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| 2. |
- Andrade, Jorge, et al.
(author)
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Using Grid Technology for Computationally Intensive Applied Bioinformatics Analyses
- 2006
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In: In Silico Biology. - 1386-6338. ; 6:6, s. 495-504
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Journal article (peer-reviewed)abstract
- For several applications and algorithms used in applied bioinformatics, a bottle neck in terms of computational time may arise when scaled up to facilitate analyses of large datasets and databases. Re-codification, algorithm modification or sacrifices in sensitivity and accuracy may be necessary to accommodate for limited computational capacity of single work stations. Grid computing offers an alternative model for solving massive computational problems by parallel execution of existing algorithms and software implementations. We present the implementation of a Grid-aware model for solving computationally intensive bioinformatic analyses exemplified by a blastp sliding window algorithm for whole proteome sequence similarity analysis, and evaluate the performance in comparison with a local cluster and a single workstation. Our strategy involves temporary installations of the BLAST executable and databases on remote nodes at submission, accommodating for dynamic Grid environments as it avoids the need of predefined runtime environments (preinstalled software and databases at specific Grid-nodes). Importantly, the implementation is generic where the BLAST executable can be replaced by other software tools to facilitate analyses suitable for parallelisation. This model should be of general interest in applied bioinformatics. Scripts and procedures are freely available from the authors.
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| 3. |
- Berglund, Lisa, et al.
(author)
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A genecentric Human Protein Atlas for expression profiles based on antibodies
- 2008
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In: Molecular & Cellular Proteomics. - 1535-9476 .- 1535-9484. ; 7:10, s. 2019-2027
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Research review (peer-reviewed)abstract
- An attractive path forward in proteomics is to experimentally annotate the human protein complement of the genome in a genecentric manner. Using antibodies, it might be possible to design protein-specific probes for a representative protein from every protein-coding gene and to subsequently use the antibodies for systematical analysis of cellular distribution and subcellular localization of proteins in normal and disease tissues. A new version (4.0) of the Human Protein Atlas has been developed in a genecentric manner with the inclusion of all human genes and splice variants predicted from genome efforts together with a visualization of each protein with characteristics such as predicted membrane regions, signal peptide, and protein domains and new plots showing the uniqueness (sequence similarity) of every fraction of each protein toward all other human proteins. The new version is based on tissue profiles generated from 6120 antibodies with more than five million immunohistochemistry-based images covering 5067 human genes, corresponding to approximately 25% of the human genome. Version 4.0 includes a putative list of members in various protein classes, both functional classes, such as kinases, transcription factors, G-protein-coupled receptors, etc., and project-related classes, such as candidate genes for cancer or cardiovascular diseases. The exact antigen sequence for the internally generated antibodies has also been released together with a visualization of the application-specific validation performed for each antibody, including a protein array assay, Western blot analysis, immunohistochemistry, and, for a large fraction, immunofluorescence-based confocal microscopy. New search functionalities have been added to allow complex queries regarding protein expression profiles, protein classes, and chromosome location. The new version of the protein atlas thus is a resource for many areas of biomedical research, including protein science and biomarker discovery.
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| 4. |
- Berglund, Lisa, et al.
(author)
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A whole-genome bioinformatics approach to selection of antigens for systematic antibody generation
- 2008
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In: Proteomics. - : Wiley. - 1615-9853 .- 1615-9861. ; 8:14, s. 2832-2839
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Journal article (peer-reviewed)abstract
- Here, we present an antigen selection strategy based on a whole-genome bioinformatics approach, which is facilitated by an interactive visualization tool displaying protein features from both public resources and in-house generated data. The web-based bioinformatics platform has been designed for selection of multiple, non-overlapping recombinant protein epitope signature tags by display of predicted information relevant for antigens, including domain- and epitope sized sequence similarities to other proteins, transmembrane regions and signal peptides. The visualization tool also displays shared and exclusive protein regions for genes with multiple splice variants. A genome-wide analysis demonstrates that antigens for approximately 80% of the human protein-coding genes can be selected with this strategy.
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| 6. |
- Berglund, Lisa, et al.
(author)
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Generation of validated antibodies towards the human proteome
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Journal article (other academic/artistic)abstract
- Here we show the results from a large effort to generate antibodies towards the human proteome. A high-throughput strategy was developed based on cloning and expression of antigens as recombitant protein epitope signature tags (PrESTs) Affinity purified polyclonal antibodies were generated, followed by validation by protein microarrays, Western blotting and microarray-based immunohistochemistry. PrESTs were selected based on sequence uniqueness relative the proteome and a bioinformatics analysis showed that unique antigens can be found for at least 85% of the proteome using this general strategy. The success rate from antigen selection to validated antibodies was 31%, and from protein to antibody 55%. Interestingly, membrane-bound and soluble proteins performed equally and PrEST lengths between 75 and 125 amino acids were found to give the highest yield of validated antibodies. Multiple antigens were selected for many genes and the results suggest that specific antibodies can be systematically generated to most human proteibs.
