Vibratome Sectioning and Clearing for Easing Studies of Cassava Embryo Formation

Abstract

This work describes the application of clearing on vibratome sections to study the embryo formation in cassava. This procedure provides high-resolution images and reduces significantly the number of sections that need to be analyzed per ovule. This methodology was instrumental for the development of the protocol for embryo rescue in cassava. It has been also applied to monitor the embryo formation response when optimizing seed setting from regular and broad crosses for cassava breeding. Broad crosses between cassava and castor bean (incompatible-euphorbiaceae species) were made aiming to induce doubled haploids through the elimination of the incompatible-male parent genome as done in cereals. Castor bean is widely available and provides continues supply of pollen. Our results suggest that this methodology is easy and effective to assess the response of hundreds of cassava ovules pollinated with castor bean pollen, allowing the identification of multicellular structures in the embryo sac without apparent formation of endosperm. The protocol is also useful when developing and optimizing a methodology to induce doubled haploids in cassava via gynogenesis or from ovules pollinated with irradiated cassava pollen.

Fifty years of a public cassava breeding program: evolution of breeding objectives, methods, and decision-making processes

“…In spite of huge advances, there remains a significant knowledge gap relative to other major crops. As one important example, critical information regarding the flowering biology of cassava (receptivity of stigmas, pollen tube growth, embryo development) has only been addressed in the past three years (Lentini et al 2020a ; 2020b ; 2020c ; Pineda et al 2020a ; 2020b ; Ramos-Abril et al, 2019 ). Unfortunately, this lack of relevant basic information resulted in a serious bottleneck for the development of new technologies such as protocols for the production of doubled-haploids.…”

“…If public sector breeding programs are to take advantage of the lessons from the private sector (as suggested by Cobb et al 2019 ), then future cassava breeding should explore options to shift to the use of fully or partially inbred lines. Partial progress in that direction has already been made, with promising results regarding flower induction (Pineda et al 2020a , 2020b ), important progress toward the development of a protocol for the production of doubled-haploids in cassava (Lentini et al 2020a , 2020b , 2020c ), and the ongoing efforts to edit the cassava genome for the expression of haploid induction.…”