Common Buffers
10X TE Buffer
100mM Tris-Cl, pH 7.4 or 7.6
10mM EDTA, disodium salt, dehydrate, MW 372.24, pH 8.0
50 X TAE
For 1 Liter:
700 ml dH2O
242 g Tris base, MW 121.14
18.6 g EDTA, disodium salt, dehydrate, MW 372.24
-Mix until EDTA goes into solution
-Add 57.1 ml Glacial Acetic Acid
-Mix and bring volume up to 1 liter (1x= 0.04M Tris-acetate 1mM EDTA)
50X TAE (low EDTA)
For 1 liter:
242 g Tris base, MW 121.14
3.72 g EDTA, disodium salt, dehydrate, MW 372.24
-Add 700 mL dH2O.
-Mix until reagents go into solution
THEN ADD
57.1 mL of Glacial Acetic Acid
-Mix and bring volume up to 1 liter (1x= 0.04M Tris-acetate 0.2mM EDTA)
20X TBE
For 1 liter:
216g Tris base, MW 121.14
14.9g of EDTA, disodium salt, dehydrate, MW 372.24
-Mix Tris base and EDTA with 500ml ddH2O until dissolved
-Add 110g Boric Acid
-Bring up to 1 liter with ddH2O
-Filter sterilize and then autoclave
SM
For 1 liter:
ingredient: amount: [final]
Tris-HCl pH 7.5: 50 ml of 1 M: 50 mM
MgSO4 7-H2O: 2 grams (or 1 gram w/anhydrous): 5 mM
NaCl: 5.8 grams: 0.2 M
gelatin: 1 gram: 0.1%
-Autoclave (gelatin will not go into solution until heated), aliquot into 50 ml polypro. tubes, reautoclave
20X SSPE
For 500ml:
3.6 M NaCl: 105.2 g
0.2M PO4 buffer: 100 mls 1 M
20mM EDTA: 20 mls of 0.5 M
-Autoclave and (optional) DEPC treat
20X SSC
For 1 liter:
NaCl: 175.3g: 3M
Na citrate: 88.2g: 0.15M
H2O: 800ml
-Add a few drops of HCl and pH at 7.0
-Autoclave
IPTG
+Final concentration in plates, 0.5mM, either use 25ul per ml of cell or spread on X-Gal plates.
+When adding to plates, per liter of plates add 5 mls of 100mM IPTG or 0.119grams of IPTG, bring up in 1 ml of water, filter sterilize with 0.2 micronfilter prior to adding to cool medium.
PHOSPHATE BUFFER
For 500ml:
1M NaH2PO4 (Monobasic): 69g
1M Na2HPO4 (Dibasic): 71g
pH 7.2
140ml of 1M Monobasic
360ml of 1M Dibasic
OR
pH 7.0
195ml of 1M Monobasic
305ml of 1M Dibasic
500ml total;
-bring vol. to 1 liter and pH at 900ml
Red Loading Dye
60% Sucrose
1mM Cresol Red
5% Yellow Food dye