Seeds, cotyledons, and leaves of the Indian Brassica juncea line CPI 81793 all contained (2-hydroxybutenyl) -, [ (4-hydroxyindolyl) methyl] -, pen ten yl-, (indolylmethyl) -, [ (4-methoxyindolyl) methyl] -, (phenylethy1)-, butenyl-, and allylglucosinolates. Butenyl-and allylglucosinolates predominated in all tissues and were also detected in callus tissue and regenerated plants derived from cotyledon cultures.The glucosinolate content of intact 7-day-old cotyledons was similar to that of whole seeds, but thereafter seedling cotyledon glucosinolates declined gradually to approximately 10% of the seed level at 21 days, just before abscission. A rapid decline in glucosinolate content occurred when 7-day-old cotyledons were excised and cultured in vitro, reaching approximately 2% of the seed level after 13 days in culture. Prolific root and shoot regeneration occurred in these cultures, but glucosinolate levels remained very low in all nontransferred cultures. Leaf glucosinolate content increased markedly when regenerant plants were transferred to fresh medium. After 31 days in fresh medium, the leaf glucosinolate content of regenerant plants was comparable to that of mature leaves from 8-week-old seedlings.Agronomic characteristics such as reduced pod-shattering, good tolerance of heat and drought, low erucic acid oil, and high-protein seed meal render modern varieties of Indian mustard potentially valuable alternatives to rapeseed (Brassica napus, Brassica campestrk) in warm and dry climates; however, the usefulness of this crop is still severely limited by the unacceptably high content of goitrogenic glucosinolates in the seed meal (Kirk and Oram, 1978, 1981).Conventional breeding and selection of low-glucosinolate Brassica sp. would be greatly facilitated by a better understanding of the physiology and biochemistry of glucosinolates and the development of improved screening techniques. The recent premature announcement of "glucosinolate-free" lines of Brassica juncea by Cohen et al. (1983) highlights the difficulties that plant breeders face in recognizing heritable differences in glucosinolate metabolism; these workers subsequently found the glucosinolate-free character to be unstable in generations grown after 1983 (W. Thies, personal communication).
Enzyme-linked immunosorbent assays (ELISA's) were developed for the determination of alkenyl glucosinolates (sinigrin, gluconapin) in crude extracts of seed meal and vegetative tissues of Indian mustard (Brassica juncea). The assays were constructed from two types of high-titer anti-sinigrin polyclonal antibodies derived from antisera raised against a conjugate of sinigrin hemisuccinate with bovine serum albumin. An ovalbumin-sinigrin hemisuccinate conjugate was used as the ELISA plate-coating antigen. In cross-reactivity studies, antisera showed high specificities toward the alkenyl side chain and the thiohydroximate moiety of the glucosinolate molecule and did not bind to desulfoglucosinolates. All antisera showed 100% cross-reactivity with gluconapin, and one type cross-reacted completely with progoitrin, the principal glucosinolate of rapeseed (Brassica napus). The most sensitive ELISA had a linear inhibition curve over 1 X mol of sinigrin, with 50% inhibition at 2.8 X lo-" mol per assay. Parallel analyses of mustard seed and cotyledon extracts using both an ELISA and a standard HPLC method gave values identical within the error limits of the two procedures. The ELISA is, however, more sensitive, less expensive, and much faster than any other glucosinolate assay of comparable specificity.to 1 X
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