Lipid peroxidation analysis
Lipid peroxide was measured using the protocol mentioned in Chen and
Zhang (2016). Briefly, 50 mg of crushed plant tissues were placed in a
microcentrifuge tube and 1 ml phosphate buffer (PBS) (100mM, pH 7.8) was
added. After vortexing briefly, tubes were centrifuged for 20 mins at
10,000 rpm. Then 100 µl of supernatant was taken in a fresh tube and 1ml
of 0.25% Thiobarbituric acid (TBA) in Trichloroacetic acid (TCA) was
added to it. PBS was used as a control. The samples were heated on a
heating block for 15 mins at 90oC. Holes were poked in
the centrifuge tube lids to avoid popping. The samples were then cooled
on ice for 5 mins and the absorbance readings were taken using a UV-Vis
spectrophotometer (Biotech Epoch microplate reader, Aligent Technologies
Inc., Santa Clara, CA) at 532nm and 600nm. The final Malondialdehyde
(MDA) concentration was calculated based on Lambert−Beer’s equation
(extinction coefficient of MDA is 155 mM/cm).