Fate of Exogenous DNA in Arabidopsis thaliana

Abstract

It has been shown that when Arabidopsis seeds are incubated with different heteropycnic labelled DNA during the first 4 days of germinating, large fragments of exogenous DNA are taken up by the seedling plant tissues (cotyledons and rootlet) and remain polymerized and double-stranded during the entire growth period. Only a small percentage of the population of exogenous DNA molecules appears to be destroyed and reutilized for DNA synthesis de novo. Most of the exogenous DNA remains free or becomes integrated with the Arabidopsis DNA as double-stranded molecules, not separable by alkali denaturation, but easily separable by ultrasonication.Analysis of Werent tissues and organs of growing plants reveals that a large amount of foreign DNA remains in the cotyledons during the vegetative growth phase, but is redistributed a t the flowering stage (when cotyledons etiolate). At this time, the foreign DNA leaves the cotyledons and migrates towards the flowering buds where it accumulates as a highly molecular and double-stranded material. I n the E", progeny, radioactive molkcules are found which do not correspond to the endogenous material and which have a density depending on that of the foreign DNA used.Arabidopsis thaliana (L.) Heyhn, is a facultative long-day autogamous crucifer (2 n = 10 chromosomes) which has a short life cycle (about 35 days under our experimental conditions), and can be mass cultured in a relatively small area. After the incubation, the seeds were washed and grown aseptically in closed glass vessels on perlite soaked with a mineral medium [5], at 24 "C, under continuous Gro-Lux (Silvana) fluorescent illumination (10000 lux).Plants were harvested a t different growth stages (Fig.1). The organs were excised, frozen and kept a t -30 "C in 20 ml ethanol for several days, which

Uptake and Transient Expression of Chimeric Genes in Seed-Derived Embryos

“…The approach described here is reminiscent of work carried out in the late 1960s and early 1970s, when seeds or seedlings were treated with bacterial or genomic plant DNA. This earlier work proved to be contentious because of the possibility of bacterial contamination, the lack of an effective molecular marker, as well as the lack of reproducibility (Hess, 1969(Hess, , 1970Ledoux and Huart, 1969;Ledoux et al, 1971Ledoux et al, , 1974Soyfer, 1980). A critical evaluation of the results described in this paper prompts several obvious questions that can be summarized as follows: Are the observed signals in embryos true NPT ll-dependent activities resulting from the uptake and expression of DNA in the cells of the plant embryos?…”

“…This general approach has been used previously (Hess, 1969(Hess, , 1970Ledoux and Huart, 1969;Ledoux et al, 1971Ledoux et al, , 1974Soyfer, 1980), but the results of these early studies were discussed controversially in the literature (Kleinhofs et al, 1975;Kleinhofs and Behki, 1977;Soyfer and Titov, 1981). In our study, defined molecular markers were used and their expression was followed over a short time interval.…”

On the question of the integration of exogenous bacterial DNA into plant DNA.

“…Foreign DNA uptake and its integration and replication in plants have been reported for several plant systems by Le-doux and his collaborators (1)(2)(3). Double stranded segments of several types of donor bacterial DNA are said to be covalently joined with recipient plant DNA in an alkali-stable linkage, and then preferentially replicated during subsequent growth of the plants (1)(2)(3) (6).…”

Exogenous DNA Uptake via Cellular Permeabilization and Expression of Foreign Gene in Wheat Zygotic Embryos.

“…Exogenous DNA uptake came into limelight in early 1970s, employing dry tissues such as mature embryos and seeds as the target material and exploited the physico-chemical and structural properties of plasmalemma of the cell (Hess et al,1970;Ledoux et al, 1971). The simplicity of the technique and elimination of skill and labour involved in tissue culture continued to allure scientists to explore its mechanism for the field of plant transformation.…”