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Reto's Buffers & Media Book

信息来源：本站原创　更新时间：2004-7-26 13:56:00
08-15 Buffer

Compound
MW
Final concentration
200 ml
Glycine 75.07 
0.1 M
1.5 g
1 M MgCl2  
 10 mM
2 ml
KOH  
to pH 9
  
H2O     to 200 ml

A

Acetic Acid 1 M

Compound
MW
Final concentration
50 ml
Acetic Acid 98% (~17 M)
 
1 M
3 ml
H2O    
to 50 ml

Acrylamide/Bisacrylamide

Compound
MW
Final concentration
500 ml
Acrylamide
71.08
20 %
100 g
Bisacrylamide
154.17
0.8 %
4 g
H2O    
to 500 ml

Alkaline Lysis Buffer A

Compound
MW
Final concentration
500 ml
Glucose
180
50 mM
4.5 g
Tris
121
25 mM
1.5 g
EDTA 0.25 M  
10 mM
20 ml
H2O    
to 450 ml
Adjust pH 8.0 with HCl
H2O, Autoclave, store at 4°C    
to 500 ml

Alkaline Lysis Solution 1

Compound
MW
Final concentration
100 ml
Lysozyme  
0.2 mg/ml
0.02 g
Alkaline Lysis Buffer A    
to 100 ml

Alkaline Lysis Solution 2

Compound
MW
Final concentration
50 ml
SDS 20 %  
1 %
2.5 ml
NaOH 10 M  
0.2 M
1 ml
H2O    
to 50 ml

Alkaline Lysis Solution 3

Compound
MW
Final concentration
50 ml
KAc
98
3 M
14.7 g
Acetic Acid 100 %  
11.5 %
5.75 ml
H2O    
to 50 ml

Amido Black

Compound
MW
Final concentration
100 ml
Amidoblack
98
0.1 %
0.1 g
Acetic Acid 100 %  
7.5 %
7.5 ml
Methanol  
20 % (v/v)
20 ml
H2O    
to 100 ml

Aminoacid Concentration in Media

Constituent
Final mg/l
Stock
ml stock for 1 liter medium
Adenine sulfate
20
2 mg/ml
10
Uracil
20
2 mg/ml
10
L-tryptophan
20
 2 mg/ml
10
L-histidine-HCl
20
2 mg/ml
10
L-arginine-HCl
20
2 mg/ml
10
L-methionine
20
2 mg/ml
10
L-tyrosine
30
3 mg/ml
10
L-leucine
30
3 mg/ml
10
L-isoleucine
30
3 mg/ml
10
L-lysine-HCl
30
3 mg/ml
10
L-phenylalanine
50
5 mg/ml
10
L-glutamic acid
100
10 mg/ml
10
L-aspartic acid
100
10 mg/ml
10
L-valine
150
15 mg/ml
10
L-threonine
200
20 mg/ml
10
L-serine
400
40 mg/ml
10

3-Aminophtalhydrazide 250 mM

Compound
MW
Final concentration
10 ml
3-Aminophtalhydrazide
177
250 mM
442 mg
H2O    
to 10 ml

Ammoniummolybdate 4.2 %

Compound
MW
Final concentration
50 ml
Ammoniummolybdate x 4H2O
236
4.2 %
2.1 g
H2O    
to 30 ml
HCl fuming 37 % (~12 M)  
4 M
16.7 ml
H2O    
to 50 ml

Ampicillin 20 mg/ml pH 8.0

Compound
MW
Final concentration
15 ml
Ampicillin
349.4
20 mg/ml
300 mg
H2O    
to 10 ml
Adjust pH 8.0 with NaOH
H2O, Filter sterilize    
to 15 ml

