High Genetic and Epigenetic Stability in Coffea arabica Plants Derived from Embryogenic Suspensions and Secondary Embryogenesis as Revealed by AFLP, MSAP and the Phenotypic Variation Rate

Abstract

Embryogenic suspensions that involve extensive cell division are risky in respect to genome and epigenome instability. Elevated frequencies of somaclonal variation in embryogenic suspension-derived plants were reported in many species, including coffee. This problem could be overcome by using culture conditions that allow moderate cell proliferation. In view of true-to-type large-scale propagation of C. arabica hybrids, suspension protocols based on low 2,4-D concentrations and short proliferation periods were developed. As mechanisms leading to somaclonal variation are often complex, the phenotypic, genetic and epigenetic changes were jointly assessed so as to accurately evaluate the conformity of suspension-derived plants. The effects of embryogenic suspensions and secondary embryogenesis, used as proliferation systems, on the genetic conformity of somatic embryogenesis-derived plants (emblings) were assessed in two hybrids. When applied over a 6 month period, both systems ensured very low somaclonal variation rates, as observed through massive phenotypic observations in field plots (0.74% from 200 000 plant). Molecular AFLP and MSAP analyses performed on 145 three year-old emblings showed that polymorphism between mother plants and emblings was extremely low, i.e. ranges of 0–0.003% and 0.07–0.18% respectively, with no significant difference between the proliferation systems for the two hybrids. No embling was found to cumulate more than three methylation polymorphisms. No relation was established between the variant phenotype (27 variants studied) and a particular MSAP pattern. Chromosome counting showed that 7 of the 11 variant emblings analyzed were characterized by the loss of 1–3 chromosomes. This work showed that both embryogenic suspensions and secondary embryogenesis are reliable for true-to-type propagation of elite material. Molecular analyses revealed that genetic and epigenetic alterations are particularly limited during coffee somatic embryogenesis. The main change in most of the rare phenotypic variants was aneuploidy, indicating that mitotic aberrations play a major role in somaclonal variation in coffee.

Assessment of genetic and epigenetic changes during cell culture ageing and relations with somaclonal variation in Coffea arabica

“…Observation of the same behaviour for the three independent cell lines strongly supported the existence of a nonrandom mechanism for SV. Low levels of SV (around 1 %) were previously found in C. arabica plants regenerated from recently obtained callus or from 6-month-old cell cultures (Etienne andBertrand 2001, 2003;Bobadilla Landey et al 2013).…”

“…Nevertheless, when applied over a short 6-month period in the presence of a low growth regulator supply, embryogenic cell cultures gave rise to very low SV rates, as observed through massive phenotypic observations in field plots (0.74 % from 200,000 plants) (Bobadilla Landey et al 2013). Molecular analyses performed on the regenerated plants revealed that genetic and epigenetic alterations were particularly limited.…”

Genetic Fidelity Testing Using SSR Marker Assay Confirms Trueness to Type of Micropropagated Coconut (Cocos nucifera L.) Plantlets Derived from Unfertilized Ovaries

“…The present study indicates that the level of hormones especially 2, 4-D and BAP used for coconut plant regeneration is good enough to retain the genetic integrity of regenerated plants indicating the above hormone levels not to be deleterious to genetic integrity of coconut cultures even though the protocol is developed through a callus phase. High genetic and epigenetic stability has been reported in coffee (Coffea Arabica) plants derived from embryogenic suspensions [30]. It has also been reported that long term culture and cell culture ageing in coffee, resulting in high rates of mutations and chromosomal re-arrangements which are directly linked to somaclonal variations (31).…”

Field Performance and Genetic Fidelity of Micropropagated Plants of Coffea canephora (Pierre ex A. Froehner)

“…Similarly, Zarghami et al [15] evaluated the genetic stability of cryopreserved and non-cryopreserved potato and obtained 97-100% similarity using AFLP markers. Recently, Landey et al [19] observed high genetic and epigenetic stability of Coffea arabica plants derived from embryogenic suspensions as well as secondary embryogenesis using AFLP and MSAP markers. The present findings of high genetic similarities between mother plants and the plants derived from somatic embryogenesis are concurrent with the earlier findings.…”

“…Various molecular markers such as RAPD [11][12][13][14], AFLP [15,16], SSR [17] and ISSR [18] are used for determining the genetic fidelity of micropropagated plants. In coffee (C. arabica), Landey et al [19] analyzed genetic fidelity of in vitro propagated plants of C. arabica using AFLP and SSAP markers and confirmed that genetic variation observed in micropropagated plants are caused by abnormal chromosome numbers. Interestingly, these authors could not detect genetic variability among micropropagated plants using AFLP markers.…”