8), with PP2C activity alone (OA and EGTA
inhibition) being 8.
In this experiment we concluded that: (a) 5 days after chelate application was the most suitable time for harvest of a Cd-laden biomass, (b) EGTA
was effective in enhancing shoot uptake of Cd by wheat when grown on a Cd-contaminated soil, and (c) 8 weeks after emergence was the best growth stage for wheat to attain maximum Cd translocation to the shoots.
5 mM MgCl2, 50 mM pyrophosphate, 1 mM sodium orthovanadate, 1 mM EGTA
, 100 mM sodium fluoride, 1% Triton X-100, 10% glycerol, 10 [micro]g/mL leupeptin, 10 [micro]g/mL aprotinin, and 1 mM phenylmethylsulfonyl fluoride.
4], 4mM EGTA
and 1%Triton X-100), and luciferase activity was determined using a luciferase assay kit.
2), 25 mM [beta]-glycerophosphate, 5 mM EGTA
, 1 mM [Na.
Enzymes, molecular biology reagents, reagent sets, and chemical reagents were obtained from the following sources: Tris-HCl and CHAPS (Sigma); MgCl,, EGTA
, and glycerol (Fisher Scientific); phenylmethylsulfonyl fluoride and 4-(2-aminoethyl)benzenesulfonyl fluoride (Calbiochem); RNA Secure RNase inhibitor (Promega); TeloTAGGG Kit (Roche Molecular Biochemicals); PCR buffer and Taq HotStar polymerase (Qiagen); TRIzol LS Reagent and 2'-deoxynucleoside 5'-triphosphates (dNTPs; Invitrogen); chloroform (Mallinckrodt).
2 mM EGTA
and washed three times with this solution at 20 min intervals.
4, 10 mM EDTA, 5 mM EGTA
, 100 mM NaF, 200 mM sucrose, 100 [micro]M Na-orthovanadate, 5 mM pyrophosphate, 4 [micro]g/mL leupeptin, 4 [micro]g/mL soybean trypsin inhibitor, 1 mM benzamidine, 20 [micro]M calpain inhibitor 1, 100 mU/mL aprotinin, and 100 [micro]M phenylmethylsulfonyl-fluoride).
One study has suggested that EGTA
is preferable to EDTA as an anticoagulant to prevent the irreversible loss of antigen-specific lymphoproliferative responses in instances in which unclotted blood may be stored for long periods (14).
6 buffer consisting of 20 mM Tris-HCl, 100 mM KCl, 5 mM NaCl, 2 mM EDTA, 1 mM EGTA
, 2 mM dithiothreitol and 2 mM PMSF.
For preparation of washed platelets, blood was collected in the citrate buffer described above, supplemented with 15 mmol/L EGTA
The cells were then washed with PBS, and 400 [micro]l of fixative containing rhodamine phalloidin (20 mM KP04, 10 mM Pipes, 5mM EGTA
, 2 [micro]M MgCl2, 0.