镉对DNA甲基化的影响

镉是人类致癌物,目前普遍认为镉的致癌机制与表观遗传学有关。DNA甲基化是重要的表观遗传学修饰之一。本文从镉暴露对基因组总甲基化水平、甲基转移酶、基因表达的影响三个方面综述镉暴露对DNA甲基化的影响。     

利用CpG DNA甲基化酶M.SssⅠ共表达载体制备限制性内切酶NotⅠ

限制性核酸内切酶是DNA重组的重要分子生物学工具。由于其本身对DNA有切割作用导致其重组表达在技术上十分困难,产率低、提纯程序复杂。而商业化生产所采用的利用专一型甲基化酶保护宿主DNA的限制酶表达技术流程繁琐、实用性有限。为表达NotⅠ限制酶,采用来源于Spiroplasma sp.MQ1的DNA甲基化酶M.SssⅠ特异性甲基化CpG序列,甲基化后的DNA会免受识别位点中包含CpG序列的限制酶NotⅠ的切割。将甲基化酶M.SssⅠ导入大肠杆菌表达宿主ER2566后,M.SssⅠ基因在宿主中持续表达并甲基化宿主DNA成CpG甲基化样式;利用此表达体系制备限制性内切酶NotⅠ获得成功。并借助引入纯化标签经过简便的Ni亲和层析和阴离子交换层析2步层析洗脱纯化,制备了高活力高纯度的重组限制酶NotⅠ。此表达体系可应用于一系列识别位点中包含CpG序列的限制酶的表达。     

银杏细胞转录组高通量测序及分析

利用Illumina的Genome Analyzer IIx对银杏(Ginkgo Biloba)细胞转录组进行高通量测序,挖掘银杏内酯和紫杉醇生物合成基因,特别是新的羟基化酶基因,为今后最终完善红豆杉细胞紫杉醇生物合成途径中未知的羟基化步骤作准备。通过测序,获得了银杏细胞69 286个contig,56 387个scaffold,32 032个unigene。Unigene平均长度636bp。另外从gap分布、GC含量、基因组coverage等方面对unigene进行评估,数据显示测序质量好,可信度高。通过分析unigene的表达和功能注释等信息,发现66个属于CYP450基因家族,726个参与次生代谢物合成,其中59个与萜类合成有关,17个与二萜类合成相关。利用生物信息学方法从Michigan State University银杏成熟叶、侧根、成熟果实、无菌苗以及次生茎的转录组数据中找到了与银杏细胞CYP450高度同源的紫杉烷羟基化酶候选基因15个,为后续研究奠定了基础。     

天花粉蛋白抑制人乳腺癌MDA-MB-231细胞生长及逆转syk基因甲基化的研究

目的:研究天花粉蛋白(trichosanthin,TCS)抑制人乳腺癌MDA-MB-231细胞增殖和诱导细胞凋亡过程中脾酪氨酸激酶(spleen tyrosine kinase,syk)基因甲基化状态的改变,以寻找新的去甲基化药物.方法:采用MTT法分析TCS对MDA-MB-231细胞增殖的影响,Giemsa染色法观察TCS作用后细胞形态学的变化,FCM检测不同浓度TCS作用48 h后MDA-MB-231细胞凋亡和细胞周期分布情况,甲基化特异性PCR(methylation-specific PCR,MSP)法检测MDA-MB-231细胞经TCS作用前后syk基因甲基化状态的改变,RT-PCR方法分析TCS作用后MDA-MB-231细胞中syk、Dnmts和MBD2 mRNA表达量的变化,比色法检测TCS对MDA-MB-231细胞DNA甲基转移酶活性的影响.结果:TCS体外对MDA-MB-231细胞增殖具有显著抑制作用(P<0.05),并呈时间和浓度依赖性;TCS可诱导MDA-MB-231细胞出现空泡样变性、核固缩和核碎裂等细胞凋亡形态学改变,能使MDA-MB-231细胞凋亡并可诱导细胞生长周期阻滞,凋亡率明显高于对照组(P<0.05),并呈时间和浓度依赖性.TCS可以逆转MDA-MB-231细胞syk基因启动子区甲基化状况;MDA-MB-231细胞syk mRNA表达阴性,经TCS作用后,syk mRNA表达阳性;MDA-MB-231细胞经TCS作用48 h前后Dnmts和MBD2 mRNA表达量没有明显变化(P>0.05),但Dnmts活性降低.结论:TCS在抑制MDA-MB-231细胞增殖、诱导细胞凋亡过程中对syk基因具有明显的去甲基化作用,因而可能成为新型的去甲基化药物.     

