Uptake and incorporation of labeled tryptophan isomers into IAA in the jsR1 mutant of Lemna gibba

Analyses of the IAA-overproducing mutant of Lemna have been initiated in order to study in vivo biosynthesis of IAA. Using radiolabelled tryptophan isomers prepared from commercial sources of 14C-D,L tryptophan by chiral separation kinetics of uptake of L and D tryptophan were determined for sterile cultures of individual jsR1 four-frond colonies. Over a 24 h period, about 50% of the radioactivity from 14C-L-TRP in media, or about 25% from 14C-D-TRP, was found in the plant tissue. Maximal rates of uptake were seen in the first six hors for both isomers. Endogenous levels of tryptophan determined in jsR1 as measures of pool sizes in vivo show 5 to 10 ug/g FW total tryptophan with less than 1% in the D isomer form. Information on uptake and endogenous pool sizes of tryptophan isomers is being used for feeding of stable isotope labeled tryptophan (13C, 14N) to jsR1 at physiological levels. Analyses of incorporation of label into IAA using GC-MS and high resolution mass spectrometry are currently underway