Title of Abstract

Comparison of Commercial DNA Extraction Kits for Use on Human Breastmilk

Poster Number

39

Submitting Student(s)

Cassidy Butler
Amy Matsumoto

Session Title

Poster Session 2

Faculty Sponsor (for work done with a non-Winthrop mentor)

Hope Lima, Ph.D., RDN, LRD, IBCLC

College

College of Arts and Sciences

Department

Human Nutrition

Abstract

For-profit donor human milk organizations have DNA-based proprietary methodology for testing incoming milk for purity. However, there is currently no standardized methodology for extracting DNA from human milk. Microbiome research has shown that DNA quality can vary depending on the extraction methodology. This study assessed the quality and quantity of DNA extracted from four commercially available DNA extraction kits – including one kit that was specific to human milk. This study was for method validation only. One donor was utilized to provide a 3-ounce sample. The sample was aliquoted into 70, 1-mL microcentrifuge tubes. Aliquots were randomized into one of three categories: fresh extraction, extraction after freezing, and extraction after purification for storage at room temperature. DNA extraction was performed using four commercially available DNA extraction kits and DNA was analyzed for quality and quantity using a NanoDrop Spectrophotometer. Results confirmed differences in DNA quality and quantity between extraction kits. The Plasma/Serum Circulating DNA Purification Mini Kit (Norgen Biotek, ON, Canada) provided significantly more DNA, consistent purity as measured by 260/280 and 260/230 ratios, and DNA quantity and quality was similar between fresh and frozen human milk samples. Our results suggest that DNA quality and quantity is highest when extracted using the Plasma/Serum Circulating DNA Purification Mini Kit. To ensure reliable quality assurance protocols for testing donor human milk, standardized methodology for extracting DNA from human milk is necessary. The Plasma/Serum Circulating DNA Purification Mini Kit is consistent, providing high quality and sufficient quantity for downstream analysis.

Start Date

15-4-2022 12:00 PM

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Apr 15th, 12:00 PM

Comparison of Commercial DNA Extraction Kits for Use on Human Breastmilk

For-profit donor human milk organizations have DNA-based proprietary methodology for testing incoming milk for purity. However, there is currently no standardized methodology for extracting DNA from human milk. Microbiome research has shown that DNA quality can vary depending on the extraction methodology. This study assessed the quality and quantity of DNA extracted from four commercially available DNA extraction kits – including one kit that was specific to human milk. This study was for method validation only. One donor was utilized to provide a 3-ounce sample. The sample was aliquoted into 70, 1-mL microcentrifuge tubes. Aliquots were randomized into one of three categories: fresh extraction, extraction after freezing, and extraction after purification for storage at room temperature. DNA extraction was performed using four commercially available DNA extraction kits and DNA was analyzed for quality and quantity using a NanoDrop Spectrophotometer. Results confirmed differences in DNA quality and quantity between extraction kits. The Plasma/Serum Circulating DNA Purification Mini Kit (Norgen Biotek, ON, Canada) provided significantly more DNA, consistent purity as measured by 260/280 and 260/230 ratios, and DNA quantity and quality was similar between fresh and frozen human milk samples. Our results suggest that DNA quality and quantity is highest when extracted using the Plasma/Serum Circulating DNA Purification Mini Kit. To ensure reliable quality assurance protocols for testing donor human milk, standardized methodology for extracting DNA from human milk is necessary. The Plasma/Serum Circulating DNA Purification Mini Kit is consistent, providing high quality and sufficient quantity for downstream analysis.