Optimized somatic embryogenesis in Pinus strobus L.

Abstract

Somatic embryogenesis (SE) initiation in Pinus strobus was optimized by the manipulation of plant growth regulator (PGR) concentrations in the culture medium. Modified Litvay medium (MLV) of Litvay et al. (1985) supplemented with lower than routinely used PGR concentration increased initiation of established embryogenic cultures from approximately 20 to 53%. The original developmental stage of zygotic embryos had a pronounced effect on the SE response. The optimum stage was the pre-to shortly post-cleavage stage. A substantial genetic influence on initiation of SE was indicated by a significant variance component due to families. Genotype  collection date and genotype  media interactions had large effects on initiation of SE. The PGR levels in the culture medium prior to maturation had a significant effect on subsequent production of mature somatic embryos. Embryogenic tissue initiated and proliferated on medium with a low level of PGR consistently produced a high number of somatic embryos, indicating that optimized initiation protocol also enhanced somatic embryo production. Somatic embryos of 93 embryogenic lines (representing five families) that were initiated on media with different PGR concentrations were converted to plants at an overall frequency of 76%, and grown in the greenhouse. With these improved protocols, application of P. strobus SE in commercial clonal forestry is feasible as an alternative to traditional breeding and reforestation.

Implementation of conifer somatic embryogenesis in clonal forestry: technical requirements and deployment considerations

“…There was no correlation among the different phases of SE. There was also a significant genetic influence in initiation of SE in P. strobus [7].…”

“…The initiation rate in pine species had been low. However, we recently achieved a high SE initiation rate in Pinus strobus by manipulating plant growth regulators [7]. There seem to be opportunities for improving SE initiation in semi-recalcitrant and recalcitrant species by manipulating initiation media, but there are large differences in SE initiation among species.…”

“…In P. strobus, for example, the most responsive stage of ZE development is immediately following fertilization, i.e., pre-and post-cleavage stage before the appearance of a dominant embryo. This is evidenced by a sharp increase in induction rate during the anticipated fertilization period and subsequent gradual decline as ZE development continues (figure 2) [7]. Careful monitoring of the ZE development stage may result in high initiation rates.…”

“…LH is optimized initiation protocol based on Litvay medium and AFC is initiation medium formulated in our laboratory with different base elements than LH medium. Both media had the same optimized levels of growth regulators [7].…”

The effect of culture medium formulation on Pinus nigra somatic embryogenesis

“…embryogenic tissues have been initiated (Salajova and Salaj 1992) and the effect of zygotic embryo developmental stage (Salajova et al 1995), different concentration as well as combination of plant growth regulators and carbon source on the initiation process have been tested (Salajova and Salaj 2005). In several conifer species it was demonstrated the mineral composition of culture medium affected the initiation of somatic embryogenesis as well as somatic embryo maturation (Becwar et al 1990, Klimaszewska et al 2001. In Pinus nigra this factor has not been studied yet.…”