Comparative Studies on Preparation of Large Plant DNA Fragments by Pulse Field Gel Electrophoresis

The preparation of large plant DNA fragments is extremely important to the construction of large insert DNA libraries (YAC, BAC, PAC and TAC). Although several techniques have been developed in each step of large plant DNA fragments preparation, the whole processing remains complicated and difficult. Based on authors research experience and the recent worldwide development in this field, the following aspects are discussed in this paper: techniques of plant high molecular weight (HMW) DNA purification by pre-electrophoresis, the optimal conditions for the partial digestion of the HMW DNA by HindIII, the isolation effects of of large plant DNA fragments (100～400 kb) with different parameters of pulse field gel electrophoresis (PFGE), and the recovery of large DNA fragments. Through comparative studies, the advantages and disadvantages of each technique are discussed and some recommendations are proposed for preparing high quality large plant DNA fragments. The suggested techniques have been used in preparing the large DNA fragments of maize, rice, moss, laver, sea tangle and peach,and similar results are obtained among all the materials. This paper only reports the results using maize as material.