Promega eNotes - Applications

Wizard^Ž Genomic DNA Purification Kit and the Isolation of Plant Genomic DNA

Genomic DNA was isolated from plant leaf tissue using the Wizard^Ž Genomic DNA Isolation Kit (Cat.# A1120). Isolated DNA is suitable for PCR and restriction enzyme digestions.

By Sonja Vorwerk, B.S.
Max Planck Institute for Plant Breeding Research, Cologne, Germany

Introduction

The Wizard^Ž Genomic DNA Purification Kit (Cat.# A1120) provides a simple and phenol-free method for the isolation of genomic DNA from different sources including bacteria, yeast, blood, and animal and plant tissues. The whole procedure can be finished in as little as two hours and the DNA obtained is suitable for many downstream applications like PCR and restriction enzyme digestion. For example, as little as 40mg of tomato leaf usually yields 7-12ľg genomic DNA. However, if more plant material is available the amount of starting material can easily be scaled up without increasing the solutions or most of the incubation times as reported here. Both monocotyledon (corn) and dicotyledon (Arabidopsis) species were tested.

Leaf material from each species was frozen in liquid nitrogen and ground extensively (approximately 10 minutes). Ground tissue (40-250mg) was transferred to a microcentrifuge tube and processed according to the Wizard^Ž Genomic DNA Purification Kit Technical Manual (#TM050) (1). There is no need to prolong the incubation times except for the RNase Solution treatment in Step 4, Section III.E. To completely remove RNA, it is better to incubate for 30 minutes instead of 15 minutes at 37°C.

Yields for Arabidopsis and corn from 40, 100, 150 and 250mg starting tissue are listed in Table 1. DNA was of high quality, with A₂₆₀/A₂₈₀nm ratios between 1.8 and 1.95. As shown in Figures 1 and 2, the DNA is suitable for PCR^(a) (Figure 1) and restriction enzyme digestion with BstX I (Figure 2). Therefore, the genomic DNA is free of contaminants and enzyme inhibitors that are often copurified when conventional methods are used.

Table 1. Yield of Genomic DNA from Different Starting Amounts of Leaf Tissue. Values represent mean (+/-S.D.)

Starting
Amount

Arabidopsis (ľg)

Maize (ľg)

40mg

8.33 (+/-1.15)

7.67 (+/-0.58)

100mg

43.33 (+/-2.52)

40.67 (+/-6.66)

150mg

49.83 (+/-1.04)

47.17 (+/-2.75)

250mg

59.67 (+/-7.57)

60.00 (+/-4.00)

Reference

Wizard^Ž Genomic DNA Purification Kit Technical Manual #TM050, Promega Corporation.

^(a)The PCR process is covered by patents issued and applicable in certain countries. Promega does not encourage or support the unauthorized or unlicensed use of the PCR process.

*Products may be covered by pending or issued patents. Please visit our patent and trademark web page for more information.

Wizard is a trademark of Promega Corporation and is registered with the U.S. Patent and Trademark Office.

Figure 1. PCR of genomic DNA following isolation from Arabidopsis and corn.

Figure 2. Restriction enzyme digestion of genomic DNA following isolation from Arabidopsis and corn.

Focus: Genomic DNA Isolation from Plants

WizardŽ Genomic DNA Purification Kit and the Isolation of Plant Genomic DNA

By Sonja Vorwerk, B.S. Max Planck Institute for Plant Breeding Research, Cologne, Germany

(a)The PCR process is covered by patents issued and applicable in certain countries. Promega does not encourage or support the unauthorized or unlicensed use of the PCR process.

*Products may be covered by pending or issued patents. Please visit our patent and trademark web page for more information.

Wizard is a trademark of Promega Corporation and is registered with the U.S. Patent and Trademark Office.

Wizard^Ž Genomic DNA Purification Kit and the Isolation of Plant Genomic DNA

By Sonja Vorwerk, B.S.
Max Planck Institute for Plant Breeding Research, Cologne, Germany

^(a)The PCR process is covered by patents issued and applicable in certain countries. Promega does not encourage or support the unauthorized or unlicensed use of the PCR process.