Antibiotics

Antibiotics
mg/ml on plates
Action
 Resistance Mechanism
Ampicillin
50
 Inhibits cell wall synthesis by inhibiting formation of the peptidoglycan crosslink b-lactamase (286 aa) is secreted into the periplasm and cleaves lactam antibiotic
Chloramphenicol
20
 Inhibits protein synthesis by interacting with the 50S ribosomal subunit and inhibiting the peptidyltransferase reaction. 216 aa protein cytoplasmic protein which acetylates chloramphenicol
Kanamycin 
Neomycin
30
 Inhibits translocation and elicits miscoding by binding to the 30S ribosomal subunit. 256 aa aminoglycoside phosphotransferase phosphorylates kanamycin, neomycin, etc.
Tetracycline
12
 Inhibits protein synthesis by preventing binding of aminoacyl tRNA to the ribosome A site. 428 aa membrane protein catalyzing efflux of tetracyclin, probably by a cation symport mechanism.

APS

Compound
MW
Final concentration
10 ml
Ammoniumpersulfate
228.2
10 %
1 g
H2O, store at 4°C    
to 10 ml

Ascorbate 0.5 M pH 6.0

Compound
MW
Final concentration
50 ml
L-Ascorbic acid
176.1
0.5 M
4.4 g
H2O    
to 22 ml
Adjust pH 6.0 with NaOH 1 M
H2O    
to 50 ml

Asolectin (fatty acid free) 50 mg/ml

Compound
MW
Final concentration
50 ml
Asolectin (aceton treated)
 
50 mg/ml
2.5 g
Diethylether, store at -20°C    
to 50 ml

ATP 200 mM

Compound
MW
Final concentration
10 ml
Adenosine-5'-Triphosphate
551.1
200 mM
1.102 g
H2O    
to 5 ml
Adjust pH 6.5 with NaOH 1 M
H2O, store at -20°C    
to 10 ml

ATP Regenerating System (ARS) 40x

Compound
MW
Final concentration
1 ml
Phospoenolpyruvate (PEP)
208.0
20 mM
4.16 mg
Pyruvate Kinase (PK) 5 U/ml  
0.1 U/ml
20 ml
H2O, store at 4°C    
to 1 ml

B

BALP Protease Inhibitor Mix 21x

Compound
MW
Final concentration
10 ml
Aprotinin 1.5 mg/ml  
0.486 g/l
3.24 ml
PepstatinA 1 mg/ml in DMSO
686
0.131 g/l
1.3 ml
Leupeptin x 1/2H2SO4 10.5 mM
476
0.04 mM
37.5 ml
4-Aminobenzamidine x 2HCl
208
190 mM
396 mg
H2O    
to 10 ml

Bis-Tris 1M pH7.0

Compound
MW
Final concentration
100 ml
Bis-Tris
209.2
1 M
20.9 g
H2O    
to 50 ml
Adjust pH 7.0 with HCl      
H2O    
to 100 ml

Brij/Doc pH 8.0

Compound
MW
Final concentration
100 ml
EDTA 0.25 M
 
1 mM
0.4 ml
Tris 1 M pH 8.0  
10 mM
1 ml
Na-Deoxycholate 8 %  
0.4 %
5 ml
Brij 58  
1 %
1 g
H2O    
to 90 ml
Dissolve at 37°C, adjust pH 8.0
H2O, Filter sterilize     to 100 ml

BS3 Buffer pH7.5
Compound
MW
Final concentration
1000 ml
HEPES
238.3
100 mM
23.83 g
Dodecylmaltoside
 510.6
0.05 %
0.5 g
EDTA 0.25 M  
1 mM
4 ml
DTT
154
10 mM
1.54 g
H2O    
to 900 ml
Adjust pH 7.5 with NaOH 10M
H2O     to 1000 ml

C

CdSO4 1 M
Compound
MW
Final concentration
1.5 ml
CdSO4
208.5
1 M
313 mg
H2O    
to 1.5 ml

Chloroform

Compound
MW
Final concentration
50 ml
Chloroform  
 
48 ml
Isoamyl alcohol  
4 %
2 ml

CIP Buffer 4x

Compound
MW
Final concentration
15 ml
EDTA 0.25 M pH 8.0
 
200 mM
12 ml
Tris 1 M pH 8.0  
0.4 mM
6 ml
H2O    
3 ml

Citrate 1M pH5.O

Compound
MW
Final concentration
50 ml
Citrate x H2O
210
1 M
10.5 g
H2O    
to 35 ml
Adjust pH 5.0 with NaOH 10M
H2O     to 50 ml

