Real-time PCR with molecular beacons provides a highly accurate assay for detection of Tay-Sachs alleles in single cells

The results presented here provide the first single-cell genetic assay for Tay-Sachs disease based on real-time PCR. Individual lymphoblasts were lysed with an optimized lysis buffer and assayed using one pair of primers that amplifies both the wild type and 1278 + TATC Tay-Sachs alleles. The resulting amplicons were detected in real time with two molecular beacons each with a different colored fluorochrome. The kinetics of amplicon accumulation generate objective criteria by which to evaluate the validity of each reaction. The assay had an overall utility of 95%, based on the detection of at least one signal in 235 of the 248 attempted tests and an efficiency of 97%, as 7 of the 235 samples were excluded from further analysis for objective quantitative reasons. The accuracy of the assay was 99.1%, because 228 of 230 samples gave signals consistent with the genotype of the cells. Only two of the 135 heterozygous samples were allele drop-outs, a rate far lower than previously reported for single-cell Tay-Sachs assays using conventional methods of PCR. Copyright © 2002 John Wiley & Sons, Ltd.     

Molecular diagnosis of a novel heterozygous 268C→T (R90C) mutation in TGIF gene in a fetus with holoprosencephaly and premaxillary agenesis

We report the sonographic diagnosis and molecular analysis of holoprosencephaly (HPE) and premaxillary agenesis in a second-trimester fetus with a 46,XY karyotype. Mutational sequence analyses for the entire coding region and exon–intron boundaries of SHH, ZIC2, SIX3 and TGIF genes identified a novel heterozygous missense TGIF mutation 268C→T (CGC→TGC change) that predicts an Arg90Cys substitution in the homeodomain region of TGIF. The proband's parents did not carry the mutation. The present case is an example of the heterogeneous entity of the HPE spectrum and demonstrates that adjunctive molecular analyses of distinct human genes for HPE can reassure genetic counselling by elucidating the genetic pathogenesis, especially in cytogenetically normal fetuses affected with HPE. Copyright © 2002 John Wiley & Sons, Ltd.     

能源植物修复镉污染土壤对根际细菌网络结构的影响

选取油脂类能源植物大豆和碳水化合物类能源植物玉米,采用高通量测序方法研究大豆、玉米修复Cd污染土壤过程中根际土壤细菌群落组成,基于高通量测序数据采用分子生态网络分析细菌相互作用.结果表明,50 mg·kg-1Cd污染土壤中两种植物根部Cd浓度和积累量最高,转移系数TF分别为玉米0.78和大豆0.35.基于细菌16S r RNA基因的群落分析表明,大豆、玉米根际土壤细菌主要包括Proteobacteria(变形菌门)、Acidobacteria(酸杆菌门)31个门细菌组成,大豆、玉米种植均能影响土壤细菌群落组成,能影响Candidate division TM7 norank、Acidimicrobiales norank、Sphingomonas等丰度.分子生态网络分析表明种植大豆和玉米增加了细菌之间的相互作用,导致其网络结构更为复杂,关键细菌从不种植物处理的1个增加到种植大豆的6个和种植玉米的10个.     

河谷型城市土壤有害金属有机酸与细菌淋溶特性

在野外调查与前期研究的基础上,根据一点多样混合采样法收集宝鸡河谷型城市污染土壤样品50个.通过等温振荡平衡法研究了人工合成的螯合剂EDTA、DTPA、低分子有机酸草酸(Oxalic acid)、丁二酸(Butanedioic acid)、柠檬酸(Citrate)和枯草芽孢杆菌对城市污染土壤有害金属As、Cr、Cu、Pb、Sb、V、Zn的淋溶解吸特性.研究结果表明,EDTA、DTPA对宝鸡城市污染土壤As、Cr、Cu、Pb、Sb、V、Zn的整体淋溶解吸效果要优于草酸、丁二酸和柠檬酸.有机酸对宝鸡城市土壤有害金属As、Cr、Cu、Pb、Sb、V、Zn淋洗速率顺序为:柠檬酸:PbSbZn≈AsCuVCr;DTPA:Pb≥Cu≈ZnSbAsVCr;EDTA:Pb≥ZnCuSbAsVCr;草酸:PbSbAsVCuCr;丁二酸:CrPbSbAsVCu(丁二酸淋洗的Cr随时间缓慢增加).枯草芽孢杆菌对土壤有害金属淋溶具有pH和温度的依赖性,特别对Pb、Sb的淋溶解吸特为敏感,较高pH(pH=6)淋溶要大于低pH(pH=2).研究表明,枯草芽孢杆菌淋溶土壤有害金属是细菌本身、有机酸和淋溶液pH共同作用的过程.重金属Pb和Sb是宝鸡市环境污染与人群健康风险最主要的因子,因此,有机酸和细菌及其淋溶pH综合作用对西部河谷型城市土壤有害金属Pb、Sb复合污染的原位修复提供了新的思路.     

