Maize with impaired DNA-repair mechanism

The maize plants have been edited using the CRISPR/Cas9 system resulting in the mutation of the ATR gene. One extra DNA base pair has been introduced into the target gene resulting in the knock-out of this gene by means of a frameshift mutation.

The ATR gene plays a role in the repair of DNA damage. Thus, DNA damage is expected to accumulate to a higher degree due to the faulty repair mechanism. The plants could acts as a biosensor for DNA damage caused by different forms of abiotic stress such as drought, heat, or environmental pollution.

Zea Mays inbred line B104

pBUN411

To introduce the CRISPR/Cas9 gene cassette, plants were transformed using Agrobacterium tumefaciens-mediated transformation. The CRISPR/Cas9 machinery introduced a single base pair insertion within the coding sequence of the ATR gene, which caused a frameshift mutation. Transformants containing the gene cassette and the desired mutations were selected, and by means of conventional crossing with wild-type plants, the CRISPR/Cas9 gene cassette was removed by selecting T1 plants that only contained the desired mutation, but no longer contained the gene cassette (null-segregants). Therefore, the plants in the field trial are not expected to contain foreign genetic material. They should only contain the desired mutation.

The gene has been edited using the CRISPR/Cas9 system resulting in a frameshift mutation. Therefore, the full length protein is not expected to be present within the modified plants.

Please note that two additional lines were also created to knock-out ATR activity. One contains the full deletion of the gene and the other a single basepair insertion in a different location to also cause a frameshift mutation.

Primarily, ATM responds to double stranded breaks, whereas ATR is activated by single stranded DNA and defects in replication fork progression.