Colony PCR Buffer 10x

Compound
MW
Final concentration
15 ml
Tris 1M pH6.8
 
125 mM
1.875 ml
KCl
74.5
0.5625 M
0.629 g
H2O, Autoclave    
to 15 ml

Coomassie Blue Solution

Compound
MW
Final concentration
1000 ml
Coomassie Blue (Serva Blau)
 
0.2 %
2 g
Acetic acid  
7.5 %
75 ml
Methanol  
50 %
500 ml
H2O    
to 1000 ml

Coumaric Acid 40 mM

Compound
MW
Final concentration
10 ml
p-Coumaric acid
164
40 mM
66 mg
H2O    
to 10 ml

CS/PBS

Compound
MW
Final concentration
200 ml
PBS, 20x
 
1x
10 ml
Newborn calf serum  
10 %
20 ml
H2O, Store frozen     200 ml

CuSO4 1 M
Compound
MW
Final concentration
25 ml
CuSO4
159.6
1 M
4 g
H2O    
to 25 ml

D

DEPC treated H2O
Compound
MW
Final concentration
1000 ml
H2O  
 
1000 ml
Diethylpyrocarbonate (DECP)  
0.1 %
1 ml
Let sit at RT for 1 h, Autoclave  

Destain Solution

Compound
MW
Final concentration
1000 ml
Acetic acid  
7.5 %
75 ml
Ethanol  
10 %
100 ml
H2O    
to 1000 ml

Developer 5x w/o

Compound
MW
Final concentration
2000 ml
Na2CO3  
10 %
200 g
H2O    
to 2000 ml

Dialysis Tubing for DNA

Product/Reagent Volume
Treatment
Visking 20/32 tubing
6 m
Cut 30 pieces of 20 cm
2 % NaHCO3
2 l
Briefly boil tubing
10 mM EDTA
2 l
Briefly boil tubing
10 mM EDTA
200 ml
Store tubing submerged
H2O  
Rinse tubing before use

DNA Loading Buffer 6x

Compound
MW
Final concentration
10 ml
Bromophenolblue
 
0.25 %
25 mg
Xylene Xyanol  
0.25 %
25 mg
Sucrose  
40 %
4 g
H2O    
to 10 ml

Dodecylmaltoside 10 %

Compound
MW
Final concentration
50 ml
Dodecylmaltoside
510.6 
10 %
5 g
H2O, Store at 4°C    
to 50 ml

DTT 1 M

Compound
MW
Final concentration
5 ml
Dithiothreitol
154
1 M
770 mg
H2O, Store at 4°C    
to 10 ml

E

ECL Solution 1

Compound
MW
Final concentration
102 ml
Tris-Cl 200 mM pH 8.5
 
200 mM
100 ml
3-Aminophtalhydrazine 250 mM  
2.5 mM
1 ml
Coumaric acid 40 mM, Filter  
0.4 mM
1 ml

ECL Solution 2

Compound
MW
Final concentration
100 ml
H2O
 
 
100 ml
H2O2 30 %  
 
60 ml

EDTA (Na-Salt) 0.25 M pH8.0

Compound
MW
Final concentration
100 ml
EDTA (Na2 salt)
372
0.25 M
9.3 g
H2O    
to 70 ml
Adjust pH 8.0 with 10 M NaOH 
H2O, Autoclave    
to 100 ml

F

FeCl3 1 M
Compound
MW
Final concentration
1.5 ml
FeCl3 x 6 H2O
270
1 M
406 mg
H2O    
to 1.5 ml