异常DNA甲基化与免疫性皮肤病

<正>表观遗传学是指在基因表达水平发生的可遗传的、无DNA序列变化的改变。其主要机制包括DNA甲基化、组蛋白修饰及非编码RNA调控,这些因素均可调控基因的转录。其中,DNA甲基化是最早被发现的一种调控机制,它在基因表达、细胞增殖、分化、发育及基因组印记等方面均起着重要作用。DNA甲基化是一项由多种酶参与的复杂分子机制,主要发生在CpG岛区域,是指S-腺苷甲硫氨酸上     

翅碱蓬响应高盐胁迫的分子机制研究

为研究翅碱蓬Suaeda heteroptera响应高盐胁迫的分子机制,试验将翅碱蓬植株在高盐胁迫(856 mmol/L NaCl)和无盐(0 mmol/L NaCl)条件下处理2 h后,对植株分别进行全转录组测序。结果表明:通过RNA-seq分析共获得转录本147 176个,拼接后获得单基因(Unigene)84 545个;比较转录组学分析显示,与无胁迫组(对照)相比,高盐胁迫下翅碱蓬共检出144个差异表达基因,其中发生表达上调的有55个,发生表达下调的有89个;GO富集分析显示,差异表达基因主要分布在次生代谢、信号转导、光合作用电子传递等途径;KEGG富集分析显示,差异表达基因涉及的代谢通路有14个;对差异表达基因发生富集的5个主要代谢通路(光合作用、蔗糖与海藻糖代谢、植物激素信号转导、苯丙基类生物合成、半胱氨酸与甲硫氨酸代谢)进行了详细阐述,从分子水平探讨了翅碱蓬响应高盐胁迫的机制。本研究结果可为更深入研究翅碱蓬耐盐分子机制、挖掘耐盐关键基因提供数据资料。     

翅碱蓬响应高盐胁迫的分子机制研究

为研究翅碱蓬Suaeda heteroptera响应高盐胁迫的分子机制,试验将翅碱蓬植株在高盐胁迫(856 mmol/L NaCl)和无盐(0 mmol/L NaCl)条件下处理2 h后,对植株分别进行全转录组测序。结果表明:通过RNA-seq分析共获得转录本147 176个,拼接后获得单基因(Unigene)84 545个;比较转录组学分析显示,与无胁迫组(对照)相比,高盐胁迫下翅碱蓬共检出144个差异表达基因,其中发生表达上调的有55个,发生表达下调的有89个;GO富集分析显示,差异表达基因主要分布在次生代谢、信号转导、光合作用电子传递等途径;KEGG富集分析显示,差异表达基因涉及的代谢通路有14个;对差异表达基因发生富集的5个主要代谢通路(光合作用、蔗糖与海藻糖代谢、植物激素信号转导、苯丙基类生物合成、半胱氨酸与甲硫氨酸代谢)进行了详细阐述,从分子水平探讨了翅碱蓬响应高盐胁迫的机制。本研究结果可为更深入研究翅碱蓬耐盐分子机制、挖掘耐盐关键基因提供数据资料。     

DNA甲基化与基因表达调控研究进展

表观遗传修饰是指不改变DNA序列的、可遗传的对碱基和组蛋白的化学修饰,主要包括DNA甲基化、组蛋白修饰、染色质重塑以及非编码RNA等。表观遗传修饰是更高层次的基因表达调控手段。DNA甲基化是一种重要的表观遗传修饰,参与基因表达调控、基因印记、转座子沉默、X染色体失活以及癌症发生等重要生物学过程。近年来随着研究方法和技术的进步,全基因组DNA甲基化的研究广泛兴起,多个物种全基因组甲基化图谱被破译,全局水平对DNA甲基化的研究不仅利于在宏观层面上了解DNA甲基化的特性与规律,同时也为深入分析DNA甲基化的生物学功能与调控奠定了基础。结合最新研究进展综述DNA甲基化在基因组中的分布模式、规律以及和基因转录的关系等。     

