Developmental and genetic variation in nuclear microsatellite stability during somatic embryogenesis in pine

Abstract

Genotypic instability is commonly observed in plants derived from tissue culture and is at least partly due to in vitro-induced stress. In this work, the issues of whether genetic instability induced by in vitro stress varies among families and if genetic instability influences the adaptation to in vitro conditions and embryo development have been addressed. By comparing the stability of four variable nuclear microsatellite loci in embryogenic cultures and zygotic embryos of Pinus sylvestris, a significant difference in genetic stability among families was found. In six out of 10 families analysed, the level of genetic stability was similar between somatic and zygotic embryos. However, for the rest of the families, the mutation rate was significantly higher during somatic embryogenesis. Families showing a low genetic stability during establishment of embryogenic cultures had a higher embryogenic potential than those which were genetically more stable. In contrast, embryo development was suppressed in genetically unstable families. The relatively high mutation rates found for some families might reflect the plasticity of the families to adapt to stress, which is important for widely distributed species such as Pinus sylvestris.

Analysis of genetic stability at SSR loci during somatic embryogenesis in maritime pine (Pinus pinaster)

“…Somatic mutations can result in the production of genetic chimeras, where unchanged alleles are retained by some cells (López et al 2004;Burg et al 2007). The variation at locus FRPP91, obtained in ECL 49/432 under proliferation for 6 months, was not detected in the subsequent time points of analysis.…”

“…Despite the high frequency of abnormal phenotypes in plants derived from SE, these morphological traits could not be directly related to the genetic variation detected. In Pinus sylvestris, high mutation rates in SSR sequences were also found in embryogenic cells lines and cotyledonary embryos (Burg et al 2007). These authors suggest that the high mutation rates found might reflect the plasticity to adapt to stress, which also includes adaptation to in vitro conditions.…”

“…However, this process may be longer in duration depending on how long the cultures must be maintained under proliferation conditions in order to obtain sufficient amounts of tissue for regeneration of emblings. Several cases of somaclonal variation have been reported in embryogenic cultures of conifers including Larix decidua (Von Aderkas et al 2003), Picea abies (Burg et al 1999), Picea glauca (DeVerno et al 1999) and Pinus sylvestris (Burg et al 2007). Therefore, it appears that routine monitoring of somaclonal variation in SE-derived conifers would be prudent in order to maintain conformity and quality of cloned plants.…”

“…Some genetic molecular markers, such as RFLP (DeVerno et al 1994), RAPD (Isabel et al 1993;Fourré et al 1997) and AFLP (Hornero et al 2001), have been used for analysis of true-to-typeness. Microsatellites can serve as highly sensitive markers for monitoring genetic variation that may signal potential deleterious mutations during in vitro culture, because they reflect a relatively high rate of mutation and corresponding degree of genetic variability (Burg et al 1999;Leroy et al 2000;Wilhelm et al 2005;Lopes et al 2006;Burg et al 2007). Microsatellites consist of tandem repeats of short sequence motifs (2-8).…”

Somaclonal Variation During Picea abies and P. omorika Somatic Embryogenesis and Cryopreservation

“…Also, simple sequence repeats (SSR), also known as microsatellite markers, are used as the markers of choice to study the genetic conformity of plant material obtained via SE and/or stored in LN (Burg et al, 2007;Hazubska-Przybył et al, 2013;Helmersson et al, 2004;Lopes et al, 2009;Marum et al, 2009). Microsatellites are 1-6 nucleotide-long DNA sequences occurring extremely commonly in the genomes of Eucaryota in the form of perfect or near-perfect tandem iterations (Ellegren, 2004;Lopes et al, 2009).…”