Purification and characterization of bovine adrenal cytochrome b561 expressed in insect and yeast cell systems

Bovine adrenal chromaffin granule cytochrome (cyt) b561 is a transmembrane hemoprotein that plays a key role in transporting reducing equivalents from ascorbate to dopamine-beta-hydroxylase for catecholamine synthesis. We have developed procedures for expression and purification of functional bovine adrenal cyt b561 in insect and yeast cell systems. The bovine cyt b561 coding sequence, with or without a hexahistidine-tag sequence at the C-terminus, was cloned into the pVL1392 transfer vector under the control of the polyhedrin promoter to generate recombinant baculovirus for protein expression in Sf9 insect cells (approximately 0.5 mg detergent-solubilized cyt b561/L culture). For the yeast system, the cyt b561 cDNA was modified with a hexahistidine-tag sequence at the C-terminus, and inserted into the pPICZB vector under the control of the alcohol oxidase promoter. The recombinant plasmid was transformed into Pichia pastoris GS115 competent cells to give methanol-inducible cyt b561 expression (approximately 0.7 mg detergent-solubilized cyt b561/L culture). Recombinant His-tagged cyt b561 expressed in Sf9 or Pichia cells was readily solubilized from membrane fractions with dodecyl maltoside and purified to electrophoretic homogeneity by one-step chromatography on Ni-NTA affinity resin. The purified recombinant cytochrome from both systems had a heme to protein ratio close to two and was fully functional, as judged by comparison with the spectroscopic and kinetic parameters of the endogenous cytochrome from chromaffin granules. A novel procedure for isolation of chromaffin granule membranes was developed to utilize frozen adrenal glands instead of fresh tissue.     

Enhanced Activity of DNA Polymerase Iota in Mouse Brain Cells Is Associated with Aggressiveness

Recent studies performed with crude extracts of mouse tissues showed that the activity of DNA-polymerase iota (Pol iota) can be detected only in brain and testis extracts. To assess whether the activity of Pol iota is associated with animal behavior, we determined Pol iota activity in brain extracts of mice of two lines sharply differing in aggressiveness (RSB and RLB). We found that Pol iota activity in the mice with aggressive behavior was three times higher than in the less aggressive mice. The possible relationship between the activity of Pol iota and animal behavior is discussed.     

Histological and immunohistochemical studies of the structure of lymph nodes in Kilis goats

Ten healthy adult Kilis goat mesenteric lymph nodes were used to examine the general structure of lymph nodes, lymphocytes, plasma cells, reticular cells and reticular fibers using histological methods. We also detected T lymphocytes using anti-CD3 [SP7], anti-CD4 [74-12-4], mouse anti-bovine CD4 [CC30] and mouse anti-bovine CD8 [CC63] monoclonal antibodies (mAb); and B lymphocytes using anti-CD79a [HM57] mAb, macrophages using anti-macrophage [MAC387] mAb and follicular dendritic cells using anti-S100 polyclonal antibody (pAb). The distribution of these cells also was studied. Although the primer antibodies we used for CD3, CD8, CD79a, MAC387 and S100 worked well, the primer antibodies for CD4 were ineffective for paraffin embedded goat lymph nodes.     

Quantitative trait loci analysis of individual and total isoflavone contents in soybean seeds

Soybean isoflavones play diverse roles in human health, including cancers, osteoporosis, heart disease, menopausal symptoms and pabulums. The objective of this study was to identify the quantitative trait loci (QTL) associated with the isoflavones daidzein (DC), genistein (GeC), glycitein (GlC) and total isoflavone contents (TIC) in soybean seeds. A population of 184 F_21:0 recombinant inbred lines derived from a ‘Xiaoheidou’ בGR8836’ cross was planted in pot and field conditions to evaluate soybean isoflavones. Twenty-one QTL were detected by composite interval mapping. Several QTL were associated with the traits for DC, GeC, GlC and TIC only. QDGeGlTIC4_1 and QDGlTIC12_1 are reported first in this study and were associated with the DC, GeC, GlC and TIC traits simultaneously. The QTL identified have potential value for marker-assisted selection to develop soybean varieties with desirable isoflavone content.     

榕树须活性部位的成分研究

采用硅胶柱层析色谱和重结晶方法对榕树须95%乙醇提取物进行分离,得到8化合物,通过理化性质和波谱技术分别鉴定为:α-香树脂醇乙酸酯(1),羽扇豆醇乙酸酯(2),木栓酮(3),木栓醇(4),正三十五烷醇(5),白桦酯酸(6),β-谷甾醇(7)和豆甾-4-烯-6β-羟基-3-酮(8)。除化合物6外,其余化合物均为首次从该植物中分离得到。     

一个重要的蒙古二叠纪牙形刺种

在蒙古南部Lugiingol剖面卢金格尔(Lugiingol)组下部发现牙形刺Mesogondolell aidahoensis,该地层应归入下二叠统空谷阶,而不是像以前那样归入上二叠统。     

β-环糊精包合雨生红球藻皂化产物可行性研究

本文以雨生红球藻皂化产物中虾青素含量为评价指标,对β-环糊精包合雨生红球藻皂化产物可行性进行了实验研究。试验结果表明,当雨生红球藻粉在优选的实验条件下皂化产物经β-环糊精包合后,HPLC检测主要成分组成未见明显变化,包合率可达到90%。于温度40℃,湿度75%条件下进行稳定性加速实验,结果表明,经皂化后包合物中虾青素稳定性较好,达到了药物和保健食品原料的稳定性要求,说明该方法可行。     

低能离子束辐照下水稻中与ABA相关基因的差异表达

本文检测了用基因芯片筛选出水稻种子被低能N＋辐照后引起的差异表达的ABA代谢和信号途径相关基因。结果显示,与ABA合成相关的ZDS、Lyc-β、ZEP、NCED、SDR这五种酶的基因表达量均为上调;受ABA调控的的H＋-ATPase、NR、Rubisco的基因表达变化显著;ABA依赖的逆境应答蛋白DREB和ASR的表达量上调;受ABA信号转导调控的蛋白LEA的表达量下调,GAD和P5CS的表达量上调。这些结果表明,6×1017N＋·cm-2剂量的辐照可能促进了ABA的合成和幼苗气孔的开放,同时促进了ABA信号系统并激活或抑制了一些相关基因的表达。