结合光谱法和计算模拟多角度分析2,2',4,4',5-五溴二苯醚与人血清白蛋白的作用机制

本文结合光谱法、动力学模拟(MD)和分子对接等手段,多角度地研究了2,2′,4,4′,5-五溴二苯醚(BDE-99)与人血清白蛋白(HSA)在pH=7.4模拟生理环境下的相互作用机制.首先采用MD从理论上模拟BDE-99与HSA相互作用的构象变化情况;然后利用同步荧光和三维荧光光谱法从实验角度进行验证,所得结果表明,BDE-99使HSA的疏水性增强从而导致其构象发生变化.同时,荧光光谱和紫外光谱得出BDE-99对HSA的猝灭机制属于静态猝灭和非辐射能量转移.另外,分子对接结果显示,BDE-99通过静电引力和疏水作用力结合在HSA的位点I处,这与竞争实验和热力学参数的分析结果是一致的.实验数据与模拟计算结果的相互印证,为进一步探究BDE-99和HSA的相互作用机制提供了重要的信息和参考依据.     

分子流态下翅片管式热沉传输几率计算分析

目的研究空间环境模拟器翅片管式热沉传输特性,为进一步开展真空等效性分析等研究工作奠定基础。方法根据热沉结构特点建立镜面反射气体输运通道模型,采用蒙特卡罗方法,对该模型的传输几率进行分析计算,并建立验证模型,对计算结果进行验证。结果对于所选择的翅片管式热沉,其平面阵列的传输几率为0.030 116 4,柱面阵列的传输几率为0.017 988 6。结论蒙特卡罗方法可以用于计算一定结构的翅片管式热沉的传输几率。通过传输几率的计算,可以将热沉等效为具有相应传输几率的可透射平面,从而在保证模拟精度的同时大大简化模拟计算强度,提高计算效率。     

两种不同硅铝比的ZSM-5型分子筛去除水中Pb~(2+)的研究

考察硅铝比不同的ZSM-5两种分子筛(25H和50H,25和50为硅铝比:SiO2/Al2O3)对水中Pb2+离子的吸附去除效能。两种材料均可有效吸附去除水中Pb2+离子,吸附等温线符合Langmuir吸附等温式,25H的最大吸附容量达22.22mg/g,吸附动力学符合假二阶动力学模型,吸附速率及吸附容量顺序为:25H>50H。随着投加量的增加,分子筛对Pb2+的去除率呈上升趋势。常见共存阳离子均对分子筛吸附去除Pb2+产生影响。     

Molecular toxicity of earthworms induced by cadmium contaminated soil and biomarkers screening

Earthworms (Eisenia fetida) were used to study the impact of low-dose cadmium in treated artificial soil (0, 0.6, 3, 6, 15, 30 mg/kg) and contaminated natural soil (1.46 mg/kg). The changes of earthworms' physiological related gene expressions of metallothionein (MT), annetocin, calreticulin and antimicrobial peptides were detected using real-time PCR after a 70-day incubation period. The results showed that low doses of cadmium could up regulate earthworms' MT and down regulate annetocin gene expression and show a significant positive and negative correlation respectively. The expression of two other genes, calreticulin and anti-microbial peptides, was induced at low doses of cadmium (highest gene expression at 0.6 mg/kg for calreticulin and 6 mg/kg for anti-microbial peptides) and inhibited at high doses. No significant correlation was found for these two genes. This study shows that MT and annetocin genes expression found in earthworms in contaminated soil have the potential to be developed as biomarkers of soil cadmium pollution.