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A guide for in-house design of template-switch-based 5' rapid amplification of cDNA ends systems
Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Photochemistry and Molecular Science.
Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Photochemistry and Molecular Science.
2009 (English)In: Analytical biochemistry, ISSN 1096-0309Article in journal (Refereed) Published
Abstract [en]

Rapid amplification of cDNA ends (RACE) is an established strategy used to determine the transcription start point(s) and the 5' untranslated region(s) of mRNA. Different approaches to perform 5' RACE are available, and one particularly simple and powerful strategy is based on a phenomenon called template-switching. We investigated different aspects of template-switch-based 5' RACE, and we describe the different steps leading to the in-house development of a complete 5' RACE system-from oligonucleotide design to polymerase chain reaction (PCR) amplification. We show that the resulting system is reliable, time-efficient, and inexpensive.

Place, publisher, year, edition, pages
National Category
Natural Sciences
URN: urn:nbn:se:uu:diva-110392DOI: 10.1016/j.ab.2009.10.022ISI: 000274163900015PubMedID: 19837043OAI: oai:DiVA.org:uu-110392DiVA: diva2:276992
Available from: 2009-11-13 Created: 2009-11-13 Last updated: 2012-08-22
In thesis
1. Development of Molecular Biology and Bioinformatics Tools: From Hydrogen Evolution to Cell Division in Cyanobacteria
Open this publication in new window or tab >>Development of Molecular Biology and Bioinformatics Tools: From Hydrogen Evolution to Cell Division in Cyanobacteria
2009 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The use of fossil fuels presents a particularly interesting challenge - our society strongly depends on coal and oil, but we are aware that their use is damaging the environment. Currently, this awareness is gaining momentum, and pressure to evolve towards an energetically cleaner planet is very strong. Molecular hydrogen (H2) is an environmentally suitable energy carrier that could initially supplement or even substitute fossil fuels.

Ideally, the primary energy source to produce hydrogen gas should be renewable, and the process of conversion back to energy without polluting emissions, making this cycle environmentally clean. Photoconversion of water to hydrogen can be achieved using the following strategies: 1) the use of photochemical fuel cells, 2) by applying photovoltaics, or 3) by promoting production of hydrogen by photosynthetic microorganisms, either phototrophic anoxygenic bacteria and cyanobacteria or eukaryotic green algae. For photobiological H2 production cyanobacteria are among the ideal candidates since they: a) are capable of H2 evolution, and b) have simple nutritional requirements - they can grow in air (N2 and CO2), water and mineral salts, with light as the only energy source.

As this project started, a vision and a set of overall goals were established. These postulated that improved H2 production over a long period demanded: 1) selection of strains taking in consideration their specific hydrogen metabolism, 2) genetic modification in order to improve the H2 evolution, and 3) cultivation conditions in bioreactors should be exmined and improved. Within these goals, three main research objectives were set: 1) update and document the use of cyanobacteria for hydrogen production, 2) create tools to improve molecular biology work at the transcription analysis level, and 3) study cell division in cyanobacteria.

This work resulted in: 1) the publication of a review on hydrogen evolution by cyanobacteria, 2) the development of tools to assist understanding of transcription, and 3) the start of a new fundamental research approach to ultimately improve the yield of H2 evolution by cyanobacteria.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2009. 66 p.
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology, ISSN 1651-6214 ; 697
Cyanobacteria, hydrogen, RNA, bioinformatics, RT-PCR, RACE, transcription
National Category
Biochemistry and Molecular Biology Bioinformatics and Systems Biology
Research subject
Molecular Biology
urn:nbn:se:uu:diva-110842 (URN)978-91-554-7678-6 (ISBN)
Public defence
2010-01-20, Häggsalen, Ångströmlaboratoriet, Lägerhyddsv. 1, Uppsala, 13:15 (English)
Available from: 2009-12-21 Created: 2009-11-26 Last updated: 2010-12-22Bibliographically approved

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