Common Solutions - Protein transport and secretion

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Protocols > Biochemical Protocols >

Common Solutions

MOPS NUPAGE RUNNING BUFFER: 104,6g MOPS, 60,6 Tris

Base, 10g SDS, 3 g EDTA in 500ml final(50mM MOPS,50mM
Tris,0,1%SDS,1mM EDTA) pH 7.7 do not use acid or base to
adjust the pH.
BLU DI COOMASSIE 125 ml di isopropanol, 30 ml acetic acid,
325 ml H20, 1,25 gr Brillant Blue. Dissolve O.N. Stirring and
finally filter on paper.
DESTAIN / FIXING SOLUTION 10% acetic acid, 10% methanol
EDTA 0.5M pH 8.0: dissolve186.1 g EDTA disodic salt in H20.
Stirr and add NaOH till pH=8.5 add water till11. Pay attention:
EDTA will dissolve only if pH= 8
NaCl 5M: dissolve 292.2 g NaCl in H20 in 1L final water. Stock
RT
LOADING BUFFER: 5 ml SAMPLE BUFFER 4X, 4 ml SDS 20%,
BpB powder
PBS 10X: prepare following solution: NaH 2P04 1M pH 4.0
(137,99 g/mol); Na 2HP04 1M pH 9.0 (177,99 g/molhard to
dissolve) mix solution starting from pH 9 solution till pH 7.3.
From buffer pH 7.3 take 50ml and add 45g NaCl and H20 to
500ml final.
SAMPLE BUFFER 4X 5,25 ml UPPER UFFER + 20 ml glycerol
100%
SDS (20% w/v): Dissolve 20 g SDS in 100 ml H20 at 60C.
STN (washing beads in pulse chase experiment) 10mM
TRISHC1 pH:7.0, 0.25% NP40, 150mM NaCl
H20 lavaggi (last wash for beads in pulse chase experiment)
5mM TRISHCl pH:7,0
STRIPPING SOLUTION 62,5mM TrisHCl pH=6,7, add
betamercaptoethanol (100 mM final); NaOH
TAE 50X (pH 8 ):Trisbase 242 g, EDTA pH 8 100ml 0.5 M,
Acido acetico glaciale 57.1 ml up to 1L with water.
TAMPONE DI CORSA TRISGLICINA 10X: dissolve 30g di tris
a di 144g glicina in 11 di H2O . Add final 0.1% SDS before use.
(4 litri: 120 g Tris; 576 g glicina)
NP40 Lysis Buffer: 150mM NaCI, 10mM TRIS (pH. 7. 5), 1%

https://sites.google.com/site/proteintransportandsecretion/protocols-2/biochemical-protocols/common-solutions[31/03/2015 13:42:20]

Common Solutions - Protein transport and secretion

NP40
TRANSFER BUFFER 25 mM Trisbase, 192 mMglicina,20%
metanolo Sciogliere 3,03 gr di Tris a 14,4 gr di glicina in circa
750 ml di H20 portare a pH 8,3; aggiungere 200 ml di metanolo
a portare a volume finale di 1 1
TRISGLICINA (10X) per RUNNING BUFFER SDSPAGE 144g
GLICINA, 30g TRIS BASE bring to 1L con H2O or 576g GLICINA,
120g TRIS BASE bring to 4L with H2O
TRISHC1 pH 6.8 (0.5M): Dissolve 6 g Tris in 40 ml H2O,
adjust pH with 48 ml HCl 1Mto 100 ml final with H2O.
TRISHCI IM pH 7.5: dissolve 121.1 g TRIS base in H2O.
Adjust pH with HCI (65ml pH 7.5) bring to 1L.
TRISHCL pH 8.8 (3M): Dissolver 36.3 g tris in 40 ml di H2O,
adjust pHwith 48 ml HCI 1Mbring to100 mlwith H2O
UPPER GEL BUFFER 4X 0, 5 M TrisHCI, 0, 4 % SDS, pH 6, 8.
Add30,3g Trisbase, 10 ml SDS 20% pH 6.8 with HCI add
H2Oto 500 ml final
LOWER GEL BUFFER 4X 1,5 M TrsHC1, 0,4 % SDS, a pH 8,8.
Add90,9g Trisbase, 10 ml SDS 20%pH 8.8with HCIadd H2O
to 500 ml final

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https://sites.google.com/site/proteintransportandsecretion/protocols-2/biochemical-protocols/common-solutions[31/03/2015 13:42:20]