Based on the article: Figueredo, E.F., da Cruz, T.A., de Almeida, J.R., Batista, B.D., Marcon, J., de Andrade, P.A.M., de Almeida Hayashibara, C.A., Rosa, M.S., Azevedo, J.L. and Quecine, M.C., 2023. The key role of indole-3-acetic acid biosynthesis by Bacillus thuringiensis RZ2MS9 in promoting maize growth was revealed by the ipdC gene knockout mediated by the CRISPR-Cas9 system. Microbiological Research, 266, p.127218.

1) Use a flow diagram to describe how the CRISPR-Cas9 system was designed and used for the indole-3-pyruvate decarboxylase (ipdC) gene that was knocked out. (8)

2) Comment on the efficacy of the reported CRISPR-Cas9 system for the ipdC gene knockout by evaluating the experimental design and the results obtained. (8)

3) Propose an alternate gene knockout technique that could have been used and briefly detail its principle.