An improved transformation protocol for studying gene expression in hairy roots of sugar beet (Beta vulgaris L.)

A transformation protocol, based on co-inoculation with two strains of Agrobacterium, Agrobacterium tumefaciens LBA4404 and A. rhizogenes 15834 containing a binary vector with the GUS gene, was established for the induction of transgenic hairy roots from sugar beet (Beta vulgaris L.) explants. It resulted in marked improvement in the formation of hairy roots and the integration of the binary vector T-DNA into the host genome. Of 250 inoculated sugar beet hypocotyls, 84% yielded hairy roots 5–7 days after inoculation, of which 70% were co-transformed with the binary vector T-DNA. To determine stable expression of alien genes in hairy roots, the nematode resistance gene Hs1 ^pro-1 was used as a reporter gene. In addition, molecular marker analysis was applied to monitor stable incorporation of a translocation from the wild beet B. procumbens. The molecular analysis and the nematode (Heterodera schachtii) resistance test in vitro demonstrated that the genomic structure and the expression of the Hs1 ^pro-1 -mediated nematode resistance were well-maintained in all hairy root cultures even after repeated sub-culture. Received: 25 November 1997 / Revision received: 26 May 1998 / Accepted: 15 June 1998     

Cloning of the genes encoding urease from Proteus vulgaris and sequencing of the structural genes

A fragment of chromosomal DNA from proteus vulgaris encoding urease was cloned and expressed in Escherichia coli. A 3 kbp region was sequenced and revealed three open reading frames with homology to jack bean (Canavalia ensiformis) urease. The smallest protein (11 kDa) was homologous to the N-terminus of the plant enzyme and the largest polypeptide (61 kDa) corresponded to the C-terminus. The large protein contained conserved regions and a cysteine residue which is known to be catalytically important in the plant enzyme. A protein of 12 kDa showed homology to residues 132 to 237 of jack bean urease.     

Biostimulant activity of humic-like substances from agro-industrial waste on Chlorella vulgaris and Scenedesmus quadricauda

The use of microalgae in a number of sectors, including biodiesel, feed and food production, is proving to be of great interest. An evaluation was made of the possible biostimulant effects on Chlorella vulgaris and Scenedesmus quadricauda of humic-like substances (HLs) extracted from agro-industrial wastes. These included digestate from the waste of an agro-livestock farm (D-HL), oil extraction residues from rape (B-HL, Brassica napus L.) and tomato residues (T-HL). The microalgal growth medium (BG11) was supplemented with HLs to evaluate their effect on biomass yield as well as carbohydrate, chlorophylls a and b, lipid and fatty acid contents. Our results showed that the HLs used in the test are effective biostimulants of C. vulgaris and S. quadricauda. The biostimulant effect seems to depend on the type of extract used for cultivating the microalgae, the concentration and the species treated. Among the extracts applied to the growth medium, D-HL and T-HL seem to have a significant effect on microalgal biomass and lipid production. Although B-HL showed no significant effect on the biomass and lipid content of C. vulgaris and S. quadricauda, its presence in the growth medium increased the saturated:unsaturated fatty acid ratio (SFA/UFA) and stimulated the sugar metabolism of the microalgae by increasing their carbohydrate and chlorophyll content.     

菜豆环氧化物水解酶的表达及其催化特性研究

【背景】光学纯环氧化物及邻二醇是一类多功能手性砌块。与化学合成法相比,环氧化物水解酶(EHs)介导的生物转化法因环境友好而成为当前的研究热点。【目的】从菜豆(Phaseolus vulgaris)中克隆一种EH基因并进行原核表达,研究重组酶催化环氧苯乙烷(Styrene oxide,SO)的水解特性。【方法】通过计算机辅助分析EHs的一级结构,推测一种菜豆来源的未知功能蛋白(PvEH4)可能具有EH活性。利用RT-PCR技术,以菜豆总RNA为模板,扩增编码PvEH4的基因pveh4,并实现其在Escherichia coli BL21(DE3)中的表达。利用重组菌E.coli/pveh4全细胞催化SO水解,分析PvEH4的对映选择性和区域选择性。【结果】一级结构分析表明,PvEH4具有α/β折叠型EH特有的保守区域。SDS-PAGE结果显示PvEH4的表观分子量为39.4 kD。PvEH4针对SO的对映选择率(E值)为10.1,区域选择性系数αS和βR分别为99.5%和82.5%。当外消旋SO转化率达到68.1%时,可同时获得99.9%ees的(R)-SO和92.3%eep的(R)-苯乙二醇(Phenyl-1,2-ethanediol,PED),两者的产率分别为31.9%和65.6%。【结论】PvEH4的挖掘不仅增加了植物类EHs的数量,同时也为EHs的蛋白分子改造提供参考。     

