Improved organogenic capacity of shoot cultures from mature pedunculate oak trees through somatic embryogenesis as rejuvenation technique

Abstract

Theoretically, complete rejuvenation of mature trees should occur through somatic embryogenesis, however, this has not been extensively studied. The main objective of the present study was to increase the efficiency of in vitro clonal propagation for mature Quercus robur (100–300 years old), by induction of somatic embryogenesis as rejuvenation step prior to establishment of shoot culture through micropropagation of somatic embryo-derived plantlets. Shoot culture lines of “mature” origin were established from epicormic shoots of two centenarian oak genotypes (Sainza and CR-0) and maintained by axillary shoot proliferation. Embryogenic lines were also initiated from epicormic leaf explants of the same genotypes and maintained by secondary somatic embryogenesis. Although the frequency of somatic embryo conversion into plantlets was low in pedunculate oak, shoot culture lines could be established and maintained by axillary branching from several germinated somatic embryos. For each genotype and shoot culture line of the two origins (mature tree and somatic plantlets), shoot multiplication rate and elongation as well as rooting ability parameters were compared. Compared with “mature-origin” shoot cultures and after more than one year propagation in vitro, shoot lines established from somatic plantlets produced a significantly higher proportion of elongated, rootable shoots (from 26.0–31.6 to 36.8–40.5%) with increased rooting ability (from 3.3–45.6% to 23.2–89.8%). In the case of 300-year-old Sainza genotype such a high organogenic capacity was similar to shoot cultures initiated from basal sprouts. Basal sprouts are considered as “mature” material that retains juvenile characteristics compared with epicormic shoots forced from crown branches. Somatic embryogenesis only slightly improved plant regeneration of shoot cultures from basal sprouts, thus validating their use as “juvenile control”. The present results provide evidence that some rejuvenation occurred during the process of somatic embryogenesis and resulted in improved shoot growth and rooting of somatic embryo-derived culture compared with “mature” shoot culture. The results reported in this study might be useful in embryogenic systems with low plant conversion rates. The proposed experimental model might also be useful in finding molecular markers of plant ontogeny.