食品微生物学实验课立体化教学体系的构建

通过对食品微生物学实验课教学内容、教学手段和方法以及考核方法等方面进行改革,构建了一套基础性实验—综合提高性实验—创新性实验的立体化实验教学体系,使其教学系统化、课程层次化、内容多样化,使学生的综合能力得到了良好的训练,培养了创新意识和综合实验能力,达到了较好的教学效果.     

Spironolactone inhibits apoptosis in rat mesangial cells under hyperglycaemic conditions via the Wnt signalling pathway

Mesangial cells (MCs) play a crucial role in maintaining structure and function of glomerular tufts, providing structural support for capillary loops and modulating glomerular filtration by their contractility. MCs apoptosis occurs in experimental diabetic nephropathy, and this correlates with worsening albuminuria. Accumulating evidence suggests that mineralocorticoid receptor (MR) blockade effectively reduces proteinuria in diabetic nephropathy; however, it is rarely known whether spironolactone (SPI), a nonspecific MR antagonist, inhibits apoptosis in MCs under hyperglycaemic conditions. The objectives of this study are to determine the relationship between SPI and apoptosis, and investigate the cell signalling pathway by which SPI inhibits apoptosis. Rat MCs were treated with 30 mM d-glucose and 10^?8, 10^?7 or 10^?6 M aldosterone (ALD) for 24 h. In some experiments, MCs were pretreated with 10^?7 M SPI or 10 mM LiCl for 1 h. Apoptosis was evaluated by cell nucleus staining and flow cytometric analyses, and caspase-3 activity was assayed. Gene and protein expression were quantified using quantitative real-time PCR and Western blotting, respectively. SPI directly inhibited high glucose and ALD-induced MCs apoptosis in a caspase-dependent manner. Importantly, SPI inhibited MCs apoptosis via the Wnt signalling pathway. SPI promoted activation of the Wnt signalling pathway in MCs, leading to upregulation of Wnt4 and Wnt5a mRNA expression, decreased GSK-3β protein expression and increased β-catenin protein expression. As a conclusion, this study suggests that SPI may inhibit apoptosis in MCs during hyperglycaemic conditions via the Wnt signalling pathway. Blockade of the ALD system may represent a novel therapeutic strategy to prevent MCs injury under hyperglycaemic conditions.     

Structural divergence is more extensive than sequence divergence for a family of intrinsically disordered proteins

The p53 transactivation domain (p53TAD) is an intrinsically disordered protein (IDP) domain that undergoes coupled folding and binding when interacting with partner proteins like the E3 ligase, MDM2, and the 70 kDa subunit of replication protein A, RPA70. The secondary structure and dynamics of six closely related mammalian homologues of p53TAD were investigated using nuclear magnetic resonance (NMR) spectroscopy. Differences in both transient secondary structure and backbone dynamics were observed for the homologues. Many of these differences were localized to the binding sites for MDM2 and RPA70. The amount of transient helical secondary structure observed for the MDM2 binding site was lower for the dog and mouse homologues, compared with human, and the amount of transient helical secondary structure observed for the RPA70 binding site was higher for guinea pig and rabbit, compared with human. Differences in the amount of transient helical secondary structure observed for the MDM2 binding site were directly related to amino acid substitutions occurring on the solvent exposed side of the amphipathic helix that forms during the p53TAD/MDM2 interaction. Differences in the amount of transient helical secondary structure were not as easily explained for the RPA70 binding site because of its extensive sequence divergence. Clustering analysis shows that the divergence in the transient secondary structure of the p53TAD homologues exceeds the amino acid sequence divergence. In contrast, strong correlations were observed between the backbone dynamics of the homologues and the sequence identity matrix, suggesting that the dynamic behavior of IDPs is a conserved evolutionary feature. Proteins 2013; 81:1686–1698. © 2013 Wiley Periodicals, Inc.     

淡足侧沟茧蜂寄生对甜菜夜蛾幼虫血淋巴总糖、蛋白质及虫体脂质含量的影响

为探讨淡足侧沟茧蜂Microplitis pallidipes Szepligeti调控寄主的生理机制,测定了淡足侧沟茧蜂寄生后,甜菜夜蛾Spodoptera exigua(Hübner)幼虫血淋巴总糖、蛋白质及虫体脂类含量的变化。研究结果表明,寄生后连续5 d的观察时间内,甜菜夜蛾幼虫血淋巴总糖含量从寄生后第1天开始就高于未寄生寄主幼虫,且在寄生后第2至5天达到显著水平;除寄生后第3天外,被寄生寄主幼虫血淋巴总蛋白质含量始终低于未寄生幼虫,且在寄生后第1、4、5天达到显著水平;甜菜夜蛾被寄生后,虫体脂质含量始终高于未寄生幼虫,且从寄生后第2天开始达到显著水平。本研究揭示,淡足侧沟茧蜂寄生刺激了寄主甜菜夜蛾幼虫血淋巴总糖合成,但抑制了其蛋白质合成,同时也刺激了甜菜夜蛾虫体脂质合成。     

