Polyethylene glycol promotes maturation but inhibits further development of Picea abies somatic embryos

Abstract

A combined application of abscisic acid (ABA) and high molecular mass osmoticum, polyethylene glycol (PEG) has become a routine method for stimulating somatic embryo maturation in some genera of Coniferales. The goals of the present study were to clarify how the PEG 4000‐attributed low osmotic potential (ψs) of the maturation medium affects the yield and morphology of mature somatic embryos as well as subsequent developmental processes during germination and ex vitro plantlet growth in different genotypes of Picea abies belonging to 3 full sib seed families. Despite high within‐ and among‐family variation, a stimulatory effect of 7.5% PEG (ψs=−0.645 MPa) on somatic embryo maturation was recorded for 13 out of 17 cell lines (F= 2.83, P= 0.1). PEG‐treated somatic embryos were more dehydrated than embryos matured in the absence of PEG. Subsequently, embryos were partially desiccated using a high relative humidity treatment (HRH‐treatment). The dynamics of embryo water content (WC) during HRH‐treatment differed between embryos developed on maturation medium for 5 or 7 weeks. These two patterns remained unchanged irrespective of the ψs of the maturation medium. In 5‐week somatic embryos, the WC decreased to the lowest level (in the range 25‐35%) within the first 8 days of HRH‐treatment and was not further substantially changed. Seven‐week embryos also lost water within 8 to 16 days (decrease to 15‐25% WC), but this drop was followed by rehydration of embryonic tissues by 24th day of HRH‐treatment up to nearly the initial WC. Thus, 7‐week embryos experienced both desiccation and slow imbibition in the course of the 24‐day HRH‐treatment. This could account for their increased germinability compared to 5‐week somatic embryos found in the present study. Addition of 7.5% PEG to the maturation medium significantly inhibited somatic embryo germination for the vast majority of genotypes (F= 7.35; P= 0.01). Moreover, even after ex vitro transfer, both radicle elongation and lateral root formation were substantially suppressed (F= 3.8; P= 0.03) in those plantlets produced from PEG‐treated somatic embryos. Alterations both in the organization of the root meristem and in the structure of the root cap were found by histomorphological analysis of PEG‐treated somatic embryos. All those embryos possessed massive root caps with numerous intercellular spaces in the pericolumn tissue. Cells of the quiescent center exhibited clear symptoms of degradation manifested in shrinkage and collapse of the protoplasm. In addition, PEG‐treated embryos were of smaller size compared to embryos matured without osmoticum. When grown in artificial substrate (up to 5 months) the PEG‐induced inhibitory post‐effect gradually decreased. At this stage, the duration of maturation was the only factor separating plantlets into slow‐ and fast‐growing categories. Somatic embryos matured for 5 weeks produced plantlets twice the size of those produced by 7‐week embryos (F= 37.8; P < 0.0001). This trend did not depend on ψs of the maturation medium, nor on the genotype.