机动车排放PM_(2.5)对EA.hy926细胞DNA甲基化效应研究

目的探讨机动车排放PM2.5对EA.hy926细胞(人脐静脉内皮细胞和人肺腺癌细胞株A549融合的细胞株)增殖功能、DNA甲基化及凋亡的影响。方法机动车排放PM2.5对EA.hy926细胞染毒24 h。采用MTS法测定机动车排放PM2.5对细胞增殖功能的影响;DNA甲基化定量检测试剂盒(比色法)测定机动车排放PM2.5对细胞DNA甲基化的影响;Annexin V-FITC/PI染色,流式细胞仪测定机动车排放PM2.5对细胞凋亡的影响。结果机动车排放PM2.5染毒24 h可抑制细胞增殖功能,当染毒质量浓度为200μg/m L时,与PBS对照组比较差异有统计学意义(P<0.05);随着染毒质量浓度的增加细胞DNA甲基化率(5-m C%)下降,在染毒质量浓度为50μg/m L和200μg/m L时,与PBS对照组比较差异有统计学意义(P<0.05);随染毒质量浓度升高细胞诱导凋亡率逐渐升高。结论机动车尾气排放PM2.5可以抑制EA.hy926细胞的增殖功能,改变EA.hy926细胞的DNA甲基化率,对EA.hy926细胞产生诱导凋亡作用。此外DNA甲基化可能是细胞增殖和凋亡的机制,定点基因的甲基化仍需要进一步研究。     

DNA甲基转移酶在苯并[a]芘致小鼠空间学习记忆功能障碍中的改变

目的研究DNA甲基转移酶在苯并[a]芘(B[a]P)致小鼠学习记忆功能障碍中的改变,为B[a]P神经毒作用机制的研究提供科学依据。方法将48只成年雄性SPF级hauschka(ICR)小鼠按体重随机分为4组,每组12只,分别为溶剂(花生油)对照组,0.5 mg/kg(低剂量)、2.0 mg/kg(中剂量),和10.0 mg/kg(高剂量)B[a]P染毒组,隔天腹腔注射染毒1次,连续染毒30次。染毒结束后,用Morris水迷宫检测小鼠的空间学习记忆能力。并在染毒第10、20和30次后,用实时荧光定量-聚合酶链式反应(q PCR)和酶联免疫吸附试验(ELISA)检测小鼠大脑皮质DNA甲基转移酶(DNMTs)的基因表达和蛋白活性的改变。结果水迷宫结果显示,随着染毒剂量的增加,小鼠到达平台的潜伏期逐渐延长,高剂量组潜伏期显著高于溶剂对照组,差异有统计学意义(P<0.05);小鼠进入目标象限的总次数和在目标象限的游泳路程逐渐减少,高剂量组显著低于溶剂对照组,差异有统计学意义(P<0.05);与溶剂对照组相比,各染毒组小鼠大脑皮质中DNA甲基转移酶DNMT1、DNMT3a和DNMT3b的基因表达量和蛋白激酶的活性均逐渐增强,呈明显的剂量和时间依赖性,差异有统计学意义(P<0.05)。结论 B[a]P诱导大脑皮质DNA甲基转移酶DNMT1、DNMT3a和DNMT3b基因表达水平和蛋白激酶的活性增强,可能是B[a]P致小鼠空间学习记忆障碍的重要机制之一。     

Vanadium in foods and in human body fluids and tissues

Using neutron activation analysis, vanadium was analysed in a range of foods, human body fluids and tissues. On the basis of these results and those of other workers, it was concluded that daily dietary intake amounts to some tens of micrograms. Analysis of body fluids (including milk, blood and excreta) and organs and tissues provided an estimate for the total body pool of vanadium in man of about 100 μg. Vanadium was not detectable in blood and urine at the level of 0.3 ng/g, while low levels were found in muscle, fat, bone, teeth and other tissues. The relationship between dietary intake to pulmonary absorption is discussed in relation to the occurrence of vanadium in man-made air particulates. The very low levels found in milks and eggs suggest minimal vanadium requirements in growth. The findings are discussed in the light of previous results and also in relation to the possible essentiality of vanadium.     

Iodine-129 in thyroid and urine in Ukraine and Denmark

Human thyroids collected from Gomel in Belarus, sheep thyroid from Jutland and human urine from Zealand in Denmark were analysed for 129I and 127I concentrations. The ratios of 129I/127I in human thyroid in Gomel are 2.65-11.0 x 10(-9) with an average of 7.21 x 10(-9), which is one order of magnitude higher than those from Asia and South America (10(-10)), but significantly lower than those observed in west Europe (10(-8)). A weak negative correlation (P < 0.05) between 129I/127I ratio in human thyroid and the age of the subjects was observed in Gomel. The average ratio of 129I/127I in sheep thyroids from Jutland of Denmark is 1.81 x 10(-7), which is two orders of magnitude higher than those in south hemisphere, and Asia. It is also significantly higher than those observed in other west European countries before 1984 and that in human thyroid in Gomel. The high thyroid 129I level in Jutland is attributed to the release of reprocessing plants in France and UK. The 129I/127I ratios in human urine in Zealand of Denmark are 0.86-2.86 x 10(-8). The possibility of using urine 129I to evaluate the thyroid exposure to 129I is investigated.     

