CROPSR: A New Tool to Accelerate Gene Discovery

The genomes of crops are tailored through generations of breeding to optimize specific traits, and until recently breeders were limited to selecting for naturally occurring diversity

CRISPR/Cas9 gene editing technology could change this, but so far, the software tools required to design and evaluate CRISPR experiments have been based on the need for editing mammalian genomes, which do not share the same features as complex crop genomes

CROPSR is the first open-source software tool for genome-wide design of CRISPR experiments and evaluation of guide RNA (gRNA) sequences, created by scientists at the Department of Energy-funded Bioenergy Research Center (BRC)

"CROPSR provides the scientific community with new methods and workflows for conducting CRISPR/Cas9 knockout experiments," said CROPSR developer Hans Müller Paul

To better meet the needs of crop geneticists, the team built software that removes the limitations imposed by other software packages when designing and evaluating gRNA sequences, the guidelines used to locate target genetic material

"Our goal was to combine features that make life easier for scientists," Müller Paul said

Many crops, especially bioenergy feedstocks, have highly complex polyploid genomes with multiple sets of chromosomes

"It sometimes takes weeks or months to realize that you're not getting the results you expected," says Müller Paul

For example, a trait may be regulated by a range of genes, and backup systems are useful especially when plant stress is involved

By using a genome-wide approach, the scientists removed the inherent biases present in existing software tools, thereby tailoring CROPSR for plant use

CROPSR can generate a database of usable CRISPR guide RNAs for the entire crop genome

So instead of searching through online databases for a target gene, scientists can search for genes in their own databases, see all available guides, and then use existing tools to design separate guides for five different sites and run multiple rounds of experiments

"You can go right into the database, get all the information you need, and get to work," says Müller Paul

For CABBI scientists, who often study repetitive plant genomes, having a gRNA tool that would give them the confidence to design functional guides "should be a step forward," he said

As the name suggests, CROPSR is designed with crop genomes in mind, but it works with any type of genome

"CROPSR is also based on human genes, as data availability for crop genes has not yet formed," Müller Paul said, "but we are investigating some collaborations with other brcs to provide a more capable prediction based on biophysics, helping to alleviate some of the problems caused by lack of data

Going forward, he hopes that researchers will document their failures and successes to help generate data to train a model for a specific crop
.
If the collaboration is successful, "we could see some very interesting advances in training machine learning models for CRISPR applications, and possibly other models as well
.
"

Hans Müller Paul, Dave D.
Istanto, Jacob Heldenbrand, Matthew E.
Hudson.
CROPSR: an automated platform for complex genome-wide CRISPR gRNA design and validation .
BMC Bioinformatics , 2022; 23 (1)