真核细胞dna提取与酶止切鉴定-liuchang.ppt

* Weigh out 0.5 g of agarose into a 250 mL conical flask. Add 50 mL of 0.5xTBE , swirl to mix. It is good to use a large container, as long as it fits in the microwave, because the agarose boils over easily. * * Microwave for about 1 minute to dissolve the agarose. The agarose solution can boil over very easily so keep checking it. It is good to stop it after 45 seconds and give it a swirl. It can become superheated and NOT boil until you take it out whereupon it boils out all over you hands. So wear gloves and hold it at arms length. You can use a bunsen burner instead of a microwave - just remember to keep watching it. * Leave it to cool on the bench for 5 minutes down to about 60°C (just too hot to keep holding in bare hands). If you had to boil it for a long time to dissolve the agarose then you may have lost some water to water vapour. You can weigh the flask before and after heating and add in a little distilled water to make up this lost volume. While the agarose is cooling, prepare the gel tank ready, on a level surface * * * * * * * Cathode:阴极 * * * Ultraviolet Transilluminator 限制性核酸内切酶 限制性核酸内切酶是识别DNA的特异序列, 并在识别位点或其周围切割双链DNA的一类内切酶。 GGATCC CCTAGG G CCTAG GATCC G + Bam HⅠ Molecular scissor 均来自微生物，并据此而进行命名； 识别序列长度常为4 － 8 核苷酸序列 ; 大部分酶识别序列呈二元旋转对称结构 即回文结构 ; 其切割后的DNA可产生不同的末端。 限制酶的特点（Ⅱ类酶） DNA的琼脂糖凝胶电泳 琼脂糖是从海洋植物琼脂中提取来的，为一种聚合链线性分子。 线性DNA分子在电场中的迁移率与其分子量的对数成反比 DNA的空间构型不同，即使分子量相同其迁移率也不相同 Making an Agarose Gel Casting tray? Gel combs? Power supply? Gel tank? ?Cover Electrical leads ? Electrophoresis Equipment Gel casting tray combs The tray is the actual mold which provides a shape for the gel as it polymerizes. The comb can make some “wells” in the gel. Seal the edges of the casting tray and put in the combs. Place the casting tray on a level surface. None of the gel combs should be touching the surface of the casting tray. Preparing the Casting Tray Agarose Agarose is a linear polymer extracted from seaweed. D-galact