BSS (1L)
3.2 g NaCl
3.0 g KCl
1.8 g MgSO4 7H2O
0.69 g CaCl2 2H2O
1.79 g Tricine
3.6 g Glucose
17.1 g Sucrose
1.0 g BSA
adjust to pH 6.95 with NaOH
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4% Formaldehyde (fixing embryos)
IMPORTANT! WEAR GLOVES AND MASK AT ALL TIMES!
Make about 200 ml in 500 ml bottle or 400 ml in 1000 ml bottle because the solution is
sensitive to acids and bases.
Add ~160 ml ddH2O to 500 ml bottle
Add 8 g (4 g/100 ml) of paraformaldehyde (cold room)
Add NaOH pellet
Stir on plate until dissolved
pH to 7.4 (be very careful!)
Fill to 200 ml
Label and keep in cold room
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Drosophila Ringer
NaCl
KCl
NaHCO3
CaCl3
g/200 ml
32.5
2.8
4.0
2.4
M
1.1*10-1
1.88*10-3 1
2.3*10-3
1.08*10-3
100 mls
4
1
1
1
NaH2PO4
0.2
1.85*10-4
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M9
Na2HPO4
KH2PO4
NaCl
NH4Cl
g/l
6g
3g
0.5 g
1.0 g
PH 7.0
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10x PBS
80 g NaCl
2 g KCl
14.4 g Na2HPO4
2.4 g KH2PO4
800 ml ddH2O
pH to 7.4 w/ NaOH
add to 1 L
autoclave!
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PEM Buffer (for 200 ml)
40 ml .5 M pipes
0.8 ml 500mM EGTA
0.4 ml 500 mM MgSO4
pH 6.95
filter sterilize
store @ 4 degrees Celsius
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0.5 M EDTA (pH 8.0)
1
90 DDW
186.1 g EDTA
800 ml H2O
keep adding NaOH pellets o clear solution
bring to 8.0 after solution dissolves
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100 mM PIPES
10mMEDTA
3.0 g pipes in ~70 ml ddH20
make a 1 M NaOH soln. (1 g/25 mls)
add NaOH soln. until pH 7.0
then add 4 mls of 0.25 M EDTA
adjust volume to 100 mls
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Potassium Acetate (for Plasmid Prep)
To 60 ml of 5 M Potassium acetate,
Add 11.5 ml of glacial acetic acid + 28.5 ml of ddH20
Resulting solution is 3 M potassium, 5 M acetate
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SOC Medium
Per Liter:
20 g
5g
5g
bactotrypone
bactoyeast extract
NaCl
Add to 950 ml H2O and shake to dissolve
Add 10ml of 250 mM KCl
Adjust to pH 7.0 (~0.2 ml of 5N NaOH)
Bring volume to 975 ml
Autoclave
Add 5 ml of sterile 2 M MgCl2 and 20 ml of sterile 1M glucose
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20x SSC (4L)
353 g NaCitrate (citirc acid trisodium salt)
701 g NaCl
dissolve in 3.5 L H2O
adjust pH to 7.0 with 1M HCl (should take ~5mls)
make up to 4 L
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20x TAE
4L
| 1L
387.2g | 96.8 g
32.8 g | 8.2 g
23.4 g | 5.84 g
|
3600ml| 900ml
136.8ml| 34.2 ml
Trizma Base
Na Acetate
Na2 EDTA
ddH2O
glacial acetic acid
Add H2O to 1L final volume
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10x TBE
108 g Tris Base
55 g Boric Acid
3.76 g Na2 EDTA (or 40mls 0.5 M pH8)
add H2O to 1 L
pH to 8.3
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TBS (1x)
Per 1 L
150 mM NaCl = 8.77 g
50 mM Tris-HCl = 50 ml 1.0 M Tris-HCl (pH 8.1)
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1M TRIS
121.1 g Tris base
mix in 800 ml ddH2O
Adjust to desired value (estimates in manual)
Bring water level to 1 L
pH
HCl
7.4
7.6
8.0
70 ml
60 ml
42 ml
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1M TRIS (250 mls total)
30.3 g Tris base
dissolve in 150 to 200 mls ddH2O
add concentrated HCl (~20 mls probably more)
allow to cool to room temp. prior to final pHing
pH 4.5
autoclave
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0.5 M TRIS
60.55 g/L Tris base
add ~800 ml ddH2O
pH to desired level -- allow to come to room temp.
fill to the liter
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YT
16 g bactotryptone
10 g yeast extract
5 g NaCl
(10 g agar/L for plates)
pH 7.4
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MINIPREP SOL 1
4.505 g glucose
12.5 mls 1M tris pH 8
10mls .5M EDTA
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TE
Per 1 L
2 ml 0.5M EDTA pH 8.0
10 ml 1M Tris pH 8.0
Bring water level to 1 L
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RNAse
To make 20ml @ 5mg/ml = 100 mg
Dissolve in 10mM Tris pH 7.5 and 15mM NaCl
Combine:
200 l Tris (1M, pH 7.5)
60 l NaCl (5M)
19.74 ml ddH2O
Aliquot 1ml/ eppy. Label "5mg/ml RNAse crude." Store in -20C in cup in door
of freezer
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TE in RNAse
50 ml TE
400 l RNAse
Boil for 5 minutes
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