气肿疽梭菌细胞毒素CctA基因的克隆及生物信息学分析

根据GenBank中气肿疽梭菌细胞毒素CctA基因的参考序列,采用PCR方法扩增细胞毒素CctA基因。在将该基因进行克隆和测序后利用生物信息学方法,对细胞毒素CctA蛋白的理化性质、结构功能域、蛋白质二级结构和三级结构及抗原表位等重要参数进行了预测和分析。结果表明:与参考序列(JQ692583)相比,存在3处突变,核苷酸序列的同源性和氨基酸序列的同源性均为99.4%;CctA蛋白的理论等电点为8.00,不存在跨膜区和信号肽序列,二级结构以β-折叠和无规则卷曲为主,主要有4个抗原表位区域。本研究为CctA蛋白功能的深入研究提供了参考,并为气肿疽梭菌单克隆抗体的制备及合成肽疫苗的研发奠定了基础。     

Estrogen treatment enhances mesenteric lymphatic contractility in rats after hemorrhagic shock

AIM To investigate the effects of 17β-estradiol （E2） treatment on the mesenteric lymphatic microcirculation and isolated lymphatic contractility in rats after hemorrhagic shock, and to explore the relationship between contractility and the difference between intra- and extracellular calcium ion concentrations （［Ca²⁺］） of lymphatic smooth muscle cells （LSMCs）. METHODS Male Wistar rats were divided into sham group, shock group and shock+E2 group. The rats were subjected to hemorrhage ［（40±2） mmHg for 90 min］ and resuscitation with or without subcutaneous injection of E2 （2 mg/kg）. After resuscitation for 3 h, the mesenteric lymphatic microcirculation in vivo was observed. Moreover, the isolated mesenteric microlymphatic rings were prepared for the observations of lymphatic contractility evaluated by the indexes including end-systolic diameter, end-diastolic diameter, contraction frequency （CF） and passive diameter. Meanwhile, the difference between intra- and extracellular ［Ca²⁺］ of LSMCs was recorded during lymphatic contraction. RESULTS Treatment with E2 significantly enhanced the CF, total contractile fraction and lymphatic dynamics index in vivo in the rats after hemorrhagic shock, and increased the CF, the fractional pump flow and the difference between intra- and extracellular ［Ca²⁺］ of LSMCs in isolated lymphatics from the shocked rats （P<0.05）. CONCLUSION Estrogen treatment enhances lymphatic contractility in rats after hemorrhagic shock, which is related to enhancement of difference between intra- and extracellular ［Ca²⁺］ of LSMCs.     

p300/p53/Smad3 pathway involved in aging-related atrial fibrosis in human atrial fibroblasts

AIM To investigate the role of p300 in aging-related atrial fibrosis in human atrial fibroblasts （HAFs） and its potential mechanism. METHODS HAFs were obtained from human left atrial tissue, and the senescence model was established by cell passage. Senescence-associated β-galactosidase （SA-β-Gal） staining was used to detect the cell senescence, and Western blot was used to determine the protein levels of p300, p53, Smad3 and other senescence and fibrosis associated proteins in HAFs. RESULTS Compared to passage 3 HAFs, the proportion of senescent cells, and the protein levels of p300, p53, p-Smad3 and other senescence and fibrosis associated proteins were increased in HAFs at passage 7 and 11 （P<0.05）. After treated with curcumin （a p300 inhibitor） or transfection with p300 small-hairpin （sh） RNA plasmid, the protein levels of p300, and the senescence and fibrosis associated proteins were decreased in HAFs at passage 7（P<0.05）. Up-regulation of p300 by transfection with p300 over-expression plasmid increased the protein levels of p53, Smad3 and MMP-2 in HAFs at passage 3 （P<0.05）. CONCLUSION p300/p53/Smad3 signaling pathway plays an important role in aging-related atrial fibrosis in HAFs.     

生物基包膜抑制型尿素对土壤温室气体排放及小青菜产量的影响

为研究生物基包膜氮肥对土壤温室气体排放及小青菜(Brassica chinensis)产量的影响,采用密闭式静态箱-气相色谱法测定盆栽试验条件下施用不同类型包膜尿素对土壤温室气体的排放特征,探究不同类型包膜尿素对土壤温室气体排放综合增温潜势(GWP)、排放强度(GHGI)及小青菜产量的影响。结果表明:抑制型尿素(I)、生物基包膜尿素(CRU)、生物基包膜抑制型尿素(CIRU)能够显著降低N_2O、CO_2、CH_4 3种温室气体的累积排放量。与普通尿素(U)相比,处理I、CRU和CIRU的N2O累积排放量显著降低了20.79%～79.52%,CO_2的累积排放量显著降低了46.53%～62.24%,CH4的累积排放量显著降低了25.38%～30.11%。与处理U相比,处理I、CRU和CIRU的GWP分别显著降低了40.44%、60.66%和65.02%;温室气体GHGI与GWP呈现出同样的趋势,处理I、CRU和CIRU分别显著降低了26.32%、70.53%和78.95%。与其他处理相比,处理CIRU具有最优的温室气体减排效果。处理CIRU的小青菜产量最高,为1 960.00 kg·hm~(-2),较处理U显著增产68.00%。研究表明,生物基包膜抑制型尿素在提高小青菜产量的同时还可以减少菜地温室气体排放及氮素气态损失。     

