Description :  

Introduction

The EZgene^TM high-capacity-plasmid DNA Isolation Kit is designed for rapid purification of BAC, PAC, cosmid and P1 from small volume of bacterial cultures. It is based on a modified alkaline lysis procedure that is specially adapted for spin column. The procedure associated with this kit has been tested using a variety of low copy cosmid, BAC, PAC and P1 in different E. coli strains. In addition, this kit can also be used for high copy plasmid isolation. Two protocols are provided in this handbook for your convenience. The first protocol provides fast and reliable method for purify BAC, PAC, cosmid, P1 and plasmid by using ezBind^TM DNA spin column. The second protocol is for the isolation of low-copy cosmid, BAC, PAC, and P1 using linear polyacrylamide.

Storage and Stability

All EZgene^TM BAC/PAC DNA Isolation Kit components are guaranteed for at least 12 months from the date of purchase when stored as follows: Buffer X1/RNase A mixture at 4℃, and all other materials at 22-25°C.

Before Starting  

Important

•  Add whole vial of RNase A to Buffer X1 and Store at 4℃.
•  Dilute DNA Wash Buffer with absolute ethanol as follows
• PD1311-00 Add 8 mL ethanol to each bottle
• PD1311-01 Add 60 mL ethanol to each bottle
• PD1311-02 Add 96 mL ethanol to each bottle
•  It's strongly recommended to use 2 x YT media for the cultivation of cosmids, BACs, PACs, and P1s.
•  Buffer X2 should be kept at room temperature. Check for SDS precipitation before use. If necessary re-dissolve SDS precipitate by warming. Close the Buffer X2 bottle immediately after use to avoid acidification that may result from air CO₂.
•  Chill Buffer X3 for precipitation enhancement.

Materials Supplied by User

•  Microcentrifuge capable of at least 14,000 x g
•  Sterile Deionized Water (or Elution Buffer)
•  Sterile 1.5 mL and 2.0 mL Centrifuge Tubes
•  10-15 mL Culture Tubes
•  96-100% Ethanol
•  96%-100% Isopropanol
•  70% ethanol

Kit Contents

* Binding Buffer: Add 1.2 mL (PD1311-00) or 16 mL (PD1311-01) or 64 mL (PD1311-02) of isopropanol before use.

**DNA Wash Buffer: Add 8 mL (PD1311-00) or 60 mL (PD1311-01) or 96 mL (PD1311-02) of absolute ethanol before use.

Operating Protocol