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1.  Targeted proteomics for validation of biomarkers in clinical samples 
The rapid rise and application of proteomic technologies has resulted in an exponential increase in the number of proteins that have been discovered and presented as ‘potential’ biomarkers for specific diseases. Unfortunately, the number of biomarkers approved for use by the Food and Drug Administration has not risen in likewise manner. While there are a number of reasons for this discrepancy, this glut of ‘potential’ biomarkers also indicates the need for validation methods to confirm or refute their utility in clinical diagnostics. For this reason, the emphasis on developing methods to target and measure the absolute quantity of specific proteins and peptides in complex proteomic samples has grown.
doi:10.1093/bfgp/eln050
PMCID: PMC2722261  PMID: 19109305
mass spectrometry; biomarker validation; targeted proteomics; multiple-reaction monitoring; AQUA; SISCAPA
2.  Comparative glycoproteomics: approaches and applications 
Glycosylation plays fundamental roles in controlling various biological processes. Therefore, glycosylation analysis has become an important target for proteomic research and has great potential for clinical applications. With the continuous development and refinement of glycoprotein isolation methods, increasing attention has been directed to the quantitative and comparative aspects. This review describes the mass spectrometry (MS)-based techniques for the comparative analysis of glycoproteins and their applications to answer a wide range of interesting biological questions.
doi:10.1093/bfgp/eln053
PMCID: PMC2673949  PMID: 19091783
biomarkers; glycoprotein quantitation; glycosylation; isotopic labelling; mass spectrometry; proteomics
3.  Big fish in the genome era 
doi:10.1093/bfgp/eln057
PMCID: PMC2722260  PMID: 19109307
4.  Transposon tools hopping in vertebrates 
In the past decade, tools derived from DNA transposons have made major contributions to vertebrate genetic studies from gene delivery to gene discovery. Multiple, highly complementary systems have been developed, and many more are in the pipeline. Judging which DNA transposon element will work the best in diverse uses from zebrafish genetic manipulation to human gene therapy is currently a complex task. We have summarized the major transposon vector systems active in vertebrates, comparing and contrasting known critical biochemical and in vivo properties, for future tool design and new genetic applications.
doi:10.1093/bfgp/eln049
PMCID: PMC2722259  PMID: 19109308
transposon; gene delivery; gene discovery; gene transfer vectors; vertebrates
5.  Emerging gene knockout technology in zebrafish: zinc-finger nucleases 
One advantage of the zebrafish model system is the ability to use forward genetics to reveal critical gene functions by their mutant phenotype. Reverse genetic tools are available, although it is more challenging and time-consuming to identify mutations in specific genes of interest and virtually impossible to induce mutations in a targeted manner. Two recent papers have shown that locus-specific zinc-finger nucleases (ZFNs) can be used to create mutations in investigator-specified loci at high frequency, generating considerable enthusiasm that the technology may be generally applicable to many zebrafish genes. The rate-limiting step in ZFN application is typically the zinc-finger protein (ZFP) design phase, partly because ZFPs that bind to intended target sequences in naked DNA may not recognize the target within chromatin, or may recognize cryptic sites. Importantly, both papers also provide new tools to validate or pre-select ZFNs that work well in vivo and thus greatly facilitate the identification of active ZFNs. Finally, work in other model systems and in cultured cells show that ZFNs can facilitate homology-directed repair, raising the exciting possibility that ZFNs may facilitate homologous recombination in zebrafish, allowing site-specific modification of endogenous genes via a method that does not require embryonic stem cell technology.
doi:10.1093/bfgp/eln043
PMCID: PMC2722258  PMID: 19109309
gene targeting; targeted mutagenesis; zebrafish; zinc-finger nuclease; non-homologous end joining; double-strand break repair
6.  Chemobehavioural phenomics and behaviour-based psychiatric drug discovery in the zebrafish 
Despite their ubiquity and impact, psychiatric illnesses and other disorders of the central nervous system remain among the most poorly treated diseases. Most psychiatric medicines were discovered due to serendipitous observations of behavioural phenotypes in humans, rodents and other mammals. Extensive behaviour-based chemical screens would likely identify novel psychiatric drugs. However, large-scale chemical screens in mammals are inefficient and impractical. In contrast, zebrafish are very well suited for high-throughput behaviour-based drug discovery. Furthermore, the vast amounts of data generated from large-scale behavioural screens in zebrafish will facilitate a systems-level analysis of how chemicals affect behaviour. Unlike serendipitous discoveries in mammals, a comprehensive and integrative analysis of zebrafish chemobehavioural phenomics may identify functional relationships that would be missed by more reductionist approaches. Thus, behaviour-based chemical screens in the zebrafish may improve our understanding of neurobiology and accelerate the pace of psychiatric drug discovery.
