Journal of Agronomy and Field Innovations

Background: Brassica juncea (L.) Czern., better known as Indian mustard, is an oilseed and condiment crop that is grown worldwide and needs a large amount of synthetic nitrogen (N) fertilizer in order to continue to produce high yields. The efficient use of nitrogen (NUE) without resulting in low yields is one of the major goals of the modern approach to crop breeding. 
Objective: This study was conducted to assess prospects of CRISPR/Cas12a based genome editing in improving NUE in B. juncea cv. Varuna by selectively altering genes involved in nitrogen uptake, transport, assimilation, and regulation. 
Methods: A Cas12a-platform-based method was conceptually used on four candidate loci—BjuNRT2.1, BjuAMT1.2, BjuGS1;2, and the nitrate-sensing silencer BjuNIGT1—based on their functional similarities with known nitrogen-responsive genes in some other Brassicaceae and cereal species. Edited and non-edited varieties were compared on features under low, moderate, and high nitrogen levels using morphological, physiological, molecular, and agronomic parameters. 
Results: Genome-edited lines demonstrated significant enhancements in root architecture, chlorophyll reserves, nitrogen uptake efficiency, as well as nitrogen utilization efficiency compared to the unedited controls, especially under low and medium nitrogen conditions. Increase in transcript levels of nitrogen transporters and assimilators in the edited lines coincided with the activity of glutamine synthetase and nitrate reductase and improved their biomass and seed yield per unit of nitrogen used. Gain in fertilizer-use efficiency was significant enough for possibility of nitrogen rate decrease in practice without losses of yield.
Significance: The research reveals that the editing of the nitrogen-responsive regulatory and transport genes using Cas12a technology can greatly improve nitrogen use efficiency in B. juncea in addition to the use of agricultural practices such as traditional breeding methods and nitrogen management.
Conclusion: CRISPR/Cas12a genome editing is a scientifically sound and translatable strategy for developing nitrogen-efficient mustard genotypes that can lead to decreased fertilizer dependency and more sustainable oilseed production systems.