• Efficient recovery of trace DNA (100-1,000 fg）
• Perform in a single tube for complete the process
• Extraction completed in 60-90 minutes
• Pretreatment protocol for samples containing high protein concentration
• Adopts Sodium Iodide method*

*Sodium Iodide method is a residual DNA extraction technique that is described in The United States Pharmacopeia (USP) 42-NF37, <509> Residual DNA Testing.

Principle of DNA extraction

i. Cell membrane and cytoplasm are destroyed by surfactant, isolating cell nuclei.
ii. The nuclear membrane and nuclear protein are decomposed by protease to expose DNA.
iii. Protein and lipids are made soluble by the action of sodium iodide, and DNA is precipitated with isopropanol.

Extraction of viral DNA in serum and an infinitesimal amount of fungus-derived DNA in biological products

• Extraction from serum

  

  Operation time: 1 to 1.5 hours
  Sample amount: 100 μL/test

• This kit is intended for use in extracting host cell-derived residual DNA remaining in biological components using the Sodium Iodide method. Extracted DNA can be quantified by qPCR. Use the DNA Extractor® kit for testing and management of the amount of DNA derived from host cells such as CHO cells, Escherichia coli, and yeast. DNA from any species extracted using this kit is suitable for using the Threshold Immunoassay Syste^® provided by Molecular Devices, LLC. The kit can also be used for DNA extraction from serum.

Recovery test of CHO cell-derived DNA

Purpose:
To examine the yields of CHO cell-derived DNA from culture supernatant.

Methods:
Using the kit, DNA was extracted from culture supernatants of PANC-1 cells to which 10 fg to 1 ng of CHO cell-derived DNA was added. A qPCR assay was then performed using the extracted DNA to obtain Cq values.
A qPCR was also performed using purified water to which CHO cell-derived DNA was added, without DNA extraction (standard conditions), to obtain Cq values. The DNA yield under each set of conditions was calculated based on a calibration curve generated from the results of the standard conditions.

Samples:
1. Purified water containing CHO cell-derived DNA (no DNA extraction): Standard conditions
2. DNA extracted from the culture supernatants of PANC-1 cells containing CHO cell-derived DNA, using the kit.

qPCR reagents:
・GeneAce SYBR^® qPCR Mix α No ROX (Nippon Gene#319-07703)
・Optical Flat 8–Cap Strips for 0.2 mL tube strips/Plates (BIO-RAD#TCS0803)
・Hard-Shell^® Thin-wall 96-well PCR plates (BIO-RAD#HSP9601)

Results:

ND: Not detected

DNA added to culture supernatants (100 fg to 1 ng) was extracted with high yield.

This test was performed in accordance with Protocol #2 described in the manual.

Recovery test of E. coli-derived DNA

Purpose:
To examine the yields of E. coli-derived DNA.

Methods:
E. coli-derived DNA (10 fg to 1 ng) was added to purified water, and Cq values were measured before and after DNA extraction using the kit. Calibration curve was generated from mean Cq values of samples before the DNA extraction to calculate the yields after the extraction.

Samples:
Purified water containing E. coli-derived DNA

qPCR reagents:
・GeneAce SYBR^® qPCR Mix α No ROX (Nippon Gene#319-07703)
・Optical Flat 8–Cap Strips for 0.2 mL tube strips/Plates (BIO-RAD#TCS0803)
・Hard-Shell® thin-wall 96-well PCR plates (BIO-RAD#HSP9601)

Results:

E. coli-derived DNA added to the samples (1 pg to 1 ng) was extracted with high yield.

This test was performed in accordance with Protocol #1 described in the manual.

Examples of using DNA Extractor^® Kit

Reference

Ishizawa, M. et al., Nucleic Acids Res., 19, 5792 (1991).