Enhancement of broccoli’s chemo-preventive agent sulforaphane production using plant tissue culture techniques

Abstract

Broccoli (Brassica oleracea var. italica) is one of the most important cruciferous vegetables that belongs to the family Brassicaceae. Broccoli has high content of glucosinolates which have positive potential in cancer treatment. The present investigation was carried out to study the effects of different treatments of abiotic and biotic elicitors on the production and accumulation of sulforaphane and the expression of myrosinase gene from broccoli plant which triggers the synthesis of sulforaphane. In addition, the cytotoxic effect of various sulforaphane treatments on the breast cancer cell line MDA-MB-231 was investigated. Moreover, the transcription levels of the apoptosis marker genes: Bcl-2, Bax, Caspase-3, Caspase-8 and Caspase-9 genes were the main objective of this work. To study these effects, the following experiments were carried out: • Germination of broccoli seeds under aseptic conditions to produce sterile seedlings and the establishment of hairy root cultures from broccoli seedlings using leaf explants. • Treatment of 7-days old seedlings and hairy root culture with different concentrations of the elicitors: zinc sulfate, copper sulfate, salicylic acid and methyl jasmonate. • Extraction of sulforaphane from control and treated cultures, determination of sulforaphane by LC-MS. • Determination of the myrosinase genes expression in control seedlings and hairy root cultures and elicitors treated seedlings and hairy root cultures using real-time PCR. • Selection of the highest sulforaphane producing cell lines.