Jurgen Logemann

Improved method for the isolation of RNA from plant tissues

A fast and efficient method for the isolation of RNA from plant tissues is described. Tuber tissue is homogenized in a guanidine hydrochloride-containing buffer followed by direct extraction with phenol/chloroform. The RNA is precipitated from the aqueous phase, washed with 3 M sodium acetate and 70% ethanol, and finally dissolved in water. The yield of RNA is up to 500 micrograms/g of tissue and several tests indicate intact and nondegraded RNA. This method can be adapted to a small-scale version by the use of 1.5-ml tubes, allowing rapid isolation of RNA from a larger number of samples. Finally, this method is of particular use for isolating RNA from tissues with a high polysaccharide and nuclease content such as wounded potato tubers.

Nucleotide sequence and regulated expression of a wound-inducible potato gene (wun1)

SummaryMechanical wounding of potato leaves, stems, roots and tubers leads to a rapid increase of wun1 mRNA. In potato leaves, the wound-induced accumulation of wun1 mRNA is inhibited by the addition of sucrose or other osmotically active agents. This inhibition is organ specific since sucrose does not prevent wun1 mRNA accumulation in wounded tubers. In contrast, expression of patatin was shown to be repressed in tubers by wounding and this repression was reversed by increasing osmotic pressure. Sequence data obtained from the analysis of a wun1 cDNA and a wun1 genomic clone show no homology to any gene known so far. Histochemical data demonstrate a striking analogy in cell specific expression of chimeric genes expressed under the control of the wun1 promoter and the cell specific production of callose in wounded tobacco leaves.

Expression of a Ribosome Inhibiting Protein (RIP) or a Bacterial Chitinase Leads to Fungal Resistance in Transgenic Plants

In order to protect plants against fungal attack, two genes (RIP, ChiA) encoding proteins with in vitro antifungal activity were expressed in transgenic tobacco plants. 1. A barley derived cDNA clone (RIP) encoding a ribosome inhibiting protein. RIP inhibits protein synthesis in fungi by specific RNA N-glycosidase modification of the 28S RNA. 2. A chitinase gene (ChiA) derived from the bacterium Serratia marcescens with the ability to destroy hyphal tips of growing fungi.