植物对重金属砷和镉胁迫适应的分子机理初探

随着现代工业的发展，重金属污染日趋严重。重金属污染引发的环境和健康问题在许多国家都有报道，我国的重金属污染状况也不容乐观。土壤和水体中的重金属污染可以通过食物链进入人体，对人类健康造成很大的危害，如诱发癌症 和畸胎等。 植物修复是一种利用植物对重金属或有机污染物的超富集能力清除或减低污染的环境生物技术。植物修复的生物学机制的研究为这项技术走向实用化奠定了基础。植物修复近期的进展可能来自于可更有效地富集重金属的植物品种的选择、土壤条件的改善等;但长远看来，植物修复技术的巨大进步将取决于新的可更好地抵抗重金属或降解有机毒物的基因的鉴定和克隆，并通过转基因技术创造一批新的植物品种，如可迅速大量富集重金属的高生物量的用作环境净化的植物，以及可排拒重金属吸收的粮食、蔬菜和水果等作物。 本研究针对砷污染的植物修复机制，以超富集砷的凤尾蕨属植物——蜈蚣草为试材取得了如下进展： 1． 以从砷污染地区采集的蜈蚣草（Pteris vittataL．）为植物材料，利用抑制消减杂交(SSH)分离了经砷诱导处理与其对照间表达有差异的cDNA片段，以期得到与砷富集密切相关的基因。其中筛选到的一个cDNA片段与ABC transporter (ATP-binding cassette transporter)有较高的同源性。通过RACE方法对该基因进行了克隆，并进行了初步的结构和功能分析。结果表明所获得的PvABCTl (Accession No. AY496966)为一全长cDNA，长度为2165 bp，其中开读框架为1791 bp，编码597个氨基酸。该基因所编码的蛋白中含有2个ABC transporter特性结构域，1个ATP-binding cassette和2个ATP/GTP结合位点(P-loop)，没有明显的跨膜区。 2． 对蜈蚣草在砷胁迫下PvABCT1基因的表达模式进行了研究。转录水平分析表明PvABCT1的表达受砷的诱导。进一步通过PvABCTl-GFP融合基因在洋葱细胞中的表达进行亚细胞定位，结果显示该基因可能定位于细胞质中。 3． 为了研究所克隆的PvABCT1基因的功能，本研究构建了PvABCT1的酵母表达载体，把该基因转入因ACR3基因缺失而对砷敏感的酵母突变株。酵母功能互补实验表明PvABCT1不仅不能与ACR3基因功能互补，反而使酵母对砷的敏感性增加，同时酵母细胞中的砷含量较未转化的酵母细胞增加。即在转入PvABCT1后，酵母细胞吸收了更多的砷。这暗示该基因与蜈蚣草中砷的高吸收有关。 针对食品重金属污染问题，本研究探讨了减低蔬菜对重金属吸收的方法及其 作用机理，取得了如下进展： 1．研究了钙离子和镧离子对镉离子胁迫下生菜种子萌发和植株生长的影响，结果表明在种子萌发时外施4 mM CaCI2或0.04 mg/L La(N03)3均可提高生菜对重金属镉的抗性。 2．通过检测0.5 mM CdCl2胁迫下生菜植株中的镉含量以及外施钙离子或镧离子后相应的镉含量，发现4 mM CaCl2可以增加镉胁迫下生菜植株中镉的积累;而0.04 mg/L La(N03)3可以降低镉胁迫下生菜植株中镉的积累。 3．对生菜中植物络合素合酶基因进行了克隆，通过RT-PCR分析以及植物络合素( phytochelatins，PCs)的检测，探讨了外施钙离子或镧离子对镉胁迫下生菜植株中植物络合素合酶基因在转录水平的表达量、植物络合素含量以及镉的积累三者之间的关系。结果表明：4 mM CaCl2可以提高镉胁迫下生菜植株中植物络合素合酶基因在转录水平的表达以及植物络合素的含量，增加镉的积累;而0.04 mg/L La(N03)3虽然同样可以提高植物络合素合酶基因在转录水平的表达以及植物络合素的含量，却能降低镉胁迫下生菜植株中镉的积累。这暗示外施钙离子可以促进用于重金属污染环境修复的植物对重金属的吸收，而外施镧离子可以用于降低叶菜类蔬菜中重金属镉的积累。

