密歇根大学生物系实验室的常用试剂配方.doc

Table of ContentsLB Medium.1NZ Medium.2SM Buffer.3SET Buffer.46X Prehyb Soln.510 X TBE.610 X TAE.720 X SSC.81% SDS, 0.2 M NaOH.914% PEG (8000), 2M NaCl, 10 mM MgSO4.1020% SDS.111.0 M Tris, pH 8.0, 1.5 M NaCl.1210mM Tris-HCl, pH 7.5, 10mM MgSO4.1310 mM Tris, 50 mM EDTA, pH 7.5.1410 mM Tris-HCl, 1 mM EDTA, pH 7.5.153 M Sodium Acetate, pH 4.8.16Electrophoresis dye.17Labelling Stop dye.18Sequencing gel dye.195% Acrylamide.206% Acrylamide in TBE, 50% Urea.2140% Acrylamide.22LB Medium (1 Liter)10g Bacto-tryptone5g Bacto-yeast extract10g NaClFor forty plates add 1% agar-1g. Autoclave media. When cool, add ampicillin and pour plates. For 1L of media, add 1.8 mL amp.NZ Medium (500 mL)5 g Bacto-tryptone2.5 g Bacto-yeast extract2.5 g NaCl1.25 g MgSO4For 20 plates add 1.2% agar-6g. Autoclave and pour plates at 50o CSM Buffer (1L)5.8 g NaCl1.2 g MgSo450 mL 1M Tris-HCl, pH 7.50.1 g Gelatin (doesnt dissolve)AutoclaveUsed for phage dilution and storage.SET Buffer50 mM Tris-HCl, pH 8.0, 50 mM EDTA, 20% w/v Sucroseto make 200mL:40 g Sucrose10 mL of 1M Tris20 mL of 0.5 M EDTA, disodium saltbring to 200 mL with H206X Prehybridization Solutionto make 500 mL300 mL ddH20150 mL 20X SSC50 mL 50X Denhardts solution1 mL 0.5 M EDTA (disodium salt)2.5 mL 20% SDS6X refers to the concentration of SSC10X TBE Buffer (for polyacrylamide gels)to make one liter:60.75 g Tris3.7 g EDTA (tetrasodium salt)30 g Boric acid10X TAE Buffer (For agarose gels)to make one liter:48.20 g Tris6.75 g NaAce3.75 g EDTA (disodium salt)Adjust pH to 7.6 with acetic acid. (Approx. 20 mL)20X SSCto make one liter:175.3 g NaCl88.2 g NaCitrateadd water to bring volume to one liter.adjust to pH 7.0 with HCl.1% SDS, 0.2 M NaOHto make 100 mL:93 mL ddH205 mL 20% SDS2 mL 10 M NaOH 14% PEG (8000), 2M NaCl, 10 mM MgSO4 to make one liter:140 g PEG117 g NaCl2.46 g MgSO4For use in phage DNA preparation.20% SDSto male 250 mL:50 g of SDS in a beakerAdd stir bar and H20 last.This solution will have to be heated for the SDS to dissolve.1.0 M Tris, pH 8.0, 1.5 M NaClto make one liter:121.1 g Trizma87.6 g NaClin a volume of water less than 1L. Adjust pH with HCl, then bring to 1L with H2010 mM Tris-HCl, pH 7.5, 10 mM MgSO4 to make one liter:10 mL 1 M Tris-HCl2.46 g MgSO4for use in phage DNA preparation 10 mM Tris, 50 mM EDTA, pH 7.5to make 200 mL:2 mL 1 M Tris20 mL 0.5 M EDTA (tetrasodium salt)178 mL ddH20adjust pH with HCl.10 mM Tris-HCl, 1 mM EDTA, pH 7.5to make 200 mL:2.0 mL 1 M Tris-HCl, pH 7.50.4 mL 0.5 M EDTA197.6 mL ddH203 M Sodium Acetate, pH 4.8to make one liter:408.1 g NaAce (trihydrate; gets cold in soln)about 700 mL H20adjust pH with glacial acetic acid (takes a lot) Measure tru pH by dilution with water; range will be between 4.8 and 5.5.Electrophoresis Dyeto make 4 mL:3 mL 50 mM EDTA, 10 mM Tris-HCl, pH 8.01 mL glycerol20 mL BPB10 mL Xylene cyanolStop dye for labelled probe1 mL 50 mM EDTA, 10 mM Tris, pH 7.5-8.5about 200 ml glyceroladd a few grains of blue dextran (8000)Sequencing gel dyefor approx 1 mL:1 mL formamide 10 mL xylene cyanol10 ml BPB3 mL 10 M NaOH5% acrylamideto make 200 mL:20 mL 10X TBE25 mL 40% acrylamide155 mL H206% Acrylamide in TBE, 50% Ureato make 500 mL:50 mL 10X TBE75 mL