儿童青少年近视的防控研究进展

儿童青少年近视率一直处于高位并持续上升,严重影响了我国儿童青少年的学习和生活,本研究通过查阅国内外治疗儿童青少年近视相关著作及文献,针对目前儿童青少年近视防控手段的进展,对各治疗方案进行分析和研究。     

德国城乡景观与中国景观问题

2003年,笔者获得德国DAAD(德国国际学术交流基金会)奖励资助,作为访问学者在德国逗留了4个月(4月～8月).在这期间,笔者还考察了法国、丹麦、瑞典和意大利,但我认为还是德国的景观最为优雅和美丽.     

广义文脉与规划设计教育

城市规划设计教育不仅应关注历史化,还应关注自然、社会和经济;不仅应关注地方和传统,还应关注全球和未来。城市规划教育应掌握正确的发展观和方法论,肩负起发展地万脉的历史使命,保护社会公众的利益,推动城市持续发展。     

Perchlorate in raw and drinking water sources in England and Wales

A well‐known use of perchlorate is as a rocket fuel propellant; however, more widespread uses include in munitions and fireworks, and it also occurs naturally. Perchlorate suppresses the thyroid, which can lead to a variety of adverse effects. It is a widespread contaminant in the United States, but limited occurrence data in the United Kingdom exist, and even less for drinking water. Monitoring of 20 raw and treated drinking water sites in England and Wales, covering four seasonal periods, showed that perchlorate is a low‐level background contaminant of raw and treated drinking water. Low concentrations (treated drinking water: <0.020–2.073 μg/L, mean 0.747 μg/L) were detected at every higher‐risk site. The concentrations were comparable in each of the four sampling exercises and no significant trends were apparent relating to the time of year, the type of risk or the method of chlorination. Limited data showed that removal by ion exchange and granular‐activated carbon may occur.     

Lead and cadmium contents in cereals and pulses in north-eastern China

It is known that, unlike Japanese, Koreans or southern Chinese who depend on rice as a major source of energy for daily life, people in north-eastern China rely not only on rice, but on wheat and other cereals and to a lesser extent also on pulses. Cereal and pulse samples were collected from open markets in north-eastern China, and analyzed by inductively-coupled plasma spectrometry (ICP-MS) for two potentially hazardous heavy metals — lead (Pb) and cadmium (Cd). The average Pb level in cereals (31.3 ng Pb/g as a geometric mean) and that of pulses (25.7 ng Pb/g) were similar to each other with no significant difference. Among the cereals, Pb contents were higher in foxtail millet (54.3 ng/g) and lower in maize (35.4 ng Pb/g; grain and flour in combination), wheat flour (28.8 ng Pb/g) and rice flour (22.7 ng Pb/g). Lead levels in two important types of pulses, kidney bean and soybean (24.6 and 30.8 ng Pb/g, respectively), were comparable to the levels in rice and wheat. In contrast, Cd levels were substantially higher in pulses (55.7 ng Cd/g) than in cereals (9.2 ng Cd/g), and among the pulses, Cd in soybean (55.7 ng Cd/g) was significantly higher than that in kidney bean (23.8 ng Cd/g). The possible public health implication of the Pb and Cd levels, especially the high Pb level in foxtail millet (54.3 ng Pb/g) and the high Cd level in soybean (73.5 ng Cd/g), is discussed.     

武汉城市景观文化生态保护建设的问题与对策

城市文化生态是形成城市特色的文化要素。历史古迹,优秀建筑,空间形态,环境特色,作为城市文化生态的显现部分,既是城市文化底蕴直观生动的反映,也是市民的情感港湾和精神家园。理应得到珍惜和保护。该文在系统地概括武汉景观文化生态特点的基础上,重点分析了当前武汉城市景观规划建设中的文化生态问题,提出了武汉今后改进城市景观文化生态规划建设的对策建议。     

创新创业教育在高校实践教学中的研究与探索

大学生创新创业教育的开展对高校人才培养体系的完善具有非常重要的意义。针对目前我国高校创新创业教育模式的问题与不足进行探讨,分析研究了创新创业教育的实质,最后重点阐述了在实践教学中如何加强创新思路的引导,建立有效激励机制,开展创新及创业教育的应对策略,促进对大学生实践创新能力、创业就业能力和持续发展能力的培养。