Identification, cDNA Cloning, and Analysis of mRNAs Having Altered Expression in Tips of Harvested Asparagus Spears

Abstract

Changes in mRNA activity in tips of harvested asparagus spears (Asparagus officinalis L.) held in light or dark for up to 48 h at 20°C were investigated as an initial step in elucidating the genetic response of asparagus spears to harvest. Total RNA was isolated from 30-mm tips of spears 180 mm in length at 0, 6, 12, 24, and 48 h after spear harvest and translated in vitro, and translation products were separated using both sodium dodecyl sulfate-polyacrylamide gel electrophoresis and two-dimensional polyacrylamide gel electrophoresis. We detected 25 consistent changes in translatable mRNAs, involving both increase and decrease in mRNA abundance. The majority of the changes occurred within 12 h of harvest. Most of the changes were not light regulated. cDNA libraries were constructed from polyadenylated mRNA extracted from tips of spears at harvest (0 h) and after 12 h in the dark at 20°C. Differential hybridization screening of the cDNA libraries isolated nine cDNA clones whose corresponding transcripts had altered expression after harvest. Investigations of mRNA activity during spear development demonstrated that the changes detected were harvest related. Possible roles for the mRNAs corresponding to the isolated clones in tips of harvested spears are discussed.