Study of the limiting factors for obtaining haploid plants from Coffea arabica Estudio de los factores limitantes para la obtención de plantas haploides de Coffea arabica

The development of an efficient method to obtain coffee haploid plants requires the study of the most limiting factors. This work of research evaluated the development of male gametes in relation to the phenological state of the C. arabica flower, flower disinfection treatments, treatments for isolation and purification of microspores. The thermal effect on the induction of androgenesis and the treatments with antimitotic agents for chromosomal duplication of haploid tissues were also evaluated. To induce the androgenesis process, flower buds between 1 and 1.7 cm in length were determined to contain the highest percentage of gametic cells in the late uninucleate and early binucleate states, crucial for ontogenetic change. It was  stablished that the lowest percentage of microbial contamination and biological oxidation was achieved with the use of calcium hypochlorite combined with antioxidants. The isolation and purification of the microspores was obtained by mechanical maceration of the anthers and filtration of the gametic cells in 40 and 70 micron-meshes together with the use of Percoll in gradients higher than 50%. The greatest androgenic induction of microspores was achieved by incubating the flower buds detached from the branch at 4°C, observing cell divisions and multicellular colonies formation. The application of Colchicine (1%) to meristems of haploid plants of C. arabica allowed to obtain branches with leaves that duplicated their chromosomal
material and changed their morphology