Preprint Article Version 1 Preserved in Portico This version is not peer-reviewed

Temporary Immersion System Improves Regeneration of in vitro Irradiated Recalcitrant Rice Embryogenic Calli

Version 1 : Received: 23 December 2021 / Approved: 24 December 2021 / Online: 24 December 2021 (14:47:29 CET)

A peer-reviewed article of this Preprint also exists.

Hernández-Soto, A.; Pérez, J.; Fait-Zúñiga, R.; Rojas-Vásquez, R.; Gatica-Arias, A.; Vargas-Segura, W.; Abdelnour-Esquivel, A. A Temporary Immersion System Improves Regeneration of In Vitro Irradiated Recalcitrant Indica Rice (Oryza sativa L.) Embryogenic Calli. Plants 2022, 11, 375. Hernández-Soto, A.; Pérez, J.; Fait-Zúñiga, R.; Rojas-Vásquez, R.; Gatica-Arias, A.; Vargas-Segura, W.; Abdelnour-Esquivel, A. A Temporary Immersion System Improves Regeneration of In Vitro Irradiated Recalcitrant Indica Rice (Oryza sativa L.) Embryogenic Calli. Plants 2022, 11, 375.

Journal reference: Plants 2022, 11, 375
DOI: 10.3390/plants11030375

Abstract

The development of gamma rays mutant rice lines would be a solution for introducing variability in already farmer using varieties. In vitro gamma (60Co) mutagenesis reduces chimeras and allows a faster selection of desired traits but requires laboratory process optimization. The objective of the present work was the in vitro establishment of a recalcitrant rice embryogenic calli, the determination of its sensitivity to gamma radiation (Co-60), sequencing MATK and Rubisco for identification purposes, as well as generation optimization. The radiosensitivity of embryogenic calli resulted in an LD50 of 110Gy, while the 20% lethal dose was 64Gy. All sequenced genes matched perfectly with already reported MATK and Rubisco O. sativa genes with a clear SNP that identifies the local variety related to the southeast Asia Region. Callus induction improves with an MS with 2mg/L 2,4D, and the regeneration was achieved with an MS medium with 3mg/L BAP and 0,5mg/L NAA. The optimized radiation condition was 60Gy with an 83% regeneration in a semisolid medium, allowing a balance between mutation and regeneration. When increased to 80Gy, the regeneration rate falls to 29%. An immersion system (RITA®) of either 60 or 120 seconds every 8hours allowed a systematic and homogeneous total regeneration of the recalcitrant line, in contrast with the semisolid medium that resulted in positive but irregular regeneration. Other well-known recalcitrant cultivars, CR1821, CR1113 also had an improving regeneration in the immersion system, demonstrating its potential use for recalcitrant materials. To our knowledge, this is the first report on using an immersion system to allow regeneration of gamma-ray mutants from recalcitrant rice materials.

Keywords

Embryogenesis; Cobalt-60 radiation-induced mutagenesis; Temporary immersion systems (TIS).

Subject

LIFE SCIENCES, Biotechnology

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