摘要:MATERIALS:2-glass plates　　1 sharks -tooth comb and spacers　　Whatman 3 mm paper　　30 or 40% acrylamide-bis (19:1)　　10X TBE　　urea　　10% ammonium persulfate　　TEMED　　60 cc. syringe　　Rain-ex Preparation of g[阅读全文]

摘要:Procedure Add the following to a sterile microcentrifuge tube:Labeled target DNA (3-6ng)1-8ulCalf Thymus DNA (0.5ug/ul)2ulTE buffer0-7ulTotal Volume10ulAdd 1ul of 4% Formic Acid and incubate for 25 mi[阅读全文]

摘要:Antibiotic Stock SolutionsAmpicillinBeta-lactam-antibiotics are not very stable when dissolved. Slow but steady degradation happens even when frozen to -20°C. Therefore commercial beta-lactam-anti[阅读全文]

摘要:1. Spin 50 ml freshly grown cells.2.Wash wash pellet twice with PBS.3. Resuspend the pellet in 1ml TNE buffer ( same buffer as above. use 2 tubes, 500 ul each).4. Add 5 ul proteinase K and incubate fo[阅读全文]

摘要:1. Spin 1.5ml fresh cells for 3 minutes.2.Wash the pellet once with TNE buffer.3. Suspend the pellet well in 20 ul TNE buffer.4. Add 2 ul proteinase K ( stock 10 ug/ul in water, -20 C).5. Add 20 ul lo[阅读全文]

摘要:Antibiotic Stock SolutionsAmpicillinBeta-lactam-antibiotics are not very stable when dissolved. Slow but steady degradation happens even when frozen to -20℃. Therefore commercial beta-lactam-antibioti[阅读全文]

摘要:These are basically Tim's procedures and have been used successfully by numerous members of our lab and by others around us.[阅读全文]

摘要:1.Ice on tray.2. Dissection tools: 1 pair of large scissors; 2 pairs of fine scissors; 1 pair of coarse forceps; 2 pairs of fine forceps; soak in 95% EtOH.3. 70% EtOH in a spray bottle.4. 2 sets of 10[阅读全文]

摘要:Three weeks prior to embryonic fibroblast isolation, a PGK-neo male mouse is mated to a heterozygote female. 13.5 days to 14.5 days after the plug is observed, sacrifice the pregnant female either by [阅读全文]

摘要:1. Dilute 2 ml of 4000 U/ml Collagenase D as follows: 1 ml into 9 ml HBSS/Ca2+/Mg2+ (=400U/ml) and 1 ml into 39 ml HBSS/Ca2+/Mg2+ (=100U/ml). Put on ice. 2. Place 5 ml of the 100U/ml solution in a 10 [阅读全文]

摘要:Buffer composition is 10 mM Tris-HCl, pH 7.5, 0.1 mM EDTA, 30 microM spermine, 70 microM spermidine, 100 mM NaCl. This buffer is more likely to produce transgenic mice with intact, unfragmented DNA mo[阅读全文]

摘要:The purity of the transgene DNA used for microinjection is critical for the successful production of transgenic founder mice. DNA impurities in the form of bacterial endotoxins or organic contaminants[阅读全文]

摘要:Injection and Holding PipettesThe glass capillary tubing used should be thin walled, borosilicate glass without a fibre. e.g. Clark Electromedical Instruments Australian Agents: SDR Clinical Technolog[阅读全文]

摘要:These procedures were originally devised in Richard Palmiter's lab for use with tail dots (DNA spotted onto a nitrocellulose filter and probed for a transgene; quicker than doing southerns), and subse[阅读全文]