Novel Application of Reversed Gel Electrophoresis: Reuse of DNA Molecular Weight Marker (100 bp DNA ladder)

100 bp DNA ladder is reused here for one time: The well corresponding to resolved DNA ladder is sealed with molten agarose. The resolved 100 bp DNA ladder is made to converge to a single band by reversing the electrode polarity or by facing the wells in gel towards the anode. The gel slice containing the DNA ladder is removed using scalpel or any other suitable device very carefully so that a thin rectangular gel piece containing the ladder is obtained. The gel slice or gel plug can be stored in water or in buffer at 4C in refrigerator.

For reuse the gel plug is placed in a fresh well of agarose gel and the well is sealed with agarose. The test DNA samples are loaded in other wells. The electrophoresis again resolves DNA ladder from gel plug.

The prior art internet search reveals commercially available low melting agarose gel plugs (eg., InCert agarose
gel plugs sold by Lonza) containing large DNA fragments such as lambda DNA ladder that are
used in Pulsed Field Gel Electrophoresis (PFGE).

We have designed methods using reversed gel electrophoresis that favourably helps in reuse of
resolved 100 bp DNA ladder.

本次实验将100 bp DNA分子量标准（100 bp DNA ladder）进行单次复用：将已完成电泳分离的DNA分子量标准对应的点样孔，以熔融琼脂糖进行密封。通过反转电极极性，或将凝胶中的点样孔朝向阳极，可使已分离的100 bp DNA分子量标准汇聚为单一条带。使用手术刀或其他合适工具小心切下含有该分子量标准的凝胶条带，即可得到一块包含该DNA分子量标准的薄长方形凝胶块。该凝胶切片或凝胶块可置于水中或缓冲液中，于4℃冰箱内保存。

复用该凝胶块时，将其置于新制备的琼脂糖凝胶的空白点样孔中，再以琼脂糖密封该孔；将待测DNA样品点样至其余孔位，再次通过电泳即可从凝胶块中分离得到该DNA分子量标准。

经现有技术网络检索可知，市面上已有商业化的低熔点琼脂糖凝胶块（low melting agarose gel plugs），例如龙沙（Lonza）公司售卖的InCert琼脂糖凝胶块（InCert agarose gel plugs），其内置有大片段DNA（如λ DNA分子量标准（lambda DNA ladder）），用于脉冲场凝胶电泳（Pulsed Field Gel Electrophoresis，PFGE）。

本研究开发了基于反向凝胶电泳（reversed gel electrophoresis）的方法，可有效实现已分离的100 bp DNA分子量标准的复用。