Table 3.

Chromatographic steps for PrSAI1 purification from growing subterranean shoots of Phelipanche ramosa parasitizing tomato plants.

	Buffer	Flow rate (mL/min)	Gradient	Collected fractions (mL)
Extraction	A: 100 mm Tris‐HCl (pH 7.0) containing 5 mm EDTA, 10 mm ascorbic acid, 1 mmβ‐mercaptoethanol, 0.1 mm PMSF, 1 mm benzamidine	—	—	—
Concanavalin‐A Sepharose	B: 10 mm sodium phosphate (pH 6.5) containing 1 mm EDTA, 10 mm ascorbic acid, 1 mm DTT, 0.1 mm PMSF, 1 mm benzamidine, 1 mm CaCl2, 1 mm MgCl2, 1 mm MnCl2 and 0.5 mm NaCl	3	0–25 mβ‐methyl‐mannoside (60 mL)	3
High Q anion exchanger	C: 50 mm Tris‐HCl (pH 8.0) containing 1 mm DTT, 0.1 mm PMSF and 1 mm benzamidine	0.5	0–0.5 m NaCl (50 mL)	2
HR Sephacryl	D: 10 mm sodium phosphate (pH 6.5) containing 5 mm EDTA, 0.1 mm PMSF, 1 mm benzamidine, 0.05% (w/v) NaN3	0.5	—	1
Liquid isoelectric focusing	50‐mL solution containing an aliquot of HR‐Sephacryl fraction, 2.5% (v/v) Bio‐Lyte® pH 3/10 ampholyte or Bio‐Lyte® pH 3/5 ampholyte (Bio‐Rad, France)	—	—	2.5

DTT, dithiothreitol; EDTA, ethylenediaminetetraacetic acid; PMSF, phenylmethylsulphonyl.