Accurate method for determining active genes
The total DNA of an organism is significantly more extensive than the actual genome used. A consortium of German and U.S. researchers involving the Max Planck Institute for Plant Breeding Research in Cologne (MPIPZ) and the Heinrich Heine University Düsseldorf (HHU) developed a method in order to determine all regions of the active genome in a single analysis. They present their results using the crop plant maize in the current issue of the journal PLoS Genetics.
Only a few percent of the genome actually serve to encode and control the structure of an organism and its functions. The vast majority of the rest have no apparent function. Moreover, depending on the location and function of a cell, the genes that are specifically activated to enable the cell's particular function differ in turn.
To read out the genes, the enzyme "RNA polymerase" attaches itself to the DNA. Starting at a specified point, it reads a defined section of the hereditary molecule by transferring the information from the DNA to a similar molecule, the "messenger RNA" (mRNA). The mRNA is then in turn transported to ribosomes, which produce proteins from the mRNA blueprint. These proteins can serve as building blocks for the cell or control certain functions within it.
Transcription factors play a crucial role in this targeted reading of genes. These proteins bind to the DNA and provide the start signal for the polymerase. Since these transcription factors often couple close ("cis") to the site to be read out, the binding sites are also called "cis elements"; the set of all these sites is the "cistrome".
Until now, it has been extremely challenging to determine the cistrome of an organism: Each transcription factor had to be examined separately using the so-called ChIP method. For a corn plant with around 2,500 transcription factors, this means the same number of experiments to analyze the entire cistrome. A team of researchers from the Florida State University in Tallahassee, USA, the Institute of Molecular Physiology at HHU and the MPIPZ in Cologne, have now developed a method called "MOA-seq" that can be used to determine the entire cistrome in a single experiment and at high resolution.
In this study, published in PlosGenetics, the research team presents how they applied the new method to the cistrome of an evolved maize cob. For the first time, they were able, with accuracy comparable to the ChIP method, to determine a cistrome of this so important crop, and to do so both faster and with less cost and material than it would have been possible with the established method.
Dr. Thomas Hartwig, head of the ‘crop yield in maize’ research team based at the Max Planck Institute in Cologne, says: "MOA-seq reliably finds the binding sites of active transcription factors in the genome that switch genes on or off. The method has very diverse applications, for example, in assigning mutations that alter cis-elements and thus controlled gene expression and traits."
Prof. Wolf B. Frommer, Ph.D., head of HHU's Institute of Molecular Physiology, added: "I congratulate Dr. Hartwig and Prof. Hank Bass of Florida State University. With a base-accurate assignment of variations in promoters to specific traits, such as yield or drought resistance, breeding of climate-adapted varieties can be made much more efficient."
The MOA sequencing method ("MOA-seq") developed in Cologne, Düsseldorf and Tallahassee is also the basis of a large-scale project by Dr. Hartwig's group to characterize the cistrome involved in drought resistance in maize and other crops. For this purpose, an international consortium named "FIND-CIS" was founded, which has also recently been supported by the German Research Foundation (DFG).