Researchers have created a robot that can analyze the effects of mutations that occur in portions of the genome that help control animal development. While lots of genetic research has focused on parts of the genome that code for protein, that's only a small part of it, and non-coding, regulatory regions can play an important part in controlling how protein-coding genes are expressed. With the robot, the researchers were able to assess the impact of hundreds of mutations. The work has been reported in Nature.
"The real star is this robot," said David Stern, a group leader at HHMI's Janelia Research Campus. "It was extremely creative engineering."
In order to determine the role of hundreds of genetic mutations, the researchers had to prepare thousands of fruit fly embryos in a standardized way; slight modifications had the potential to change the experimental results. The project leader, former Stern lab postdoctoral fellow Justin Crocker, now a group leader at the European Molecular Biology Laboratory (EMBL) in Heidelberg, Germany collaborated with the Janelia Experimental Technology team, jET, to develop a sample-preparing robot.
"The timing of many of these steps is critical," Crocker said. "For us, the goal was to be able to remove the human as much as we could from the equation, to make things very standardized."
The robot was created after jET engineers observed Crocker preparing the samples, and learned to do it themselves. After two failed prototypes and a few years of work, they came up with a machine the size of a toaster called the Hybridizer, which preserves and stains fly embryos by the hundreds at various stages of development, and does so in a consistent way.
The genetic sequence under study is an enhancer region, and the team engineered many variants of it. Over 800 different strains of flies were created, with every one carrying a handful of different mutations in the enhancer. With the robot, the team was now able to look at the impact of hundreds of different changes instead of only a few at a time, like in previous work.
The researchers were surprised to find that most of the mutations had an effect on gene expression, no matter where they occurred; virtually the entire enhancer seemed to be carrying important information. "The regulatory regions of genes encode information much more densely than previously appreciated," Stern said.
Single mutations were also found to have a variety of impacts in different areas of the fly; one mutation might elevate gene expression in one part and lower in in another.
"Both of these facts may constrain how regulatory regions can evolve," added Stern. "We hope it will lead people to start thinking about these regions differently and designing new kinds of experiments to explore this problem more deeply."
Crocker and EMBL engineers are now working on creating new tools and improving the Hybridizer. "It's allowing us to do entirely new types of experiments and answer questions that people haven't been able to touch before," Crocker said.