热带普通小球藻培养模式的筛选及其培养基的优化

【背景】从海南热带海区中分离得到一株微藻,其生长速度快、适应力强,经鉴定该微藻为普通小球藻。【目的】提高热带普通小球藻的生长速率。【方法】以"宁波大学3#微藻培养液配方"为基础培养液,分别添加有机碳(C6H12O6和CH3COONa)对热带普通小球藻进行自养、兼养及异养培养,获得促进热带普通小球藻快速生长的培养方式。在"宁波大学3#微藻培养液配方"的基础上对热带普通小球藻的兼养培养基配方进行优化,并用优化兼养培养基与"宁波大学3#微藻培养基"对比培养热带普通小球藻。【结果】添加6 g/L CH3COONa的兼养模式促进热带普通小球藻生长效果最好;优化的兼养培养基配方为:6 g/L CH3COONa,20 mg/L(NH4)2SO4-N,5 mg/L Na H2PO4-P,3 mg/L Fe SO4-Fe,1 mg/L Vitamin B1和0.000 5 mg/L Vitamin B12。对比培养实验结果显示,培养第6天,兼养培养液收获的生物量(细胞密度)达4.20×107 cells/m L,是"宁波大学3#配方微藻培养液"的2.30倍。【结论】兼养培养模式为热带普通小球藻的最佳培养模式,优化的兼养培养基极显著地提高了热带普通小球藻的生物量(P0.01)。     

Sulfoximines as potent RORγ inverse agonists

Progress in the identification of suitable RORγ inverse agonists as clinical candidates has been hampered by the high lipophilicity that seems required for high potency on this nuclear receptor. In this context, we decided to focus on the replacement of the hydroxymethyl group found on known modulators to determine if more polarity could be tolerated in this position. SAR of the replacement of this moiety is presented in this article leading to the identification of sulfoximine derivatives as potent modulators with pharmacological activity in the in vivo mouse Imiquimod psoriasis model.     

Discovery of super soft-drug modulators of sphingosine-1-phosphate receptor 1

The oral S1PR1 agonist ponesimod demonstrated substantial efficacy in a phase II clinical trial of psoriasis. Unfortunately, systemic side effects were observed, which included lymphopenia and transient bradycardia. We sought to develop a topical soft-drug S1PR1 agonist with an improved therapeutic index. By modifying ponesimod, we discovered an ester series of S1PR agonists. To increase metabolic instability in plasma we synthesised esters described as specific substrates for paraoxonase and butyrylcholinesterases, esterases present in human plasma.     

对称性肢端角化病临床研究-附9例报道

目的:报道一组以肢端出现对称性、棕褐色、角化过度性斑片病例,探讨发病原因和治疗方法。方法:系统收集自2012年10月至2016年6月期间诊疗的以对称性肢端角化性皮损为表现的9例患者的临床资料,包括年龄、性别、职业、家族成员情况、临床表现、皮损组织病理学改变、治疗、治疗转归等。结果:9例患者中男8例,女1例,平均年龄27.4岁(13~43岁),病程2个月至15年,平均(4.5±4.3)年。有1例男性患者有一同胞姐姐有类似病史。临床主要表现为红褐色角化性斑片,每年7、8月开始发病,10月左右开始缓解并渐完全消退。部分手部有轻度瘙痒不适。皮肤组织病理学检查表现为明显的网篮状角化过度,轻度乳头瘤样增生,真皮浅层血管周围稀疏淋巴细胞浸润。1例用异维A酸治疗短期有效。结论:对称性肢端角化病主要以肢端出现对称性角化斑为主,常伴随寻常型鱼鳞病;遗传因素和环境因素在发病中的作用有待阐述;异维A酸可作为一种治疗选择。