Stable overexpression and targeted gene deletion of the causative agent of ash dieback Hymenoscyphus fraxineus

Table 1 Media and buffers used in this study

Denomination	Description	Components
AM_L	Liquid ash medium	50 g ash leaves incl. petioles in ddH₂O^a
AM_S	Solid ash medium	AM_L 1.8% (w/v) microagar^a,b
AM_RegL pH 5.8	Liquid ash medium for regeneration	AM_L 500 mM sucrose
AM_RegS pH 5.8	Solid ash medium for regeneration	AM_RegL 1.8% (w/v) microagar^b
AM_HygS	Solid ash medium with hygromycin	AM_S 100 µg/ml hygromycin B^b
AM_GenS	Solid ash medium with geneticin	AM_S 375 µg/ml geneticin disulphate (G418) solution^c
Water agar		ddH₂O 1.8% (w/v) microagar^b
MgSO₄-buffer pH 3.5/5.0/5.8	Cell wall digestion	1 M MgSO₄ 50 mM tri-sodium citrate
STC pH 8.0	Collection of floating protoplasts	500 mM sucrose 10 mM Tris–HCl 50 mM CaCl₂
PEG/STC pH 8.0	Transformation	STC 40% (w/v) PEG 4000
CTAB-buffer pH 8.0	gDNA isolation	2% (w/v) CTAB^d 100 mM Tris–HCl 20 mM EDTA 1.4 M NaCl
TE-buffer/RNase pH 8.0	gDNA isolation	10 mM Tris–HCl 1 mM EDTA 1 mg/ml RNaseA

ISSN: 2054-3085