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GENOMIC DNA PREP KIT for ANIMAL & PLANT

ORDERING INFORMATION

Catalog #

Product

Amount

Price (EUR)

# GEN 50 AP
Genomic DNA Prep Kit for Animals & Plants
50 preparations


# GEN 250 AP
Genomic DNA Prep Kit for Animals & Plants

250 preparations


PRODUCT COMPONENTS

Buffer

# GEN 50 AP

# GEN 250 AP

Animal-Plant Lysis Buffer

20 ml

100 ml

Binding Buffer

35 ml

175 ml

Proteinase K (final conc. 10 mg/ml)

5 mg

5 x 5 mg

Column Activation Solution

6 ml

30 ml

Elution Buffer

5 ml

25 ml

Spin Columns & 2ml Tubes

50

250

DESCRIPTION

The spin column based Animal-Plant DNA Preparation Kit is designed for rapid and high purity isolation of genomic DNA from animal cells and plant tissue. The spin column based method completely removes PCR inhibitors such as divalent cations and proteins resulting in a high purity preparation of genomic DNA. There is no use of phenol or chloroform, handling is safe and does not produce any harmful waste. Column based genomic DNA purification kits yield up to 30 μg DNA sized from 200 bp to 50 kb per preparation.

FEATURES
  • Spin column type
  • Purification size: 200 bp - 50 kb
  • DNA amounts up to 30 g

APPLICATIONS

  • real-time PCR
  • PCR
  • southern blot analysis
  • genotyping
  • discovery or validation of SNP/SSR marker

BASIC PROTOCOL

Before start, prepare the following components:

  • Add double distilled water to Proteinase K. Proteinase K solution should be stored at -20C.
    • for 50 preparations (# GEN 50 AP): add 500 l dd-water to 5 mg Proteinase K.
    • for 250 preparations (# GEN 250 AP): add 500 l dd-water to each tube of 5 mg Proteinase K.
  • Prepare an 80% Ethanol (V/V) solution as Washing buffer. Please note that the Ethanol concentration of Washing Buffer may decrease during long time storage resulting in a possible drop of the final DNA yield.
    • for 50 preparations (# GEN 50 AP): 60 ml
    • for 250 preparations (# GEN 250 AP): 300 ml
A. DNA PREPARATION FROM ANIMAL CELLS

It is essential to use the correct amount of starting material in order to obtain optimal DNA yield and purity. A maximum amount of 80 mg animal material can generally be processed. We recommend starting with not more than 80 mg fresh or frozen animal tissue. Tissues can be stored at -70C for several months.

Fresh-freeze tissues in liquid nitrogen and grind it thoroughly with a mortar and pestle. Transfer tissue powder into a 1,5 ml microcentrifuge tube cooled with liquid nitrogen.

1. Cell Lysis

  • Add 350 l Animal-Plant Lysis Buffer to the tissue powder.
  • Vortex vigorously for 30-60 sec.
  • Add 8 l (10 mg/ml) Proteinase K solution to the tissue lysate.
  • Mix by pipetting.
  • Incubate at 70C for 10 min and cool down for 5 min.
  • Centrifuge for 5 min at 12,000 g at room temperature.
  • Transfer supernatant to new 1.5 ml microcentrifuge tube.

2. Adding Binding Buffer

  • Add 400 l Binding Buffer to the tissue lysate.
  • Vortex vigorously for 30-60 sec.
  • Centrifuge for 5 min at 12,000 g at room temperature.

3. Column Activation

  • Place a Spin Column into a 2 ml collection tube.
  • Add 100 l of Column Activation Solution into the Spin Column.
  • Centrifuge at 10,000 g for 30 sec in a microcentrifuge.

4. Column Loading

  • Pipette the lysate directly into the Spin Column.
  • Centrifuge for 1 min at 12,000 g.
  • Discard the flow-through.

5. Primary Washing

  • Add 500 l Washing Buffer (80% Ethanol) into the Spin Column.
  • Centrifuge for 30 sec at 12,000 g.
  • Discard the flow-through.
6. Secondary Washing
  • Add 500 l Washing Buffer into the Spin Column.
  • Centrifuge for 30 sec at 12,000 g.
  • Discard the flow-through.
  • Centrifuge again at 12,000 g for 1 min to remove residual Washing Buffer.
  • Discard the 2 ml wash tube and place the column in the elution tube.

7. Elution of DNA

  • Add 40-50 μl Elution Buffer to the center of the column.
  • Incubate at room temperature for 1 min.
  • Centrifuge at 12,000 g for 2 min to elute DNA.
  • Store DNA at 4C (for prolonged storage, keep it at -20C or -80C).

B. DNA PREPARATION FROM PLANT TISSUE

It is essential to use the correct amount of starting material in order to obtain optimal DNA yield and purity. A maximum amount of 80 mg plant material can generally be processed. We recommend starting with not more than 80 mg fresh or frozen plant tissue. Tissues can be stored at -70C for several months.

Fresh-freeze tissues in liquid nitrogen and grind it thoroughly with a mortar and pestle. Transfer tissue powder into a 1,5 ml microcentrifuge tube cooled with liquid nitrogen.

1. Cell Lysis

  • Add 250 l Animal-Plant Lysis Buffer to the tissue powder.
  • Vortex vigorously for 30-60 sec.
  • Add 8 l (10 mg/ml) Proteinase K to the tissue lysate.
  • Mix by pipetting.
  • Incubate at 70C for 10 min and cool down for 5 min.
  • Centrifuge for 5 min at 12,000 g at room temperature.
  • Transfer supernatant to new 1.5 ml microcentrifuge tube.

2. Adding Binding Buffer

  • Add 250 l Binding Buffer to the tissue lysate.
  • Vortex vigorously for 30-60 sec.
  • Add 8 l (10 mg/ml) Proteinase K solution to the tissue lysate.
  • Mix by pipetting.
  • Incubate at 70C for 10 min and cool down for 5 min.
  • Centrifuge for 5 min at 12,000 g at room temperature.

3. Column Activation

  • Place a Spin Column into a 2 ml collection tube.
  • Add 100 l of Column Activation Solution into the Spin Column.
  • Centrifuge at 10,000 g for 30 sec in a microcentrifuge.

4. Column Loading

  • Pipette the lysate directly into the Spin Column.
  • Centrifuge for 1 min at 12,000 g.
  • Discard the flow-through.

5. Primary Washing

  • Add 500 l Washing Buffer (80% Ethanol) into the Spin Column.
  • Centrifuge for 30 sec at 12,000 g.
  • Discard the flow-through.
6. Secondary Washing
  • Add 500 l Washing Buffer into the Spin Column.
  • Centrifuge for 30 sec at 12,000 g.
  • Discard the flow-through.
  • Centrifuge again at 12,000 g for 1 min to remove residual Washing Buffer.
  • Discard the 2 ml wash tube and place the column in the elution tube.

7. Elution of DNA

  • Add 40-50 μl Elution Buffer to the center of the column.
  • Incubate at room temperature for 1 min.
  • Centrifuge at 12,000 g for 2 min to elute DNA.
  • Store DNA at 4C (for prolonged storage, keep it at -20C or -80C).

STORAGE

Store at room temperature (15 - 25 C) for 12 months.

Proteinase K solution should be stored at -20C.

For long term storage place Proteinase K lyophilisate at -20C.