doi:10.1093/bfgp/eln040
PMCID: PMC2722257  PMID: 18784194
phenomics; chemical genetics; zebrafish
7.  The neurogenetic frontier—lessons from misbehaving zebrafish 
One of the central questions in neuroscience is how refined patterns of connectivity in the brain generate and monitor behavior. Genetic mutations can influence neural circuits by disrupting differentiation or maintenance of component neuronal cells or by altering functional patterns of nervous system connectivity. Mutagenesis screens therefore have the potential to reveal not only the molecular underpinnings of brain development and function, but to illuminate the cellular basis of behavior. Practical considerations make the zebrafish an organism of choice for undertaking forward genetic analysis of behavior. The powerful array of experimental tools at the disposal of the zebrafish researcher makes it possible to link molecular function to neuronal properties that underlie behavior. This review focuses on specific challenges to isolating and analyzing behavioral mutants in zebrafish.
doi:10.1093/bfgp/eln039
PMCID: PMC2722256  PMID: 18836206
zebrafish; behavior; mutagenesis
8.  Using retroviruses as a mutagenesis tool to explore the zebrafish genome 
We review different uses of the retroviral mutagenesis technology as the tool to manipulate the zebrafish genome. In addition to serving as a mutagen in a phenotype-driven forward mutagenesis screen as it was originally adapted for, retroviral insertional mutagenesis can also be exploited in reverse genetic approaches, delivering enhancer- and gene-trap vectors for the purpose of examining gene expression patterns and mutagenesis, making sensitized mutants amenable for chemical and genetic modifier screens, and producing gain-of-function mutations by epigenetically overexpressing the retroviral-inserted genes. From a technology point of view, we also summarize the recent advances in the high-throughput cloning of retroviral integration sites, a pivotal step toward identifying mutations. Lastly, we point to some potential directions that retroviral mutagenesis might take from the lessons of studying other model organisms.
doi:10.1093/bfgp/eln038
PMCID: PMC2722255  PMID: 18977782
genetics; moloney murine leukemia virus; enhancer traps; gene traps; linker-mediated PCR; zebrafish
9.  Reverse genetics in zebrafish by TILLING 
TILLING, for Targeting Induced Local Lesions in Genomes, is a reverse genetics strategy that identifies mutations in specific genes of interest in chemically mutagenized populations. First described in 2000 for mutation detection in Arabidopsis, TILLING is now used in a wide range of plants including soybean, rice, barley and maize as well as for animal model systems, including Arabidopsis, Drosophila, Caenorhabditis elegans, rat, medaka and zebrafish and for the discovery of naturally occurring polymorphisms in humans. This review summarizes current TILLING methodologies as they have been applied to the zebrafish, ongoing TILLING projects and resources in the zebrafish community, and the future of zebrafish TILLING.
doi:10.1093/bfgp/eln046
PMCID: PMC2899843  PMID: 19028802
zebrafish; TILLING; Cel1 mismatch cleavage; resequencing; reverse genetics
10.  Quantitative matrix-assisted laser desorption/ionization mass spectrometry 
This review summarizes the essential characteristics of matrix-assisted laser desorption/ionization (MALDI) time-of-flight mass spectrometry (TOF MS), especially as they relate to its applications in quantitative analysis. Approaches to quantification by MALDI-TOF MS are presented and published applications are critically reviewed.
doi:10.1093/bfgp/eln041
PMCID: PMC2722264  PMID: 19106161
quantification; quantitative analysis; MALDI; mass spectrometry; biomarkers

Results 1-10 (10)