发布者：Primerbank 时间：2017-09-29 浏览量：974
LB培养基
NaCl 10 g
Yeast extract 5 g
Peptone 10 g
Add dH2O to 1000 ml
Aliquot to 500ml flasks (250 ml per flask). Seal with sealfilm
and autoclave them. Store at room temperature.
LA固体培养基
NaCl 10 g
Yeast extract 5 g
Peptone 10 g
Agar powder 13～15 g
Add dH2O to 1000 ml
Aliquot to 500 ml flasks (250 ml per flask). Seal with sealfilm
and autoclave them. Store at room temperature.
X-gal (20mg/ml)：
20mg X-gal溶于1ml 二甲基甲酰胺中，-20℃避光保存；
IPTG (200mg/ml)：
1g IPTG溶于4ml去离子双蒸水中，定容至5ml，过滤灭菌后-20℃保存；
Amp（100mg/ml）：
1g Amp溶于4ml去离子灭菌双蒸水中，定容至5ml，-20℃保存
Solution Ι：
Cell resuspension solution (50 mM Tris-HCl, pH 7.5, 10 mM EDTA, RNase A 100 μg/ml)
1 M Tris-HCl (pH 7.6) 2.5 ml
0.25 M EDTA 2.0 ml
ddH2O 45 ml
sterile by autoclave
add 1% RNase 0.5ml
store at room temperature
Solution Ⅱ：
Cell lysis solution (0.2 N NaOH, 1% SDS)
Mix 0.4 N NaOH and 2% SDS in same volume.
Solution Ⅲ：
Neutralization solution (1.32 M potassium acetate, pH 5.2)
5 M potassium acetate 13.2 ml
Hac 27 ml
adjust pH to 5.2
add ddH2O to 50 ml
sterile by autoclave
store at room temperature
RNaseA: 10mg/ml溶于TE中，在沸水中煮10－30分钟之后分成小份储存于－20℃
1.5×CTAB：
CTAB 15 g
1 M Tris·Cl (pH 8.0) 75 ml
0.5 M EDTA 30 ml
NaCl 61.4 g
add ddH2O to 1000 ml
0.5 M EDTA (pH 8.0)：
EDTA-Na·2H2O 186.1 g
NaOH ~20 g
Adjust to pH 8.0
dH2O to 1000 ml
sterilize by autoclaving
1 M Tris·HCl：
pH 7.4 pH7.6 pH 8.0
Tris base 121.1 g 121.1 g 121.1 g
Concentracted HCl ~70 ml ~64 ml ~42 ml
dH2O to 1000 ml 1000 ml 1000 ml
Sterilize by autoclaving
TE (pH 8.0) ： Stock vol.
10 mM Tris·HCl (pH 8.0) 1 M 10 ml
1 mM EDTA (pH 8.0) 0.5 M 2 ml
dH2O to 1000 ml
sterilize by autoclaving
10 M NH4Ac ：
NH4Ac 385 g 770 g
H2O to 500 ml 1000 ml
10×PCR buffer：
stock vol.
500 mM KCl 2.5 M(sterilized) 200 ml
100 mM Tris-HCl 1 M pH 9.0(sterilized) 100 ml
1% Triton X-100 100% 10 ml
ddH2O 690 ml
sterilize by autoclaving
5×TBE：
Tris 54 g
Boric acid 27.5 g
0.5 M EDTA (pH 7.9) 20 ml
dH2O to 1000 ml
10×TAE：
Tris 121.1 g 484.4 g
EDTA(0.5 M) 20 ml 80 ml
NaAc·3H2O 17 g 68 g
glacial acetic acid ~30 ml ~200 ml
adjust to pH 8.1
dH2O to 1000ml 4000ml
NaOH ：
10 N 4 N
NaOH 400 g 160 g
dH2O to 1000 ml 1000 ml
2 N HCl：
concentrated HCl 3 65 ml 182.5 ml
dH2O to 2000 ml 1000 ml
5 mg/ml ssDNA：
Salmon sperm DNA 1 g
ddH2O to 200 ml
0.5 M P.B (phosphate Buffer) pH 6.8：
Na2HPO4 16.44 g 131.52 g
NaH2PO4 16.11 g 128.88 g
dH2O to 500 ml 4000 ml
20×SSC：
NaCl 175.3 g 701.2 g
Na3Citrate 88.2 g 352.8 g
dH2O to 1000 ml 4000 ml
Sterilize by autoclaving
10% SDS ：
SDS 100 g
dH2O to 1000 ml
Heat to 68 ℃ to assist dissolution
50×Denhart’s Solution：
Ficoll 400 10 g
PVP-360 10 g
BSA (Fraction V) 10 g
ddH2O to 1000 ml
Southern Blot Hybridization Buffer (Saghai,s Lab)：
Final conc. Stock Vol.
