Lawrence Yves Uy | Yvonne Ludwig | Merlyn S. Mendioro | Ma Carmina Manuel | Jorge Gil Angeles | Inez Slamet-loedin
An estimated 4 billion people depend on rice as their staple food and this number increases rapidly. As such, increasing its production is a major challenge. A suggested solution for this is gene editing yield-related regions of interest in elite indica rice varieties such as IR64 and Samba Mahsuri (SM) since these varieties are already widely accepted in the market. One promising region of interest is the THOUSAND GRAIN WEIGHT 6 (TWG6) because it has shown promising influence in grain weight when knocked out in japonica varieties. In this study, TGW6 was knocked down in IR64 and SM rice varieties using Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-Cpf1 system. A CRISPR Cpf1 vector construct (IRS-1466) was assembled to target TGW6 in both IR64 and SM. The resulting knockout IR64 and SM were analyzed through PCR screening for the presence of CRISPR gene while cutting efficiency and mutational changes in the region of interest were analyzed using T7 assay and sequencing, respectively. Knockout SM had 81.7% cutting efficiency while knockout IR64 had 94.34% cutting efficiency. Sequence analysis revealed minimal single-nucleotide insertion-deletions (indels) in the region of interest. Phenotypic data showed a significant increase in total grain weight of both knockout SM (increased up to 33.5%) and IR64 (105.11%) compared to the wild-type controls. In addition, an increase in grains per panicle was also observed in both SM and IR64 knockouts. These results reveal that targeted knockdown via CRISPR-Cpf1 of TGW6 increases grain weight thus, increasing rice grain yield which has great potential as a possible solution in the foreseeable rice production challenges ahead.
ISSN 2719-1990 (Print)
Philippine E-Journals
