Archived | DNA Extraction and Quantitation for Forensic Analysts | Advantages and Disadvantages

Extraction methods are evaluated based on their yield and the quality of results. Early studies using HLA DQ-alpha compared Chelex^® 100 with traditional organic extraction methods, and demonstrated the method's suitability for forensic samples.^01, 02, 04 An advantage of this method is that there are fewer tube manipulations required when compared to traditional organic extraction methods, reducing contamination risks. Automated extraction procedures will diminish this concern even further.

Chelex^® 100 extraction methods are performed in a harsh environment that may not be suitable for highly degraded or low-level DNA samples.⁰⁷ The alkaline environment and high temperatures could further degrade the DNA, which may result in an inability to obtain results.

One study assessed DNA samples that were isolated with Chelex ^®100, stored long-term (approximately 1 year), and subjected to several freeze-thaw cycles. When these samples were characterized using STR amplification, they showed decreased signal, peak imbalance, and allelic drop out. The authors speculated that the unbuffered suspension of the DNA, coupled with multiple freeze-thaw cycles, could accelerate the degradation process of isolated DNA samples.⁰⁸

DNA isolated using the Chelex^® 100 extraction is denatured. This was problematic when laboratories used restriction fragment length polymorphism (RFLP) for characterization since double-stranded DNA is required (RFLP). This is no longer a concern with current STR methods.

The Chelex^® 100 procedure does not include a purification step. If the sample contains inhibitors and contaminants, increasing the sample size increases their concentration, which can inhibit PCR. Some laboratories have incorporated the use of a filter unit such as a Microcon^® or Centricon^® prior to amplification so that some PCR inhibitors can be removed and the sample can be concentrated. Because the resin is a PCR inhibitor, it is important to ensure that when removing the supernatant, no resin is removed from the tube.⁰⁹

**Chelex^®100 Resin Extraction Method**
**Advantages and Disadvantages**
*Advantages*	*Disadvantages*
Fast	Harsh extraction environment (pH between 10-11 and temperature approximately 100° C)
Simple	Potential degradation concerns for long-term storage of isolated DNA samples
Inexpensive	Resin remaining in extracted DNA sample can inhibit PCR process
No hazardous chemicals	Less effective extraction of some sample types
Few tube manipulations	Isolated DNA is single-stranded
Binds heavy metal cations	Resin loses its chelating capacity after a few hours in suspension
Removes some PCR inhibitors

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Date Created: July 6, 2023