5×SSC 20× 250 ml
50 mM PB (pH 6.8) 0.5 M 100 ml
5×Denhardt’s 50× 100 ml
2.5 mM EDTA (pH 8.0) 0.5 M 5 ml
100 μg/ml ssDNA 5 mg/ml 20 ml
0.4%SDS 20% 20 ml
Dextran sulfate 50 g
ddH2O to 1000 ml
(Place a beaker on a stirrer, add these solution in the order of appearance one by one. SDS should be the very last item.)
Washing off Probe for Re-hybridization of Blots (I)：
Washing time: 10 min
Final conc. Stock Vol.
0.1×SSC 20×SSC 20 ml
0.1% SDS 10% SDS 40 ml
dH2O to 4000 ml
Washing off Probe for Re-hybridization of Blots (II)：
Washing time: 3 min
Final conc. Stock Vol.
0.1 N NaOH 10 N NaOH 40 ml
0.2% SDS 10% SDS 80 ml
dH2O to 4000 ml
Washing off Probe for Re-hybridization of Blots(Ⅲ)：
Washing time: 20 min
Final conc. Stock Vol.
0.2 M Tris. (pH 7.5) 1 M Tris. (pH 7.5) 800 ml
0.1×SSC 20×SSC 20 ml
0.2% SDS 10% SDS 80ml
dH2O to 4000ml
Blue Juice：
Final conc. Stock Vol. Vol.
70% Glycerol 100% 35 ml 70 ml
0.5×TBE 5× 5 ml 10 ml
0.2% SDS 10% 1 ml 2 ml
20 mM EDTA 0.5 M 2 ml 4 ml
5 mg/ml Bromphenol Blue 0.25 g 0.5 g
5 mg/ml Xylene cyanol 0.25 g 0.5 g
dH2O to 50 ml 100 ml
EB (10 mg/ml)：
ehidium bromide 1 g
dH2O to 100 ml
Stir on a magnetic stirrer for several hours. Transfer the solution to
a dark bottle and store at 4℃.
The concentration of work solution: 0.5 μg/μl (50 μl stock solution
In 1000 ml dH2O).
Decontamination of EB：
Reduce the concentration of EB <0.5 mg/ml, add 1 volume of
0.5 M KMnO4,mix carefully then add 1 volume of 2.5 N HCl,
mix carefully and allow the solution to stand at room temperature
for several hours. Add 1 volume of 2.5 N NaOH, mix and discard.
1M NaAc(PH7.0)：82g NaAc 先加一定量ddH2O，用NaOH调PH值至7.0，再用ddH2O定容至1L，灭菌
0.5M EDTA(PH8.0): 186.1g EDTA先加一定量ddH2O, 用NaOH调PH值至8.0,再用ddH2O定容至1L，灭菌
10×MOPS buffer: (用DEPC水配，再灭菌)
MOPS 41.85g
1M NaAc(PH7.0) 50ml
0.5M EDTA(PH8.0) 20ml
先加一定量DEPC水，再用4N NaOH 调PH 至7.0（约加7ml），再用DEPC水定容至1L，灭菌
1M NaH2PO4 buffer ( PH7.2):
NaH2PO4 71g
H3PO4(85%) 4ml
用灭菌的DEPC水定容至1L
20×SSC:
NaCl 175.3g
Na3Citrate 88.2g
用ddH2O定容至1L，灭菌
10%SDS:
SDS 100.0g
用灭菌的ddH2O定容至1L(将水加热到68℃有助于溶解)
Sample buffer：
Deionized formamide 1000 ml
10×MOPS buffer 200 ml
37% formaldehyde 320 ml
Blue juice(loading buffer):
Glycerol 70ml
5×TBE 10ml
10%SDS 2ml
0.5M EDTA(PH8.0) 4ml
Bromphenol Blue 0.5g
Xylene Cyanol 0.5g
加灭菌的ddH2O定容至 100ml
Hybridization buffer:
1M NaH2PO4 buffer 500ml
0.5M EDTA(PH8.0) 2ml
10% SDS 70g
BSA 10g
用灭菌的DEPC水定容至1L，贮存于室温下
洗膜液Ⅰ:（for 1 littre）
20×SSC 100ml
10%SDS 10ml
洗膜液Ⅱ:（for 1 littre）
20×SSC 25ml
10%SDS 10ml
洗膜液Ⅲ:（for 1 littre）
20×SSC 5ml
10%SDS 10ml
4×SSC:
20×SSC 200ml
用灭菌的DEPC水定容至1L
2×SSC:
20×SSC 100ml
用灭菌的ddH2O定容至1L