This video demonstrates the seedling growth inhibition assay in the Arabidopsis thaliana plant. The impact of increased concentration of immune peptide elicitor on seedling growth is determined.
Protocol
1. Seedling Growth Inhibition Assay Sterilize seeds and sow on Murashige and Skoog (MS) plates. Prepare sterile half-strength (0.5x) MS medium (2.16 g/L) containing 0.8% agar [w/v]. Pour media into plates (90 x 15 mm) in a laminar flow hood. Sterilize approximately 100 seeds in a microcentrifuge tube by washing with 1 mL of 70% ethanol for 2 min and remove by aspiration. Add 1 mL of 40% bleach and gently rock for 17 mins at room temperature. Remove bleach by aspiration …
Representative Results
Figure 1. Elicitor-induced seedling growth inhibition assay in Arabidopsis. (A) Sow seedlings on MS agar and grow for 3-4 days under standard short-day conditions. (B) Transplant seedlings to 48-well plates containing MS medium or MS containing different concentrations of elf18. (C) After 8-12 days, visually assess the seedling size and then measure fresh weight using an analytical scale…
Declarações
The authors have nothing to disclose.
Materials
8-Well Sterile Plates with Lid
Sigma-Aldrich
CLS3548
Analytical Scale with Draft Sheid
VWR
VWR-225AC
Any standard analytical scale can be used for growth inibition assays, however, a direct computer output is optimal.
BioHit mLine Mechanical 12 Multichannel Pipette (30-300 uL
Sartorius
725240
Any multichannel pipette can be used, as can a single pipetter if necessary.
elf18 (AcSKEKFERTKPHVNVGTIG)
EZ Biolab
cp7211
Store 10 mM stock peptide at -80 deg C in low protein binding tubes. When thawed, store 100 uM working stock at -20 deg C.