Ferrozine 100 mM

Compound
MW
Final concentration
1 ml
Ferrozine
492.5 
100 mM
49.2 ml
NaOH 1 M    
2 ml
H2O  
 
to 1 ml

Folin 50 %

Compound
MW
Final concentration
50 ml
Folin Reagens
 
50 %
25 ml
H2O  
 
to 50 ml

FPLC Solution A pH 7.4

Compound
MW
Final concentration
500 ml
Tris
121.14
50 mM
3.03 g
Glycerol 87 %  
10 %
57.5 ml
Dodecylmaltoside 10 %  
0.1 %
5 ml
H2O
to 460 ml
Adjust pH 7.4 with 1 M H2SO4
H2O    
to 500 ml

FPLC Solution B pH 7.4

Compound
MW
Final concentration
500 ml
Imidazol
68
200 mM
6.8 g
Tris
121.14
50 mM
3.03 g
Glycerol 87 %  
10 %
57.5 ml
Dodecylmaltoside 10 %  
0.1 %
5 ml
H2O
to 460 ml
Adjust pH 7.4 with 1 M H2SO4
H2O    
to 500 ml

FPLC Solution C pH 7.4

Compound
MW
Final concentration
500 ml
K2SO4
174.3
200 mM
17.4 g
Tris
121.14
50 mM
3.03 g
Glycerol 87 %  
10 %
57.5 ml
Dodecylmaltoside 10 %  
0.1 %
5 ml
H2O
to 460 ml
Adjust pH 7.4 with 1 M H2SO4
H2O    
to 500 ml

French Press Buffer (FPB)

Compound
MW
Final concentration
500 ml
Tris-SO4 1 M pH 7.4
 
50 mM
25 ml
Glycerol 87 %  
10 %
57.5 ml
H2O, Autoclave    
to 500 ml

G

Glutathione 100 mM

Compound
MW
Final concentration
1.5 ml
Glutathione (GSH)
307.4
 100 mM
46 mg
H2O, Store at 4°C    
to 1.5 ml

Glycerol 50 %

Compound
MW
Final concentration
50 ml
Glycerol 87 %  
50 %
28.75 ml
H2O, Autoclave    
to 50 ml

H

HCl 1 M

Compound
MW
Final concentration
50 ml
HCl 37 % (~12M)
 
 1 M
4.2 ml
H2O    
to 50 ml

HIS-Stock 2 mg/ml

Compound
MW
Final concentration
100 ml
L-Histidine
155
 2 mg/ml
200 mg
H2O, Autoclave    
to 100 ml

I

IPTG 1M

Compound
MW
Final concentration
5 ml
Isopropyl-b,D-thiogalactoside
238.3
1 M
1.19 g
H2O, Filter sterilize    
to 5 ml

J

K

Kanamycin 10 mg/ml

Compound
MW
Final concentration
15 ml
Kanamycin 582.6
10 mg/ml
150 mg
H2O, Filter sterilize  
 
to 15 ml

KH2PO4 0.4 M
Compound
MW
Final concentration
50 ml
KH2PO4 136.1
0.4 M
2722 mg
H2O  
 
to 50 ml

K-Na-Tartrate 2 %

Compound
MW
Final concentration
50 ml
K-Na-tartrate x 4 H2O
 
 2 %
1 g
H2O    
to 50 ml

L

Laemmli Electrophoresis Buffer

Compound
MW
Final concentration
10 liters
Tris
121.14
25 mM
30 g
Glycine
75
192 mM
144 g
EDTA (Triplex II)
292
1 mM
2.9 g
SDS
288.4
3.5 mM
10  g
H2O  
 
to 10 liters

LB Media

Compound
MW
Final concentration
1000 ml
Yeast Extract (BBL)  
0.5 %
5 g
Trypticase Peptone (BBL)  
1 %
10 g
NaCl  
1% (0.2 M)
10 g
H2O, Autoclave  
 
to 1000 ml

LB Plates

Compound
MW
Final concentration
1000 ml
Yeast Extract (BBL)  
0.5 %
5 g
Trypitcase Peptone (BBL)  
1 %
10 g
NaCl  
0.2 M
10 g
Agar   
1.5 %
15 g
H2O, Autoclave  
 
