Accurate and superaccurate gene mapping.

Highly accurate gene mapping techniques need to be developed to clone disease genes with unknown defective products. The classical pedigree method and methods based on cytologically observable chromosome aberrations share definite limits in resolution. We quantify the limits in resolution for the pedigree method. We also discuss a technique for gene localization that exploits the possible presence of minute depletions overlapping the disease locus. One can search for such submicroscopic deletions by aiming random probes at them. We show quantitatively that relatively few probes may suffice to hit a target deletion. Choosing which probes to aim should be guided by pedigree studies and by close examination of relevant cytologically observable translocations and deletions.     

Gene expression profiling of human diseases by serial analysis of gene expression

Until recently, the approach to understanding the molecular basis of complex syndromes such as cancer, coronary artery disease, and diabetes was to study the behavior of individual genes. However, it is generally recognized that expression of a number of genes is coordinated both spatially and temporally and that this coordination changes during the development and progression of diseases. Newly developed functional genomic approaches, such as serial analysis of gene expression (SAGE) and DNA microarrays have enabled researchers to determine the expression pattern of thousands of genes simultaneously. One attractive feature of SAGE compared to microarrays is its ability to quantify gene expression without prior sequence information or information about genes that are thought to be expressed. SAGE has been successfully applied to the gene expression profiling of a number of human diseases. In this review, we will first discuss SAGE technique and contrast it to microarray. We will then highlight new biological insights that have emerged from its application to the study of human diseases.     

Multiple interval mapping for gene expression QTL analysis

To find the correlations between genome-wide gene expression variations and sequence polymorphisms in inbred cross populations, we developed a statistical method to claim expression quantitative trait loci (eQTL) in a genome. The method is based on multiple interval mapping (MIM), a model selection procedure, and uses false discovery rate (FDR) to measure the statistical significance of the large number of eQTL. We compared our method with a similar procedure proposed by Storey et al. and found that our method can be more powerful. We identified the features in the two methods that resulted in different statistical powers for eQTL detection, and confirmed them by simulation. We organized our computational procedure in an R package which can estimate FDR for positive findings from similar model selection procedures. The R package, MIM-eQTL, can be found at .     

基因表达系列分析技术在动物科学中的研究进展

基因表达系列分析（serialanalysisofgeneexpression,SAGE）是一种快速分析特定组织或细胞内基因表达信息的技术,不但可以比较不同组织细胞在不同时间、空间条件下基因表达的差异,还能发现新基因。近几年来,SAGE技术在动物基因表达研究中的应用取得了飞速发展。就SAGE技术的原理、实验路线、优缺点和改进以及SAGE在动物科学研究中的研究现状及应用前景作一简要介绍。     

The genetics of gene expression and gene mapping

A recent publication has shown that a significant portion of gene expression levels are under genetic control in different organisms, that there are hotspot regions in the genome that control the expression of many other genes, and how gene expression data can be used to localize genes that affect clinical traits.     

Correct identification of genes from serial analysis of gene expression tag sequences

SAGE (serial analysis of gene expression) is a remarkable technique for genome-wide analysis of gene expression. It is crucial to understand the extent to which SAGE can accurately indicate a gene or expressed sequence tag (EST) with a single tag. We analyzed the effect of the size of SAGE tag on gene identification. Our observation indicates that SAGE tags are in general not long enough to achieve the degree of uniqueness of identification originally envisaged. Our observations also indicate that the limitation of using SAGE tag to identify a gene can be overcome by converting SAGE tags into longer 3' EST sequences with the generation of longer cDNA fragments from SAGE tages for gene identification (GLGI) method.     

Accurate and high resolution in situ hybridization analysis of gene expression in secondary stem tissues

Accurate in situ hybridization analysis in secondary stem tissues of plants has been hindered by specific characteristics of these tissues. First, secondary cell walls non-specifically bind probes used for in situ hybridization thus preventing gene expression analysis in the lignified regions of the stem, such as the xylem. Second, the mRNA in the cambial meristem and its recent derivatives are prone to inadequate fixation when conventional techniques are used. Here we describe an in situ hybridization technique which uses fast freezing and freeze substitution to cryoimmobilize the mRNA followed by embedding in a methacrylate resin for high-resolution analysis of gene expression. By using a transgenic poplar line harbouring rolC:uidA, rolC:iaaM, the gene expression pattern could be compared with histochemical GUS staining. This in situ hybridization technique results in superior preservation of cellular contents, retention of mRNA in all cell types in the poplar stem, a significant reduction of non-specific binding to secondary cell walls and a resolution not previously possible in secondary tissues. This technique will be particularly valuable for the expression analysis of genes involved in xylogenesis and wood formation.     

Gene expression profiling of resting and activated vascular smooth muscle cells by serial analysis of gene expression and clustering analysis

Migration and proliferation of vascular smooth muscle cells (SMCs) are key events in atherosclerosis. However, little is known about alterations in gene expression upon transition of the quiescent, contractile SMC to the proliferative SMC. We performed serial analysis of gene expression (SAGE) of cultured, human SMCs, either grown under resting circumstances or activated with an atherogenic stimulus. Analysis of tags, representing 47,209 and 47,259 mRNAs from a library of resting and activated SMCs, respectively, identified 105 tags induced and 52 tags repressed greater than fivefold. To evaluate the relevance in SMC biology of unmatched, regulated tags, we performed hierarchical clustering analysis, based on their expression profiles in public SAGE databases, and clustered these novel genes in distinct groups. The regulation in SMCs was confirmed by Northern blotting for representative genes of these groups. Plasminogen activator inhibitor-2 has not been associated with atherosclerosis before and was localized to atherosclerotic lesions.