Recycling of anion-exchange resins for plasmid DNA purification.

Plasmid DNA of high purity is critical for molecular biology studies such as subcloning, DNA sequencing, vector construction and expression and functional analysis. The classical method of plasmid DNA purification using cesium chloride/ethidium bromide buoyant-density centrifugation is generally time-consuming and expensive and has gradually been replaced by the use of silica-based, anion-exchange resins. Our laboratory (and many others) has been using the anion-exchange resin tips (QIAGEN-tips; Qiagen, Valencia, CA, USA) in our routine work of purification of plasmid DNA with satisfactory results. However, the frequent use of QIAGEN-tips can also be very costly if each tip is discarded after a single use as suggested by the manufacturer (1). To minimize the cost of purchasing new tips, we have developed a protocol to recycle them after each use. We have found that the plasmid DNAs purified from the recycled QIAGENtips are equally high quality as those isolated from the new tips. No crosscontaminations from the previous usages were found in the plasmid DNA obtained from the recycled tips. Here, we describe our protocol for recycling the QIAGEN-tips. After elution of plasmid DNA following the procedures suggested by the manufacturer, the used tips can be air-dried without further treatment and stored at room temperature until the next use. Alternatively, the tips can be rinsed briefly with deionized water to remove any particulate materials that are trapped on top of the packed resins and salt deposits on the outer surface of the tips before storage. However, a thorough rinse of the resins with deionized water is not necessary. Immediately before applying the used tips for plasmid DNA purification, one volume [as recommended for the equilibrium buffer in the Qiagen handbook (1)] of “renew” buffer (3 M NaCl, 0.15% Triton X100) is added into the tips and allowed to flow through to remove any residual material from the previous use. To minimize the possibility of contamination from the plasmid DNA that was purified during the previous application, we generally wash the used tips with the renew buffer 2–3 times. The refreshed tips can then be equilibrated with equilibrium buffer for plasmid DNA isolation according to the procedures recommended by the manufacturer. Based on our experience, these tips can be recycled for a minimum of 20 times without a noticed deterioration in performance. Figure 1 shows that the yields of the plasmid DNA from the new and recycled QIAGEN-tip 500 (Maxi) are equivalent and are at 2.75 and 2.30 μg/mL of bacterial culture, respectively. It was noticed that the flowrate of the recycled tips tends to gradually slow down after multiple use, due to the settlement of resins during each use. This inconvenience can be circumvented by flushing the renew buffer from the bottom of the tips using a plastic tubing connected to a syringe to dislodge the packed resins. Finally, note that, although this protocol of recycling has been developed for the QIAGENtips, it is conceivable that the same method could also be applied in recycling similar plasmid purification systems manufactured by other commercial firms.