New tools for accelerating the genetic discovery of crops

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Commercially viable biofuel crops are essential for reducing greenhouse gas emissions, and new tools developed by the Advanced Bioenergy and Bioproduct Innovation Center (CABBI) will accelerate their development. , Should promote the overall progress of gene editing.

Crop genomes have been tuned by breeding generations to optimize specific traits, and until recently breeders were limited to naturally occurring diversity choices. CRISPR / Cas9 gene editing technology can change this, but so far the software tools needed to design and evaluate CRISPR experiments are so far. Mammalian genomeDo not share the same characteristics as the complex crop genome.

The first open-source software tool for genome-wide design and evaluation of guide RNA (gRNA) sequences for CRISPR experiments, created by scientists at CABBI, a bioenergy research center (BRC) funded by the Ministry of Energy. Join CROPSR, which is. A genome-wide approach significantly reduces the time required to design CRISPR experiments, reduces crop handling work, and accelerates the design, evaluation, and validation of gRNA sequences, according to a study published in. BMC Bioinformatics..

“CROPSR provides the scientific community with new ways and new workflows for performing CRISPR / Cas9 knockout experiments,” said Hans Müller Paul, a molecular biologist and CROPSR developer with a PhD. Co-author Matthew Hudson, a student with a professor of crop science at the University of Illinois at Urbana-Champaign. “We hope that new software will accelerate discovery and reduce the number of failed experiments.”

To better meet the needs of crop geneticists, the team has built software that lifts the restrictions imposed by other packages on the design and evaluation of gRNA sequences. This is a guide used to find the genetic material of interest. Team members have also developed a new machine learning model that does not avoid the problem of existing tools, the guide to repetitive genomic regions commonly found in plants. The CROPSR scoring model provided much more accurate predictions, even in non-crop genomes, the authors said.

“The goal was to incorporate features that would make life easier for scientists,” says Muller Paul.

Many crops, especially bioenergy sources, have a very complex polyploid genome with multiple sets of chromosomes. Also, some gene editing software tools based on the diploid genome (such as those of humans) have problems with the characteristics of the crop genome.

“It can take weeks or months to realize that you don’t get the results you expect,” Muller Paul said.

For example, traits can be regulated by a collection of genes, especially those with stressful plants whose backup system is useful. Scientists may design experiments so that one gene is knocked out and another gene that performs the same function is unnoticed. This problem may not be discovered until the plant matures without changing its traits. This is a problem specific to crops that require specific weather conditions to grow and can mean a year’s delay if the season is missed.

Using a genome-wide approach, scientists were able to tailor CROPSR to plant use by removing the built-in bias found in existing software tools. Because they are based on human or mouse genomes where multiple copies of the gene are less common, these tools have genomes at multiple locations to avoid the occurrence of mutations in unintended locations. Penalizes gRNA sequences that hit. But for crops, the goal is often to change multiple positions to knock out all copies of the gene. Previously, scientists had to design four or five mutation experiments to knock out each gene individually, which required extra time and effort.

CROPSR can generate a database of CRISPR-guided RNAs that can be used throughout the crop genome. This process is computationally intensive and time consuming, usually several days, but researchers only need to run it once to build the database and it can be used for continuous experiments.

So instead of searching an online database for the gene of interest, using current tools to design individual guides in five different locations, and conducting multiple experiments, scientists can find the gene in their database. You can search and see all the guides available. CROPSR also indicates other targeted locations within the genome. Researchers can choose a guide that applies to all genes, making experiment design much easier and faster.

“You can access the database to get all the information you need, get ready, and get started,” says Muller Paul. “The less time you spend planning an experiment, the more time you can spend on it.”

For CABBI scientists who often work with repetitive plant genomes, having a gRNA tool that can confidently design a working guide “should take a step forward,” he said.

As the name implies, CROPSR was designed with the crop genome in mind, but it can be applied to all types of crops. genome..

“CROPSR is also based on human genes because we don’t have crop gene data yet, but we are considering collaborating with other BRCs to provide more competent predictions based on biophysics. Helps mitigate some of the problems caused by the lack of. “

In the future, we hope that researchers will be able to record failure results and successes and generate data to train crop-specific models. If the collaboration is successful, “training machine learning models for CRISPR applications, and even other models, can make very interesting advances.”

The other co-author of this study is Davey Stunt, a former CABBI graduate student at Hudson, University of Illinois, School of Crop Science. Jacob Helden Brand, a former CABBI research programmer at the National Supercomputing Application Center in Illinois. Hudson and Muller Paul are also affiliated with the Illinois Institute of Informatics and the Karl R. Wars Institute for Genomic Biology.

Genome editing tool TALEN is superior to CRISPR-Cas9 in dense DNA

For more information:
Hans Müller Pauletal, CROPSR: An automated platform for the design and validation of complex genome-wide CRISPR gRNAs BMC Bioinformatics (2022). DOI: 10.1186 / s12859-022-04593-2

Quote: CROPSR: A new tool to accelerate gene discovery in crops (February 17, 2022) from https: // in 2022 Obtained on February 17th.

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New tools for accelerating the genetic discovery of crops

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