BRCA1 mutation screening using restriction endonuclease fingerprinting-single-strand conformation polymorphism in an automated capillary electrophoresis system

Efficient mutation scanning techniques are needed for the rapid detection of novel disease-associated mutations and rare-sequence variants of putative importance. The large size of the breast cancer 1 gene (BRCA1) and the many mutations found throughout its entire coding sequence make screening for mutations in this gene particularly challenging. We have developed a method for screening exon 11 of the BRCA1 gene based on restriction enzyme digestion of fluorescence-labeled polymerase chain reaction (PCR) products followed by single-strand conformation polymorphism (SSCP) using an automated capillary electrophoresis system, denoted capillary restriction endonuclease fingerprinting (REF)-SSCP electrophoresis. Using this strategy on a control set of samples, we were able to detect 17 of 18 known sequence alterations. The method was then applied to screen 73 Norwegian females with family histories of breast and/or ovarian cancer. A total of 172 sequence alterations were detected, including substitutions, insertions, and deletions. One novel substitution of unknown function was identified. Sequencing of all samples negative in the capillary REF-SSCP system gave no additional mutations confirming the high sensitivity of the described methodology. Capillary REF-SSCP electrophoresis appeared as a technically convenient technique, requiring amplification of fewer PCR fragments than traditional SSCP. The novel strategy allows high-throughput mutation scanning without radioactive labeling and polyacrylamide gel electrophoresis (PAGE).     

极化效应指数与脂肪族胺、醇和醚气相碱性的关系

脂肪族胺、醇和醚的气相质子亲合能(PA)与N, O原子所带电荷(qx)以及烷基的极化效应指数(PEI)的关系可表示为其中a、b、c为系数。回归分析结果表明, 上式较好地表达了脂肪族胺、醇和醚的气相碱性变化规律。     

Microfiltration of protein solutions at thin film composite membranes

An experimental study of the interaction of the enzyme yeast alcohol dehydrogenase (YADH) with polysulfone thin film composite microfiltration membranes (Dow-Danmark) has been carried out. It was found that the membranes adsorbed only 3/4 of a monolayer of the enzyme under the conditions studied. Even so, under filtration conditions, the membrane permeation rate decreased continuously with time. This decrease in permeation rate was due neither to concentration polarisation nor to protein adsorption alone. However, it could be quantified using the standard blocking filtration law, which describes a decrease in pore volume due to deposition of protein in the interior structure of the membrane. Reversal of the membrane, so that the supporting matrix faced the feed solution, gave more stable permeation rates. Implications for the microfiltration of industrial fermentation broths are discussed.     

Dicyano-Functionalized MCM-41-Supported Palladium Complex as An Efficient Catalyst for Allylation of Aldehydes and Ketones

Dicyano-functionalized MCM-41-supported palladium complex was prepared from dicyano-functionalized MCM-41 and palladium chloride. This complex exhibited high catalytic activity in the allylation of aldehydes and ketones with allylic chlorides in the presence of SnCl2. This polymeric palladium complex can be recovered and reused without noticeable loss of activity.     

Markov processes in single molecule fluorescence

This article examines the current status of Markov processes in single molecule fluorescence. For molecular dynamics to be described by a Markov process, the Markov process must include all states involved in the dynamics and the first-passage time (FPT) distributions out of those states must be describable by a simple exponential law. The observation of non-exponential FPT distributions or other evidence of non-Markovian dynamics is common in single molecule studies and offers an opportunity to expand the Markov model to include new dynamics or states that improve understanding of the system.     

Kinetic resolution of vic-amino alcohols catalyzed by a chiral Cu(II) complex

Kinetic resolution of N-benzoylated vic-amino alcohols was achieved by benzoylation in the presence of copper triflate and (R,R)-Ph-BOX as catalysts. The observed enantioselectivity was moderate to high. The method was applied to a kinetic resolution of racemic prolinol and piperidinemethanol derivatives as well as an asymmetric desymmetrization of 2-amino-1,3-diol derivatives.     

Stereoselective synthesis of the ABCD ring framework of azaspiracids

A stereoselective synthesis of the ABCD ring framework of azaspiracid-1 and azaspiracid-3 has been achieved using a tandem bis-spiroketalization protocol in the presence of a mild proton source from 1,4-diketone precursor. A tetrahydrofuran intermediate with the correct stereochemistry for the D ring of azaspiracids-1 and 3 was then taken through a linear sequence of reactions to afford the desired diketone precursor. The D-ring of azaspiracid-1 was then constructed by employing a Sharpless asymmetric dihydroxylation followed by etherification using a homoallyl derivative. The structure of the ABCD ring framework with four contiguous rings was established by extensive NMR analysis.     

以交联聚乙烯醇为载体的离子交换剂对蛋白质的分离性能

本文对阴离子交换剂ＤＥＡ—ＰＶＴ常压液相离子交换色谱分离蛋白质的性能、分离条件进行了探讨。结果表明其对蛋白质的分离性能良好，容易洗脱。与载体交联聚乙烯醇相比，ＤＥＡ—ＰＶＴ对蛋白质的非特异性吸附明显降低。     

LaNi4.7Al0.3单颗粒微电极电化学行为的研究

报道了一种单颗粒微电极制备新方法, 并研究了LaNi4.7Al0.3球形单颗粒微电极的电化学行为.     

Single cell manipulation, analytics, and label-free protein detection in microfluidic devices for systems nanobiology

Single cell analytics for proteomic analysis is considered a key method in the framework of systems nanobiology which allows a novel proteomics without being subjected to ensemble-averaging, cell-cycle, or cell-population effects. We are currently developing a single cell analytical method for protein fingerprinting combining a structured microfluidic device with latest optical laser technology for single cell manipulation (trapping and steering), free-solution electrophoretical protein separation, and (label-free) protein detection. In this paper we report on first results of this novel analytical device focusing on three main issues. First, single biological cells were trapped, injected, steered, and deposited by means of optical tweezers in a poly(dimethylsiloxane) microfluidic device and consecutively lysed with SDS at a predefined position. Second, separation and detection of fluorescent dyes, amino acids, and proteins were achieved with LIF detection in the visible (VIS) (488 nm) as well as in the deep UV (266 nm) spectral range for label-free, native protein detection. Minute concentrations of 100 fM injected fluorescein could be detected in the VIS and a first protein separation and label-free detection could be achieved in the UV spectral range. Third, first analytical experiments with single Sf9 insect cells (Spodoptera frugiperda) in a tailored microfluidic device exhibiting distinct electropherograms of a green fluorescent protein-construct proved the validity of the concept. Thus, the presented microfluidic concept allows novel and fascinating single cell experiments for systems nanobiology in the future.