NEWLY SYNTHESIZED MESSENGER-RNA AS A PROBE FOR IDENTIFICATION OF WOUND RESPONSIVE GENES FROM POTATOES

A method for the selective isolation of wound induced RNA from potato tuber has been standardised by labeling with 4-thiouridine and subsequent affinity chromatography on Affigel 501 phenyl-mercury Sepharose column. About 30% of the total RNA isolated from wounded potato tissue was found to be synthesized de novo. The relative abundance of wound induced RNAs was confirmed by using two well identified wound inducible gene probes of hydroxyproline-rich glycoproteins (HRGP) and proteinase inhibitor I. The selectively separated wound induced mRNA was used to make cDNA probes. By screening a boolean AND gt10 cDNA library from wounded potato tissue with these cDNA probes, several wound responsive genes were identified. Wound inducible expression of these genes was further confirmed by Northern hybridization and their tissue specific expressions were examined. Thus, a simple method for the direct identification of wound responsive genes from potato was developed.