to 1000 ml

LB Low Salt Liquid

Compound
MW
Final concentration
1000 ml
Bacto Yeast Extract  
0.5 %
5 g
Bacto Peptone  
1 %
10 g
NaCl  
0.1 M
5 g
H2O, Autoclave  
 
to 1000 ml

LB Low Salt Plates

Compound
MW
Final concentration
1000 ml
Bacto Yeast Extract  
0.5 %
5 g
Bacto Peptone  
1 %
10 g
NaCl  
0.1 M
5 g
Agar   
1.5 %
15 g
H2O, Autoclave  
 
to 1000 ml

LEU-Stock 3 mg/ml

Compound
MW
Final concentration
50 ml
L-Leucine  
3 mg/ml
150 mg
H2O, Filter sterilize  
 
to 50 ml

LiAc 1 M

Compound
MW
Final concentration
100 ml
LiAc-dihydrate
102
1 M
10.2 g
H2O    
to 60 ml
TE 10x    
10 ml
H2O, Autoclave    
to 100 ml

LiCl 1 M

Compound
MW
Final concentration
50 ml
LiCl
42.4
1 M
2.12 g
H2O, Autoclave    
to 50 ml

Lowry Component

Compound
MW
Final concentration
200 ml
Na2CO3
106
3 %
6 g
NaOH 40
0.1 M
800 mg
H2O  
 
to 200 ml

LYS-Stock 3 mg/ml

Compound
MW
Final concentration
50 ml
DL-Lysine
182.6
3 mg/ml
150 mg
H2O, Autoclave  
 
to 50 ml

M

M17 Media

Compound
MW
Final concentration
1000 ml
Neopeptone Difco
 
0.5 %
5 g
Bacto-Soytone  
0.5 %
5 g
Beef Extract  
0.5 %
5 g
Yeast Extract  
0.25 %
2.5 g
Ascorbic Acid
176
2.8 mM
0.5 g
2-Glycerolphosphate x 5 H2O, disodium salt
Note: we used 19 g/l up to Dec. 2002
306
0.1%
1 g
H2O, Autoclave, Chill to RT    
to 1000 ml
Glucose 50 %
 
1 %
20 ml
MgSO4 1 M  
1 mM
1 ml

Malachit Green 1.25 mM

Compound
MW
Final concentration
50 ml
Malachit Green
482.6
1.25 mM
30 mg
H2O    
to 50 ml

MES 100 mM pH 6.4

Compound
MW
Final concentration
250 ml
MES
195.2
100 mM
4.88 g
H2O    
to 200 ml
Adjust pH 6.4 with NaOH 
H2O, Autoclave    
to 250 ml

MgSO4 1 M
Compound
MW
Final concentration
15 ml
MgSO4
246.5
1 M
3.70 g
H2O    
to 15 ml
MRS

Compound MW Final concentration 1000ml
Trypticase Peptone (BBL) 1 % 10 g
Beef Extract (Difco) 0.3 % 3 g
Yeast Extract 0.5 % 5g
K2HPO4 174 11.5 mM 2 g
NH4Acetate 77 39 mM 3 g
tri-Na-Citrate  x 2H2O 294 10.2 mM 3 g
MgSO4 1 M 246 0.8 mM 0.8 ml
MnSO4 0.4 M 0.4 mM 1 ml
Tween 80  0.1 % 1 ml
Adjust pH 6.3 with 5N HCl
H2O, Autoclave to 1000 ml
Glucose 50%
(or Lactose = MRSL)		 1 % 20 ml
N

Na-Acetate 2.5 M pH 5.2

Compound
MW
Final concentration
50 ml
NaAc x 3H2O
136
2.5 M
17 g
H2O    
to 35 ml
Adjust pH 5.2 with Acetic Acid 
ca. 10 ml
H2O, Autoclave    
to 50 ml

Na-Citrate 34 %

Compound
MW
Final concentration
50 ml
Na-citrate x 2H2O
294
34 %
17 g
H2O    
to 50 ml

NaCl 5 M

Compound
MW
Final concentration
50 ml
NaCl
58.5
5 M
14.6 g
H2O    
to 50 ml

Na-Deoxycholate 8%