Figure 1.
Schematic representation of two somatic embryogenesis processes applied at the industrial level.
The first somatic embryogenesis process (upper section of the flow diagram) involved a proliferation step based on secondary embryogenesis in RITA® temporary immersion bioreactors (photos 1A, 1B). The second process (lower section of flow diagram) included a proliferation step based on embryogenic suspensions (photos 1C, 1D). 1A, initial developmental stages of secondary embryos at the root pole of primary somatic embryos; 1B, clusters of primary and secondary embryos; 1C, clusters of embryogenic cells in suspension; 1D, embryogenic cells in suspension.
Table 1.
Frequency of coffee phenotypic variants detected in the nursery and field on three year-old plants from two C. arabica hybrids depending on the type of proliferation system used in the industrial somatic embryogenesis process.
Figure 2.
Proportions (%) of the different types of phenotypic variants in comparison to the total number of variants.
Variants representing less than 1% of somatic embryogenesis-derived plants were observed in C. arabica embling batches at nursery (A) and field (B) levels, depending on the proliferation system used, i.e. secondary embryogenesis (SCE) or embryogenic suspension (ESP). In the nursery, the data were obtained from 239 009 emblings derived from SCE and 402 576 emblings from ESP. In the field, the data were obtained through the observation of 100 257 emblings derived from secondary embryogenesis and 108 784 from embryogenic suspensions.
Figure 3.
Examples of different C. arabica phenotypic variants in plants mass propagated through somatic embryogenesis.
A, plant exhibiting a normal phenotype in the nursery; B, Angustifolia variant in the nursery; C, Variegata variant in the field; D, plant showing a normal phenotype in the field; E, Dwarf variant in the field; F, Bullata variant in nursery.
Table 2.
Primer combinations used for AFLP and MSAP analyses.
Table 3.
Summary of AFLP data and observed polymorphisms among mother plants and emblings derived from secondary embryogenesis or embryogenic suspensions.
Figure 4.
Representation of MSAP electropherograms observed for coffee mother plants and embling progeny using the isoschizomers HpaII and MspI.
Illustration of the pattern variation obtained for normal and variant phenotypes within the embling progeny.
Table 4.
MSAP patterns corresponding to different methylation states of the symmetric sequence CCGG, as revealed by the specificity of the restriction enzymes Hpa II and Msp I.
Table 5.
MSAP methylation patterns in mother plants and modified patterns in emblings.
Table 6.
Overall MSAP data and methylation polymorphism among mother plants and emblings of C. arabica hybrids derived from secondary embryogenesis or embryogenic suspensions.
Figure 5.
Methylation polymorphism accumulation in coffee emblings showing a normal vs. variant phenotype depending on the somatic embryogenesis process used.
For the secondary embryogenesis process, data were derived from the analysis of 59 phenotypically normal and 19 variant emblings. For the embryogenic suspension process, 59 phenotypically normal and 8 variant emblings were studied.
Figure 6.
Mitotic cells at metaphase or prometaphase stages and observed ploidy levels of some normal and variant emblings from the allotetraploid C. arabica species (2n = 44).
Karyotype analyses were performed by counting chromosomes on four to eight clear metaphase spreads obtained from root tips of three year-old plants.
Table 7.
Summary of chromosome counting in some normal versus variant C. arabica hybrids derived from somatic embryogenesis.
Table 8.
Plant material used in molecular marker analyses.