Preparation of commonly used reagents in molecular biology.

1. LB (Luria-Bertani)
culture
liquid, plate preparation
preparation per liter of LB culture, should be added in 950 ml of deionized water:
bacteria culture with
yeast
extract (bacto-yeastextract) 5g
bacteria culture with totalin(bac-tryptone) 10g
NaCl10g
shakes the container until the solute dissolves completely, adjusts the pH to 7.0 with 5mol/LNaOH (about 0.2ml), adds deionized water to a total volume of 1L, and steams sterilise 20min at 151bf/in2 (1.034×105Pa) pressure.
L
agar
plate preparation:
first according to the above formula to make liquid
fase
, before autocultication to add agarose 15g/L, the same method of autobaric steam sterilization 20min. When removing the medium from the autocultension sterilizer, gently spin so that the melted agar sugar can be evenly distributed throughout the medium solution, and when the medium is cooled to 50 degrees C, add heat-resistant substances such as
igre antibiotics
. To avoid bubbles, the medium should be whirlwinded when mixing, and then the medium-paved plate can be poured directly from
smotts
. A 90mm diameter petri dish requires about 30-50ml of a culture base, which, after complete condensation, should be flattened upside down and stored in a spare of 4C.2. Preparation of agarose gel
take agarose according to the desired gel concentration scale, add the corresponding electrophoresis buffer, microwave oven
heat
boil until agar sugar is completely dissolved, add the appropriate amount of E mixing Proper cooling and dumping into the gel cast tank, insert comb, gel thickness does not exceed the comb hole, if there are bubbles produced by the glass rod to remove, can not be prematurely removed comb, should wait for the gel to fully gel before removing the comb.3. P1, P2, P3 preparation
P1 preparation: in 800 ml of deionized water dissolved tri-alkali 6.06g, Na2
EDTA
2H2O3.72g, with HCl adjustment H to 8.0, with deionized water adjustment volume to 1 liter, per liter P1 add RNaseA100mg.
P2: dissolved in 950ml deionized water aOH8.0g, 20% SD50ml, adjusted volume to 1 litre.
P3 preparation: in 500 ml of deionized water dissolved potassium acetate 294.5g, with ice acetic acid to adjust the H value to 5.5, with deionized water to adjust the volume to 1 liter.4. Common buffers:
TE
pH7.4
10mmol/LTris-HCl (pH7.4)
1mmol/LEDTA (pH8.0)
pH7.) 6
10mmol/LTris-HCl (pH7.6)
1mmol/LEDTA (pH8.0)
pH8.0
10mmol/LTris-HCl (pH8.0)
1mmol/LEDTA (pH8.0)
STE (also known as TEN)
0.1mmol/LaCl
10mmol/LTris-HCl (pH) 8.0)
1mmol/LEDTA (pH8.0)
STET
0.1mmol/LaCl
10mmol/LTris-HCl (pH8.0). )
1mmol/LEDTA (pH8.0)
5% TritoX-100
TNT
10mmol/LTris-HCl (pH8.0)
150mmol/LaCl
0.05% Twee20
Electrophoretic buffer
Tris-acetic acid (TAE): 50× thick storage (per liter): 242g Tri-alkali
5. 7.1ml ice acetic acid
100ml 0.5mmol/LEDTA (pH8.0)
diluted 50 times when used..