The ∆ 1 -pyrroline-5-carboxylate synthetase (P5CS) is the rate-limiting enzyme in proline (Pro) synthesis and is involved in drought and salt stress tolerance in plants. An OsP5CS gene was isolated from a stress-treated commercial rice variety, BC15.
The length of rice OsP5CS was 2173 nucleotides, containing an ORF encoding for 716 amino acids with two regulatory amino acid residues Asp and Phe. Amino acid sequence alignment of P5CS proteins indicated that P5CS has homology among rice, corn, tobacco, bean and tomato. BC15 OsP5CS also showed the 99.6% similarity to OsP5CS of GeneBank-registed Nipponbare rice (AC111016.2).
Based on the DNA isolated sequence, four gRNA (guide RNA) constructs were designed for CRISPR-Cas9 editing of BC15 OsP5CS in order to increase the Pro accumulation in cells. This study laid the foundation for the development of high-yielding rice varieties resistant to drought and salinity using genome editing technology.