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使用微型垂直电泳槽从琼脂糖凝胶中高效小规模电洗脱高分子量DNA

Efficient, small scale electroelution of high molecular weight DNA from agarose gels by a miniature vertical electrophoresis cell.

作者信息

Pascali V L, Pescarmona M, Dobosz M, d'Aloja E

机构信息

Immunohematology Laboratory, Istituto di Medicina Legale, Università Cattolica del S. Cuore, Roma, Italy.

出版信息

Electrophoresis. 1991 Apr;12(4):317-20. doi: 10.1002/elps.1150120417.

Abstract

A miniature, vertical electrophoretic system is described that allows quick elution and efficient harvesting of double-stranded, high molecular weight DNA fragments from agarose gels. A gel slice containing the band of interest is excised from the agarose gel after submarine electrophoresis and placed into an electrophoresis cell, assembled by inserting a truncated micropipette tip into a 2.0 mL polypropylene tube. The tip supports the agarose slice and is connected to dialysis tubing. Platinum wire electrodes are placed over the tip and at the bottom of the test tube. DNA molecules are quickly eluted into the dialysis bag (60 s to 10 min for bands ranging from 2 to 23 kb) and then easily recovered. The method is highly efficient and allows considerable time saving. Of special interest is its applicability to small starting amounts of DNA (less than 0.5 micrograms per band). Purity of the DNA is comparable, if higher, to that obtainable by conventional electroelution on dialysis bags and DEAE-cellulose membrane electrophoresis. The system is simple enough to be used as a routine method for eluting multiple bands in short times.

摘要

本文描述了一种微型垂直电泳系统,该系统能够从琼脂糖凝胶中快速洗脱并高效回收双链高分子量DNA片段。在水下电泳后,从琼脂糖凝胶中切下含有目标条带的凝胶块,将其放入电泳槽中,电泳槽是通过将截断的微量移液器吸头插入2.0 mL聚丙烯管中组装而成。吸头支撑琼脂糖凝胶块,并与透析管相连。铂丝电极分别置于吸头上方和试管底部。DNA分子可快速洗脱到透析袋中(2至23 kb的条带洗脱时间为60秒至10分钟),然后便于回收。该方法效率高,可节省大量时间。特别值得关注的是,它适用于起始DNA量较少的情况(每条带少于0.5微克)。所获得DNA的纯度与通过传统的透析袋电洗脱法和DEAE - 纤维素膜电泳法相当,甚至更高。该系统足够简单,可作为在短时间内洗脱多个条带的常规方法。

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