砷超富集植物蜈蚣草(Pteris vittata L.)中砷解毒机制的研究

砷是一种具有致癌、致畸、致突变的有毒元素，在地表的含量本来很低。然而，随着现代社会的发展和工业活动的增加导致砷污染日趋严重。土壤和水体中的砷污染可以通过食物链进入人体，对人类的健康造成极大的危害。植物修复是一种利用植物对污染物的超富集能力来清除或减低污染的新型环境生物技术。植物修复的实际应用依赖于超富集植物的发现和超富集机制的阐明，特别是砷解毒过程(砷的吸收、还原和区域化)的研究及相关基因的克隆。砷超富集植物蜈蚣草(Pteris vittata L.)中砷解毒机制的阐明将为砷污染的植物修复及新型工程植物的研发提供理论基础。 本论文以蜈蚣草为试材，针对蜈蚣草的砷解毒机制取得了如下研究进展： 1.以砷超富集植物蜈蚣草为材料，建立了一个适于研究蜈蚣草砷吸收和解毒机制的新系统—愈伤组织悬浮培养体系。首次证明蜈蚣草愈伤组织与其孢子体及配子体一样具有对砷的抗性和砷超富集的能力。 2.以蜈蚣草愈伤组织为材料，通过比较亚砷酸盐、砷酸盐和二甲基胂酸盐对蜈蚣草和拟南芥植物毒性的差异，表明砷的还原可能是蜈蚣草对砷解毒的重要机制之一而砷的甲基化对蜈蚣草的砷解毒作用甚微。 3.以蜈蚣草愈伤组织为材料，通过对砷在蜈蚣草愈伤组织细胞中的亚细胞定位，首次直接证明植物液泡对砷具有非常明显的区隔化作用。暗示区隔化作用在蜈蚣草对砷的解毒过程中发挥着重要的作用。 4.通过测定蜈蚣草愈伤组织对不同化学态的砷处理下抗氧化物质的变化发现酸溶性巯基在蜈蚣草砷解毒中也发挥着重要作用。 5.以蜈蚣草愈伤组织为材料，发现磷和砷的吸收在高浓度范围下（﹥0.2 mM）存在明显的协同效应。对蜈蚣草高亲和磷酸盐转运蛋白基因－PvPHT基因功能的初步分析则表明PvPHT参与了蜈蚣草对磷和砷的吸收过程。

Evolution from point defects to arsenic clusters in low-temperature grown GaAs/AlGaAs multiple quantum wells

Optical transient current spectroscopy (OTCS), photoluminescence (PL) spectroscopy and excitonic electroabsorption spectroscopy have been used to investigate the evolution of defects in the low-temperature grown GaAs/AlGaAs multiple quantum well structures during the postgrowth rapid thermal annealing. The sample was grown at 350 degrees C by molecular beam epitaxy on miscut (3.4 degrees off (001) towards (111)A) (001) GaAs substrate. After growth, the sample was subjected to 30s rapid thermal annealing in the range of 500-800 degrees C. It is found that the integrated PL intensity first decreases with the annealing temperature, then gets a minimum at 600 degrees C and finally recovers at higher temperatures. OTCS measurement shows that besides As,, antisites and arsenic clusters, there are several relatively shallower deep levels with excitation energies less than 0.3 eV in the as-grown and 500 degrees C-annealed samples. Above 600 degrees C, OTCS signals from As,, antisites and shallower deep levels become weaker, indicating the decrease of these defects. It is argued that the excess arsenic atoms group together to form arsenic clusters during annealing. (C) 2000 Elsevier Science B.V. All rights reserved.

Effect of arsenic precipitates on Fermi level in GaAs grown by molecular-beam epitaxy at low temperature

A simple model is presented to discuss the effect of As precipitates on the Fermi level in GaAs grown by molecular-beam epitaxy at low temperature (LT-GaAs). This model implements the compensation between point defects and the depletion of arsenic precipitates. The condition that the Fermi level is pinned by As precipitates is attained. The shifts of the Fermi level in LT-GaAs with annealing temperature are explained by our model. Additionally, the role of As precipitates in conventional semi-insulating GaAs is discussed. (C) 2000 American Institute of Physics. [S0021-8979(00)09905-9].

Arsenic clusters on the surface of vertically aligned InAs islands on GaAs substrate by annealing

The formation of arsenic clusters in a system of vertically aligned InAs quantum islands on GaAs during thermal annealing under As overpressure has been investigated by transmission electron microscopy (TEM) and Raman scattering. Semicoherent arsenic clusters, identified by TEM examination, have been formed on the surface of the GaAs capping layer. The existence of arsenic precipitates is also confirmed by Raman spectra, showing new peaks from the annealed specimen at 256 and 199 cm(-1). These peaks have been ascribed to A(1g) and E-g Raman active phonons of crystalline arsenic. The phenomenon can be understood by a model of strain-induced selected growth under As overpressure. (C) 1999 American Institute of Physics. [S0003-6951(99)02045-8].

Rapid thermal annealing of arsenic implanted Si1-xGex epilayers

Rapid thermal annealing of arsenic implanted Si1-xGex was studied by secondary ion-mass spectroscopy (SIMS) and spreading resistance probe (SRP) over a wide range of Ge fractions (0-43%). Redistribution of the implanted arsenic was followed as a function of Ge content and annealing temperature. Arsenic concentration profiles from SIMS indicated that the behavior of implanted arsenic in Si1-xGex after RTA was different from that in Si, and the Si1-xGex samples exhibited box-shaped, concentration-dependent diffusion profiles with increasing Ge content. The maximum concentrations of electrically active arsenic in Si1-xGex was found to decrease with increasing Ge content. Experimental results showed that the arsenic diffusion is enhanced with increasing temperature for certain Ge content and strongly dependent on Ge content, and the higher Ge content, the faster As diffusion.

Excess arsenic in GaAs grown at low temperatures by molecular beam epitaxy

The structural properties of GaAs grown at low temperatures by molecular beam epitaxy (LTMBE GaAs) were studied. The excess arsenic atoms in LTMBE GaAs exist in the form of arsenic interstitial couples (i,e, two ns atoms share the one host site), and cause an increase in the lattice parameter of LTMBE GaAs. Annealing at above 300 degrees C, the arsenic interstitial couples decomposed, and As precipitates formed, resulting in a decrease in the lattice parameter.