The stimulatory effect of ROCK inhibitor on bovine corneal endothelial cells

Reagents which can promote the proliferation, adhesion and migration of cultured corneal endothelial cells (CECs) will be helpful for the treatment of reduced visual acuity due to CECs deficiency. The objectives of this study were to investigate the potential use of an inhibitor of Rho-associated protein kinase (ROCK), Y-27632, to cultured bovine corneal endothelial cells (B-CECs) and evaluated its effects on the proliferation, adhesion and migration of B-CECs. The proliferation of cultured B-CECs was moderately enhanced by 10 μM Y-27632. Y-27632 induced fibroblast-like morphological changes in the cultured B-CECs and normal cell morphology could recover after Y-27632 removal. In addition, Y-27632 was found to significantly enhance the adhesion and migration of B-CECs. Furthermore, the hanging drop aggregation assay showed that Y-27632 promoted B-CECs to form cellular networks and sheets, which proliferated along the liquid–air interface and migrated to the surface of the lid of dish. Our study demonstrated that Y-27632 is a potentially powerful reagent which can enhance the proliferation of cultured B-CECs. Y-27632 will be useful in CEC injection therapy and topical application for CEC deficiency.     

茅苍术叶片可培养内生细菌多样性及其促生潜力

对江苏省道地药材茅苍术叶片可培养内生细菌的多样性及其固氮、解磷、解钾、产生长素的能力进行研究。依据菌落形态的不同,共分离得到52株内生细菌。能正常传代培养的45株内生细菌经ARDRA分析后归入14个聚类簇,簇内菌株的BOX-PCR指纹图谱相似度不高,在属水平上显示出茅苍术内生细菌丰富的多样性。各聚类簇代表菌株16S rDNA的序列分析表明分离得到的内生细菌与泛菌属、微杆菌属、短杆菌属、农杆菌属、假单胞菌属、芽孢杆菌属细菌亲缘关系相近,优势内生细菌与假单胞菌属细菌亲缘关系相近。45株能正常传代培养的内生细菌中,有10株能够在无氮培养基上正常生长,具固氮潜力。使用nifH基因通用引物对其基因组进行扩增后,除ALEB 33外,其它9株内生细菌均可获得与nifH基因片段大小相近的条带。分别使用NBRIP培养基和蒙金娜有机磷培养基筛选后获得19株和15株能够溶解磷酸钙和卵磷脂的内生细菌,其中ALEB 43溶解无机磷的能力最强,达(251.43±6.55)mg/L;ALEB 4A溶解有机磷的能力最强,达(23.63±1.46)mg/L。部分内生细菌溶解无机磷的能力与其产酸能力呈正相关,而菌株溶解有机磷的能力却无此相关性。通过硅酸盐培养基的筛选,获得具有解钾潜力的菌株24株。43株内生细菌能够将色氨酸转化为生长素,其中ALEB 44产生长素的能力最强,达(268.44±10.12)μg/mL。本研究首次揭示了江苏省道地药材茅苍术体内丰富的内生细菌资源及其促生长潜力,对进一步阐述茅苍术与内生菌之间的相互关系具有重要意义。     

氢化物发生—原子荧光光谱法测定桉树叶、皮、躯干、根中硒含量

以铁氰化钾为掩蔽剂,1.5% KBH4为还原剂,10%的盐酸为载流液,微波消解处理样品,氢化物发生-原子荧光法(HG-AFS)分别测定桉树叶、皮、躯干和根中硒元素含量。加标回收验证了结果的准确性。考察了仪器测定硒的检出限。并对铁氰化钾,聚环氧琥珀酸(PESA),酒石酸,柠檬酸,乙二胺五种掩蔽剂对11种常见干扰元素的掩蔽效果进行了探究,为优良掩蔽剂的选择提供了参考性资料。     

宽口光唇鱼微卫星位点的筛选与特征分析

通过FIASCO法(Fast Isolation by AFLP Sequences Containing Repeats)筛选宽口光唇鱼Acrossocheilus monticola(Günter)基因组微卫星位点,利用生物素标记的寡核苷酸探针(AC)8、(CT)8、(GGT)8、(GATA)8和(GATT)7首次成功构建宽口光唇鱼基因组微卫星富集文库。从文库中共筛选495个克隆测序,成功设计163对微卫星引物,经PCR扩增检测,获得了18个多态性微卫星标记。利用这18对多态性引物,分析了赤水河赤水市宽口光唇鱼种群的遗传多样性,数据显示:18对多态性微卫星引物的等位基因数为8～31个,平均等位基因数为19.6个,平均期望杂合度为0.8701,平均观测杂合度为0.8312,其中引物Am07、Am35、Am47、Am69、Am78和Am127显著偏离哈迪温伯格平衡(P<0.05),以上数据表明赤水河赤水市宽口光唇鱼种群的遗传多样性水平较高。筛选的微卫星标记对于宽口光唇鱼的遗传背景分析和生物种质资源的保护有重要作用。