The stock solution of common chemicals
| Final volume | 100 ml | 500 ml | 1000 ml |
| Acrylamide | 29 g | 145 g | 290 g |
| Bisacrylamide | 1 g | 5 g | 10 g |
Weigh out the components above, and begin dissolve them in about 60% of the final volume of ddH2O. Heat to 37¡ãC until completely dissolved. Check pH: if above 7.0, add HCI until at or below 7.0. Filter through 0.45 um filter. Wrap bottle in foil, and store in dark at 4¡ãC.
0.7% Agarose for gel electrophoresis
Mix:
2.8 g agarose
8 ml 50xTAE
400 ml ddH2O
Dissolve agarose completely by microwaving. Cool at 60 ¡ãC
and add 20 ml ethidium bromide solution (10 mg/ml) before use.
10% Ammonium persulphate
Weight 1 g of ammonium persulphate in 15 ml Corning Tube and add about 9.5 ml of sterile ddH2O. Vortex until fully dissolved, and store at 4¡ãC.
1000x Ampicillin stock solution
Dissolve 1 g ampicillin in 10 ml of sterile ddH2O. Filter through a 0.22 um filter, Store at -20¡ãC.
1 M Dithiothreitol (DTT)
Dissolve 15.45 g DTT in 100 ml H2O. Store at -20¡ãC
6x DNA Loading dye
25mg bromophenol blue (0.25%), 25mg xylene xyanol (0.25%), 4g sucrose (40%), adjust volume to 10 ml with H2O. Store at room temperature.
0.5 M EDTA, pH 8.0 (disodium ethylenediamine tetraacetate):
Add 186.1 g EDTA (Sigma # E-5134) to 800 ml of ddH20. Add about 20g of NaOH pellets while stirring to bring the pH to 8.0. Add the last few grams slowly to avoid overshooting the pH. Note that the EDTA won’t completely dissolve until the pH is around 8. Filter with 0.5 micron filter and autoclave.
10 mg/ml ethidium bromide (EtBr):
Add 100 mg EtBr (Sigma E-8751) to 10 ml ddH2O. Wrap in foil, store in dark.
40% Glucose
Dissolve 200 g (D)-glucose in a final volume of 500ml ddH2O. Filter sterilize (0.22 um filter), and store at room temperature.
1 M HEPES, pH 7.5
Add 23.83 g HEPES (Sigma H-3375) to 100 mo of ddH2O. Adjust pH to 7.5 with potassium hydroxide (KOH) and store at 4¡ãC
500x Kanamycin stock
Dissolve 0.5 g kanamycin in 10 ml of sterile ddH2O. Filter through a 0.22 um filter. Store at -20¡ãC.
LB plates:
Mix the following in a 2 liter flask:
Weigh out 32 g pre-made LB agar mix, which is composed of:
10 g Bacto-Tryptone
5 g Bacto-yeast extract
5 g NaCl
12 g Bacto-agar
Add magnetic stirring bar, add 1 liter ddH2O. Autoclave immediately, and cool to 50¡ãC in water bath. Add antibiotic if desired, and quickly pour into sterile petri dishes until dish surface is completely covered. After solidification, open the dishes in the hood and dry the plate for about 1 hour.
For ampicillin, use 0.5 ml of 1000x amp stock per 500 ml media.
For kanamycin, use 1 ml of 500x kan stock per 500 ml media
10 x PBS (Phosphate buffered saline)
| Chemical | amount | 10x conc | 1x conc |
| NaCl | 80 g | 1.37 M | 137 mM |
| KCl | 2 g | 27 mM | 2.7 mM |
| Na2HPO4-7H2O | 11.5 g | 43 mM | 4.3 mM |
| KH2PO4 | 2 g | 14 mM | 1.4 mM |
Dissolve in ddH2O to a final volume of 1 liter. There is no need to adjust pH for this solution. (The final pH will be ~7.3.) Autoclave and store at room temperature. The 10x solution can be diluted as 1:10 with sterile ddH20 to prepare a 1xworking solution.
PMSF (phenylmethylsulphonyl fluoride), 100 mM
Dissolve 174 mg PMSF in 10 ml isopropanol. Store at -20¡ãC.
Q-solution (5 M betaine)
Dissolve 29.275 g betaine in final volume of 50 ml sterile ddH2O. Sterilize by 0.22 um filter, and store at -20¡ãC. This solution is helpful for high GC content template in PCR.
Sodium acetate, 3 M
Dissolve 408 g sodium acetate(x3H2O) in 800 ml ddH2O. Adjust pH to 5.0 with glacial acetic acid. Adjust to 1 L final volume with ddH2O.
Sodium chloride, 5 M
Weigh out 292 g NaCl, and add ddH2O to one liter final volume. Autoclave and store at room temperature.
SSC (sodium chloride/sodium citrate), 20x
Dissolve 175.3 g NaCl and 88.2 g sodium citrate in 800 ml ddH2O. Adjust pH to 7.0 with 10 N NaOH. Adjust final volume to 1 liter, and autoclave. Store at room temperature.
SDS, (sodium dodecyl sulphate), 10%
Dissolve 100 g SDS in 900 ml ddH2O. Heat solution to about 68¡ãC to assist in dissolution. Adjust pH to 7.2 with 1 N HCl. Adjust final volume to 1 liter.
Sodium hydroxide, 10 N
Dissolve 400 g NaOH in a final volume of 1 liter ddH2O. Store at room temperature.
TAE (Tris/acetate/EDTA) electrophoresis buffer
50x stock solution:
| 242 g Tris base | 40 mM Tris-acetate |
| 1.86 g Na2EDTA-2H2O | 0.1 mM Na2EDTA-2H2O |
| 57.1 ml glacial acetic acid | pH ~8.5 |
Add ddH2O to 1 liter, autoclave
Make 1 x TAE by diluting 1:50 (eg.: 20 ml 50 x TAE/980 ml ddH2O)
1x TE (Tris/EDTA) buffer
10 mM Tris-Cl, pH 8.0
1 mM EDTA, pH 8.0
100 ml 1 M Tris-Cl, pH 7.5
9 g NaCl
Add ddH2O to final volume of 1 liter. Store at 4¡ãCIPTG and X-gal for blue/white selection
IPTG: isopropyl thiogalactoside, or isopropyl beta-D-thiogalactopyranoside
pStock Solution Concentration: 0.1 Mp
Dissolve: the molecular weight is 238.3, so this is 0.238 g in 10 ml of water. Sterilize by filtration, then store at -4¡ãC.
X-gal: 5-bromo-4-chloro-3-indolyl-beta-D-galactopyranoside
Stock Solution Concentration: 20 mg/ml
To 100 mg X-gal add 5 ml dimethylformamide (DMF) or DMSO and store at -20C.
It must be dissolved in DMSO (dimethyl sulfoxide) or dimethyl formamide, not water! It must be wrapped in foil to protect it from the light, and stored in the freezer. X-gal solutions do not need to be sterilized.
