Research Article

Conformational-Dependent and Independent RNA Binding to the Fragile X Mental Retardation Protein

Table 1

Binding conditions used to measure the interaction between FMRP and RNA.

PublicationAssayConditions

Ashley et al. (1993) [51]Pull-down16 mM HEPES-KOH pH 7.9, 120 mM KCI, 0.04% Nonidet P-40, 1 mg/mL BSA, 0.16 mM dithioerythritol, 0.4 mM phenylmethylsulfonyl fluoride

Brown et al. (1998) [49]Pull-down10 mM Tris-HCl pH 7.5, 2.5 mM MgCl2, 100 mM NaCl, 2.5% Trition X100, 1 mg/mL heparin

Price et al. (1996) [52]Pull-down20 mM Hepes, pH 7.9, 2 mM MgCl2, 10 mM ZnCl2, 70 mM NH4Cl, 0.02% Nonidet P-40, 5 mg/mL yeast tRNA

Sung et al. (2000) [21]Pull-down Filter-Binding20 mM Hepes, pH 7.9, 2 mM MgCl2, 10 mM ZnCl2, 70 mM NH4Cl, 0.02% Nonidet P-40, 5 mg/mL yeast tRNA

Denman and Sung (2002) [57]Pull-down20 mM Hepes, pH 7.9, 2 mM MgCl2, 10 mM ZnCl2, 70 mM NH4Cl, 0.02% Nonidet P-40, 5 mg/mL yeast tRNA

Schaeffer et al. (2001) [26]EMSA50 mM Tris-HCl pH 7.4, 1 mM MgCl2, 1 mM EDTA, 150 mM KCl, 1 mM DTT, 0.25 mg/mL of E.coli tRNA, 0.01% BSA, 8 U of RNasin

Sung et al. (2003) [17]Pull-down
EMSA
50 mM Tris-HCl, pH 7.0, 2 mM MgCl2, 150 mM NaCl, 1 mM DTT, 0.25 mg/mL E.coli tRNA, 0.25 mg/mL BSA

Bechara et al. (2006) [61]EMSA50 mM Tris-HCl pH 7.4, 1 mM MgCl2, 1 mM EDTA, 150 mM KCl, 1 mM DTT, 0.25 mg/mL of E.coli tRNA, 0.01% BSA, 8 U of RNasin

Didiot et al. (2008) [54]EMSA50 mM Tris-HCl pH 7.4, 1 mM MgCl2, 1 mM EDTA, 150 mM KCl, 1 mM DTT, 0.25 mg/mL of E. coli tRNA, 0.01% BSA, 8 U of RNasin

Zalfa et al. (2003) [36]EMSA10 mM HEPES pH 7.9, 3 mM MgCl2, 10 mM DTT, 100 mM KCl, 750 mM NaCl, 5% glycerol, 7 mM β-Mercaptoethanol, 1 mg/mL Albumin, 1.3 mg/mL Heparin

Zalfa et al. (2005) [40]EMSA20 mM HEPES-KOH, pH 7.6, 5 mM MgCl2, 300 mM KCl, 2 mM DTT, 5% glycerol, and 500 ng of total yeast tRNA or 20 μg of heparin.

Darnell et al. (2001) [22]Filter-Binding10 mM Tris-OAc pH 7.7, 200 mM KOAc, 5 mM MgOAc2

Darnell et al. (2005) [30]Filter-Binding50 mM Tris-OAc at pH 7.7, 50 mM KOAc, 10 mM DTT, 5 mM Mg(OAc)2, 30 μg/mL tRNA

Gabus et al. (2004) [42]EMSA20 mM Tris-HCl pH 7.5, 30 mM NaCl, 0.2 mM MgCl2, 5 mM DTT, 10 μM ZnCl2

Laggerbauer et al. (2001) [16]Pull-downPBS, 0.02% IGEPAL, 1% BSA

Siomi et al. (1993) [15]Pulldown10 mM Tris-HCl pH 7.4, 2.5 mM MgCl2, 0.5% Triton X-100, 100–1000 mM NaCl

Stetler et al. (2005) [56]Pulldown2 M KOAc, 100 mM Tris-OAc pH 7.7 and 50 mM MgOAc2, 1 μL of yeast tRNA, 1 μL of RNAsin

Menon and Mihailescu (2007) [62]EMSA50 mM Tris-HCl pH 7.5, 150 mM NaCl and protease inhibitors

Fahling et al. (2009) [20]EMSA10 mM Hepes pH 7.2, 3 mM MgCl2, 5% glycerol, 1 mM DTT, 150 mM KCl, 2 U/μL
RNaseOUT, 0.5 μg/μL rabbit rRNA

Zou et al. (2008) [63]AGESA20 mM Tris-HCl pH 7.2, 150 mM NaCl

Iacoangeli et al. (2008) [47]EMSA50 mM Tris-HCl pH 7.6, 150 mM KCl, 1 mM MgCl2, 1 mM EDTA, 1 mM DTT, 0.2 U/μL
RNase inhibitor, 100 ng/μL total yeast tRNA, and 100 ng/μL BSA

Sets of binding conditions are grouped by the different laboratories that used them. Each set of conditions is differentiated from the next by a dotted line. In some instances the same group used multiple sets of binding conditions in multiple publications.