Indirect somatic embryogenesis in Coffea with different ploidy levels: a revisiting and updating study

Indirect somatic embryogenesis (ISE) is required for plant propagation and a prerequisite for applications that may provide new germplasms. Genetic, epigenetic and physiological features of the explant donor are barriers for ISE establishment, hindering its wide use. Despite the identification and/or expression analysis of genes during ISE, no approach to establish the karyotype aspects has been performed so far. So, this study aims to establish the ISE and compare the in vitro responses between diploid (Coffea canephora and Coffea eugenioides), allotriploid (“Híbrido de Timor”—HT) and true allotetraploid (Coffea arabica) Coffea in a taxonomic and evolutive scenario. Under the same in vitro conditions, the four Coffea differed from each other during ISE. Leaf explants of the true allopolyploids yielded the highest mean number of friable calli (FC) in relative short time and visually exhibiting more pronounced length. FC of the allotetraploid C. arabica presented the highest mean number of mature cotyledonary somatic embryos (MCSE), which were also recovered faster in this species. However, MCSE mean number in HT was the same or lower than diploid Coffea. Besides, intraspecific variation related to the ISE responses was observed in each Coffea, mainly the mean number of FC obtained from ex vitro and in vitro C. arabica and C. eugenioides explants. So, epigenetic and physiologic features may also have influenced the ISE responses. The findings provide the basis for performing other approaches considering the ploidy level, epigenetic and physiological backgrounds. Besides, the data also contributed for understanding about the consequences of polyploidy

Indirect somatic embryogenesis in Coffea with different ploidy levels: a revisiting and updating study

Indirect somatic embryogenesis (ISE) is required for plant propagation and a prerequisite for applications that may provide new germplasms. Genetic, epigenetic and physiological features of the explant donor are barriers for ISE establishment, hindering its wide use. Despite the identification and/or expression analysis of genes during ISE, no approach to establish the karyotype aspects has been performed so far. So, this study aims to establish the ISE and compare the in vitro responses between diploid (Coffea canephora and Coffea eugenioides), allotriploid (“Híbrido de Timor”—HT) and true allotetraploid (Coffea arabica) Coffea in a taxonomic and evolutive scenario. Under the same in vitro conditions, the four Coffea differed from each other during ISE. Leaf explants of the true allopolyploids yielded the highest mean number of friable calli (FC) in relative short time and visually exhibiting more pronounced length. FC of the allotetraploid C. arabica presented the highest mean number of mature cotyledonary somatic embryos (MCSE), which were also recovered faster in this species. However, MCSE mean number in HT was the same or lower than diploid Coffea. Besides, intraspecific variation related to the ISE responses was observed in each Coffea, mainly the mean number of FC obtained from ex vitro and in vitro C. arabica and C. eugenioides explants. So, epigenetic and physiologic features may also have influenced the ISE responses. The findings provide the basis for performing other approaches considering the ploidy level, epigenetic and physiological backgrounds. Besides, the data also contributed for understanding about the consequences of polyploidy

From chromosome doubling to DNA sequence changes: outcomes of an improved in vitro procedure developed for allotriploid “Híbrido de Timor” (Coffea arabica L. × Coffea canephora Pierre ex A. Froehner)

Since 1966, chromosome doubling has been performed mainly in vitro, associating anti-tubulin treatment and different plant tissues showing proliferative cells. Despite the achieved improvements, some bottlenecks have been pointed out, such as the low rate of polyploids and high rate of mixoploid plantlets. To overcome these hurdles, some approaches have indicated that indirect somatic embryogenesis (ISE) constitutes an alternative trigger for chromosome doubling, especially for homoploid and anorthoploid germplasms. In this way, a guideline has been developed for hexaploidization of the Coffea line “Híbrido de Timor” (HT) ‘CIFC 4106’ (anorthoploid, 3x = 33 chromosomes, 1C = 2.10 pg, Coffea canephora × Coffea arabica) from friable embryogenic calli (FEC) treated with colchicine. From this, a relatively high percentage (49.3%) of HT hexaploids (6x = 66 chromosomes, 2C = 4.20 pg) was obtained, without recovery of mixoploids. Besides confirmation of endomitosis induction through the obtained hexaploids, SSR markers revealed that the FEC/colchicine strategy also resulted in loss of allelic diversity in 39 regenerated HT plantlets, demonstrating its genotoxic effect. Considering these results, the present procedure resolved the main bottlenecks for chromosome doubling, which have been reported since the discovery and isolation of the anti-tubulin colchicine in 1930. Hexaploid HT plantlets have enriched Coffea germplasm banks as a new genetic resource since the resolution of their karyotype and DNA sequence. Just as the true allotetraploid C. arabica and the allotriploid HT ‘CIFC 4106’, the hexaploid HT is relevant to investigate the genomic and phenotypic changes arising from polyploidization events