甘薯矿质元素吸收与分配特性研究

在大田条件下,研究了甘薯品种北京553不同时期不同器官干物质的积累与分配规律,并分析其中各矿质元素含量的变化情况。结果表明,甘薯干物质前期积累少,中后期快速积累,并逐渐由地上部转移至块根中。各矿质元素在甘薯不同器官的积累量随植株生长而增加,但分配率不同,前期各元素集中于地上部,中后期逐渐转移至块根中。甘薯一生吸收最多的元素是氮、钾和钙,其中氮和钙主要在前中期积累且分布在地上部,钾则主要在中后期积累且分布于块根。微量元素中,甘薯对铁的需求较大,主要时期是生长中期并多分布于地上部。单位产量的需肥规律与矿质元素的吸收规律一致,可以根据需肥规律来指导生产,合理施肥。     

干旱胁迫与正常供水钾肥影响甘薯光合特性及块根产量的差异

【目的】干旱胁迫影响甘薯叶片光合特性及块根产量,研究通过施肥缓解干旱胁迫机理可为甘薯抗旱高产栽培提供理论依据。【方法】选用食用型甘薯品种“泰中6号”为材料,以硫酸钾(K2SO4)为供试肥料,水分处理设为土壤最大持水量的60%~70%(正常供水W1)和30%~40%(干旱处理W0); 钾肥设K0、 K1、 K2、 K3四个水平,K2O用量分别为0、 12.0、 24.0和36.0 g/m2。分析不同钾肥用量对不同生长时期甘薯叶片相对含水量、 叶绿素荧光参数、 光合特性及收获期块根产量的影响。【结果】在干旱胁迫和正常灌水条件下,施钾处理均显著增加了甘薯叶面积和叶片叶绿素含量,提高净光合速率(Pn),增加光合产物的生产和积累,提高块根产量和收获指数。两种水分条件下,块根产量均以K2处理最高, 干旱胁迫下K2与K3处理差异显著,正常灌水处理不显著。两种水分条件下,甘薯叶片光合参数对钾肥的响应存在显著差异,干旱胁迫下施钾使叶片水分利用效率(WUE)增大,气孔导度(Gs)降低,气孔阻力增大,蒸腾速率(Tr)和胞间CO2浓度(Ci)降低,水分蒸腾量减少; 而正常灌水条件下上述指标对钾肥的响应趋势相反。两种水分条件下施钾均可以增大叶片相对含水量(RWC),提高实际光化学效率(ΦPSII)和最大光化学效率(Fv/Fm),但是干旱胁迫下施钾增幅较大。【结论】干旱胁迫下适量施钾可以提高甘薯的抗旱性,增加甘薯产量,过量施钾使甘薯产量显著降低,而正常水分供应时,稍多钾肥对产量影响不显著。干旱胁迫与正常灌水条件下施钾对叶片光合参数的调控效应存在显著差异。施用钾肥可增大叶面积,提高叶绿素含量和光合性能,调节叶片气孔关闭,增大叶片气孔阻力,减少水分蒸腾损失,增加叶片相对含水量,提高水分利用效率和净光合速率; 施钾还能提高叶片PSⅡ原初光能转换效率和实际光化学效率,减少过剩激发能对光合机构的破坏,提高甘薯叶片的光合能力。干旱条件下钾肥的调节功能优于正常水肥供应。     