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| 8. |
- Berglund, Lisa, 1978-
(author)
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Selection of antigens for antibody-based proteomics
- 2008
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Doctoral thesis (other academic/artistic)abstract
- The human genome is predicted to contain ~20,500 protein-coding genes. The encoded proteins are the key players in the body, but the functions and localizations of most proteins are still unknown. Antibody-based proteomics has great potential for exploration of the protein complement of the human genome, but there are antibodies only to a very limited set of proteins. The Human Proteome Resource (HPR) project was launched in August 2003, with the aim to generate high-quality specific antibodies towards the human proteome, and to use these antibodies for large-scale protein profiling in human tissues and cells. The goal of the work presented in this thesis was to evaluate if antigens can be selected, in a high-throughput manner, to enable generation of specific antibodies towards one protein from every human gene. A computationally intensive analysis of potential epitopes in the human proteome was performed and showed that it should be possible to find unique epitopes for most human proteins. The result from this analysis was implemented in a new web-based visualization tool for antigen selection. Predicted protein features important for antigen selection, such as transmembrane regions and signal peptides, are also displayed in the tool. The antigens used in HPR are named protein epitope signature tags (PrESTs). A genome-wide analysis combining different protein features revealed that it should be possible to select unique, 50 amino acids long PrESTs for ~80% of the human protein-coding genes. The PrESTs are transferred from the computer to the laboratory by design of PrEST-specific PCR primers. A study of the success rate in PCR cloning of the selected fragments demonstrated the importance of controlled GC-content in the primers for specific amplification. The PrEST protein is produced in bacteria and used for immunization and subsequent affinity purification of the resulting sera to generate mono-specific antibodies. The antibodies are tested for specificity and approved antibodies are used for tissue profiling in normal and cancer tissues. A large-scale analysis of the success rates for different PrESTs in the experimental pipeline of the HPR project showed that the total success rate from PrEST selection to an approved antibody is 31%, and that this rate is dependent on PrEST length. A second PrEST on a target protein is somewhat less likely to succeed in the HPR pipeline if the first PrEST is unsuccessful, but the analysis shows that it is valuable to select several PrESTs for each protein, to enable generation of at least two antibodies, which can be used to validate each other.
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| 9. |
- Berglund, Lisa, et al.
(author)
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The epitope space of the human proteome
- 2008
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In: Protein Science. - : Wiley. - 0961-8368 .- 1469-896X. ; 17:4, s. 606-613
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Journal article (peer-reviewed)abstract
- In the post-genome era, there is a great need for protein-specific affinity reagents to explore the human proteome. Antibodies are suitable as reagents, but generation of antibodies with low cross-reactivity to other human proteins requires careful selection of antigens. Here we show the results from a proteomewide effort to map linear epitopes based on uniqueness relative to the entire human proteome. The analysis was based on a sliding window sequence similarity search using short windows (8, 10, and 12 amino acid residues). A comparison of exact string matching (Hamming distance) and a heuristic method (BLAST) was performed, showing that the heuristic method combined with a grid strategy allows for whole proteome analysis with high accuracy and feasible run times. The analysis shows that it is possible to find unique antigens for a majority of the human proteins, with relatively strict rules involving low sequence identity of the possible linear epitopes. The implications for human antibody-based proteomics efforts are discussed.
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| 10. |
- Berglund, Martin, 1985-, et al.
(author)
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Evaluation of a microplasma source based on a stripline split-ring resonator
- 2013
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In: Plasma sources science & technology. - : Institute of Physics (IOP). - 0963-0252 .- 1361-6595. ; 22:5, s. 055017-
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Journal article (peer-reviewed)abstract
- In this paper, a stripline split-ring resonator microwave-induced plasma source, aimed for integration in complex systems, is presented and compared with a traditional microstrip design. Devices based on the two designs are evaluated using a plasma breakdown test setup for measuring the power required to ignite plasmas at different pressures. Moreover, the radiation efficiency of the devices is investigated with a Wheeler cap, and their electromagnetic compatibility is investigated in a variable electrical environment emulating an application. Finally, the basic properties of the plasma in the two designs are investigated in terms of electron temperature, plasma potential and ion density. The study shows that, with a minor increase in plasma ignition power, the stripline design provides a more isolated and easy-to-integrate alternative to the conventional microstrip design. Moreover, the stripline devices showed a decreased antenna efficiency as compared with their microstrip counterparts, which is beneficial for plasma sources. Furthermore, the investigated stripline devices exhibited virtually no frequency shift in a varying electromagnetic environment, whereas the resonance frequency of their microstrip counterparts shifted up to 17.5%. With regard to the plasma parameters, the different designs showed only minor differences in electron temperature, whereas the ion density was higher with the stripline design.
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