme having two dissimilar catalytic domains capable of acting on xylan.     

Breeding bacterial blight-resistant hybrid rice with the cloned bacterial blight resistance gene Xa21

The cloned bacterial blight (BB) resistance gene Xa21 was transferred into Minghui63, a widely used restorer line of indica hybrid rice in China, through an Agrobacterium-mediated system. Molecular and resistance analyses revealed that the Xa21 gene was integrated in the genomes of transgenic plants and their progeny inherited resistance stably. For the purpose of hybrid breeding, Xa21 transgenic homozygous restorer lines were selected through `within-lane' dosage comparison of hybridization signal in combination with PCR and resistance analyses. The selected transgenic restorer lines were then crossed with a commonly used sterile line, Zhenshan97A, to produce Xa21 transgenic hybrid rice, Shanyou63-Xa21. The hybrid rice plants with Xa21 displayed high broad-spectrum resistance to Xanthomonas oryzae pv. oryzae (Xoo) races and maintained elite agronomic characters of Shanyou63. The propagation of this BB-resistant hybrid variety with Xa21 will benefit rice production.     

花生根瘤菌类菌体氢氧化体系的研究

花生根瘤菌类菌体含有吸H₂酶,以O₂或亚甲蓝为电子受体均表现吸H₂活性。类菌体还原减氧化的吸收差示光谱在424、500、520、550、560、585及595 nm处呈现吸收峰,表明细胞色素c、b、和o参与H₂的氧化过程。CN-明显影响520、550、560 nm处的吸收峰,意味着有些细胞色素c和b对CN^-敏感,在H₂代谢中充当末端氧化酶的作用。抗霉素A、HONO、CN^-、N₃^-和DBMIB均强烈抑制吸H₂活性,但鱼藤酮不抑制吸H₂活性。说明细胞色素b、c、a和泛配参与H₂氧化的电子传递,而NADH脱氢酶不参与。花生根瘤菌类菌体的H₂氧化系统是个复杂具分支的电子传递体系。     

人骨髓间充质干细胞分化为运动神经元过程中Nestin与Hb9表达的研究

目的探讨全反式维甲酸(RA)与音猬因子(Shh)诱导人骨髓间充质干细胞向神经干细胞乃至运动神经元方向分化过程中Nestin与Hb9的表达变化情况。方法采用传至第3代的人骨髓间充质干细胞,经含有RA与Shh的诱导液处理后,应用免疫细胞化学及荧光定量PCR方法分析Nestin与Hb9表达的变化情况。结果人骨髓间充质干细胞经过RA与Shh诱导后Nestin与Hb9的表达明显高于未加RA与Shh的对照组及空白组。结论 hBMSC具有向神经干细胞乃至运动神经元方向分化的潜能,RA与Shh可以在体外诱导hBMSC定向向该方向分化。     

银杏苦内酯B对缺血豚鼠心室肌动作电位、L-型钙电流和延迟整流钾电流的作用

目的:研究银杏苦内酯B对正常和缺血心室肌细胞动作电位(action potential,AP),L-型钙电流(L-type calcium current,ICa-L)、延迟整流钾电流(Delayed Rectifier Currennt,IK)的影响.方法:用常规细胞内微电极方法记录豚鼠心室肌细胞动作电位,用全细胞膜片钳技术记录游离心室肌细胞离子流.结果:①在生理条件下,银杏苦内酯B可缩短心室肌细胞动作电位时程 (action potential duration,APD),但对AP其他参数无影响,银杏苦内酯B可增大IK,呈浓度依赖性,但对ICa-L无显著作用;②在缺血条件下,APD50、APD90明显缩短,RP、APA减小,Vmax减慢,而银杏苦内酯B则可延缓和减轻缺血所引起上述参数的变化;3.在缺血条件下,IK和ICa-L均受到抑制,但加入银杏苦内酯B后可逆转缺血所造成这两种离子流的减小.结论:银杏苦内酯B可对抗心肌缺血所引起的心肌电生理的变化,提示银杏苦内酯B可预防心律失常的发生.     

乙醇胁迫抑制毕赤酵母表达外源蛋白的转录组学分析

在5 L发酵罐中进行毕赤酵母发酵表达猪?干扰素的实验,发现甘油培养末期乙醇的积累会抑制外源蛋白的表达。从转录组学角度系统分析不同浓度乙醇胁迫条件下,毕赤酵母甘油培养期和甲醇诱导期细胞的生理状态变化。研究结果表明,在甘油培养期,乙醇胁迫使得毕赤酵母细胞中的545个基因发生了显著差异表达(265个基因表达上调,280个基因表达下调),这些差异表达基因的功能主要涉及蛋白质合成、能量代谢、细胞周期和过氧化物酶代谢。乙醇胁迫增加了蛋白质错误折叠的情况,降低了核糖体和线粒体的结构完整性,使得甘油培养末期无法得到大量具有健全功能的酵母细胞。在甲醇诱导期,与甲醇代谢、蛋白质加工合成、氨基酸代谢等途径相关的294个基因发生了显著差异表达(171个基因表达上调,123个基因表达下调),导致内质网胁迫不能被及时解除,破坏了细胞内的氨基酸正常代谢。