长期施肥对栗褐土有机碳矿化的影响

【目的】 土壤有机碳矿化是土壤中重要的生物化学过程,与土壤养分的释放、土壤质量的保持以及温室气体的形成密切相关。本文以 25 年长期定位施肥试验为依托,对栗褐土土壤有机碳矿化速率、有机碳累积矿化量的动态变化进行研究,为科学管理土壤肥力、增加栗褐土碳汇、减少温室气体排放提供依据。 【方法】 田间试验开始于 1988,共设置 8 个施肥处理:不施肥 (CK);单施氮肥 (N);氮磷肥合施 (NP);单施低量有机肥(M₁);低量有机肥与氮肥合施 (M₁N);低量有机肥与氮磷肥合施(M₁NP);高量有机肥与氮肥合施 (M₂N);高量有机肥与氮磷肥合施 (M₂NP)。于 2013 年玉米播种前,采集耕层 (0—20 cm) 土壤样品,采用室内培养方法,对土壤碳矿化释放 CO₂ 的数量和速率进行测定,并利用一级动力学方程计算出土壤有机碳库潜在矿化势和周转速率。 【结果】 各肥料处理不同程度地提高了栗褐土总有机碳含量,以高量有机肥与化肥配施作用最为显著。与 CK 相比,M₂N、M₂NP 处理土壤总有机碳含量增加了 121.1%、166.8%。不同处理土壤样品培养有机碳矿化速率均在第一天达到峰值,随后急剧下降。5 d 后,下降趋缓,不同处理 CO₂ 产生速率趋于一致。培养期间,各处理矿化速率变化符合对数函数关系。长期施用不同肥料均可以提高栗褐土有机碳的矿化速率,其大小顺序为:有机肥与化肥配施 > 单施有机肥 > 单施化肥 > 对照。培养 57 d 后,各处理土壤有机碳累积矿化量为 555.0～980.3 mg/kg,以 M₂NP、M₁N 的累积量较高,为对照的 1.77 倍、1.73 倍。长期施肥栗褐土有机碳矿化率呈下降趋势,以处理 M₂NP 下降最明显,与对照相比,降低了 6.3 个百分点。施肥处理土壤的潜在矿化势均高于对照,M₁N、M₂NP 最高,为 923.7 mg/kg 和 926.4 mg/kg,较对照增加了 74.0% 和 74.5%。不同施肥处理均可明显提升土壤有机碳的周转速率,减少周转时间,其中处理 M₁NP、M₂NP 效果最为明显。 【结论】 长期施用化肥、有机肥及有机无机肥配施可有效促进栗褐土有机碳的积累,提高有机碳的矿化速率和周转速率,降低有机碳的矿化率 (累积矿化量占有机碳总量的比率),加强了土壤的固碳能力,以 M₂NP 处理的效果更佳。      

新疆野苹果群体遗传结构和遗传多样性的SRAP分析

 采用SRAP标记,对中国新疆野苹果4个种下居群的群体遗传结构和遗传多样性进行了研究。结果表明：10对SRAP引物总共扩增了209条带,其中206条是多态性带（98.56%）。巩留群体、新源群体、霍城群体和裕民群体分别扩增了180、169、178和165条多态性带,巩留群体的随机交配杂合度（hs = 0.3037 ± 0.0058）最高,其次为霍城群体。UPGMA聚类分析和群体间遗传分化系数显示,巩留群体和新源群体之间,以及霍城群体和裕民群体之间,遗传关系最近,群体间遗传变异最低。新疆野苹果群体内遗传变异高于群体间,占总变异的87.9%,主坐标轴分析显示4个群体是相对独立,其中巩留群体和新源群体,霍城群体和裕民群体之间,有较高的基因交流。所有参数分析表明,巩留群体遗传多样性最丰富,故在制定新疆野苹果原地和异地种质保护计划时应优先考虑巩留群体。     

苹果属小金海棠MxIrt1基因特异性片段的原核表达及多克隆抗体的制备

 MxIrt1是从苹果属植物小金海棠中克隆出的二价阳离子转运膜蛋白基因。为进一步研究该基 因的功能, 利用MxIrt1基因位于第3和第4跨膜区之间的162 bp片段, 构建了原核表达载体pGEX-MxIrt1,经原核表达、亲和层析、获得GST2MxIRT1融合蛋白, 以融合蛋白为抗原, 制备多克隆抗体, ELISA方法检测抗体效价阳性, 蛋白质印迹检测植物体内总蛋白, 获得与预期大小一致的特异性条带。上述结果表明,表达的目的蛋白可用于免疫组织化学、蛋白质印迹检测。     

Mechanism of mesenteric lymph reperfusion aggravates multiple organ injury in superior mesenteric artery occlusion shock rats

AIM: To explore the mechanism of mesenteric lymph reperfusion (MLR) aggravates multiple organ injury in superior mesenteric artery occlusion (SMAO) shock rats. METHODS: Male Wistar rats were randomly divided into 4 groups: in sham group, only anesthetization and operation were performed; in MLR group, occlusion of mesenteric lymphatics (ML) for 1 h followed by 2 h of reperfusion; in SMAO group, occlusion of superior mesenteric artery (SMA) for 1 h followed by 2 h reperfusion; in MLR+SMAO group, occlusion of SMA and ML for 1 h followed by 2 h of reperfusion. The homogenates of liver, kidney, myocardium and lung were prepared for determining the activities of free radical, nitric oxide, myeloperoxidase (MPO) and cell membrane ATPase. RESULTS: The MDA, NO contents and NOS, MPO activities of multiple organic homogenate in SMAO and MLR+SMAO group were higher than those in sham and MLR group, and these indexes in MLR+SMAO were increased significantly than those in SMAO group. The SOD and ATPase activities of muliple organic homogenate in SMAO and MLR+SMAO group were lower than those in sham and MLR group, and those in MLR+SMAO group was decreased obviously than those in SMAO group. CONCLUSION: The MLR enhances the multiple organ free radical injury, NO synthesis and release, PMN detention and decreases the activity of cell membrane ATPase, aggravating the major organs injury in SMAO shock rats. Intestinal lymphatic pathway plays an important role in the pathogenesis of SMAO shock.