Abstract
Background: The low yield and quality of buccal-derived genomic DNA have reduced its applicability in various genetic research. The aim of this study was to assess the quantity, purity and genotyping efficiency of genomic DNA isolated from neonatal buccal swabs. Methods: Paired buccal swabs and whole blood samples were collected from 60 neonates with the mean age 5 days (SD=1.57). The genomic DNA quantity and purity were measured by using Infinite® 200 PRO NanoQuant reader and agarose gel electrophoresis. High-resolution melting (HRM) analysis was used to analyse the sequence variants present in uridine diphosphate glucuronosyltransferase 1A1 (UGT1A1 c.211G>A) and nuclear receptor subfamily 1, group I, member 3 (NR1I3 IVS8+116T>G) genes. Results: Buccal swabs provided lower mean genomic DNA concentration (18.78 ± 8.39 ng/µl versus 40.02 ± 13.03 ng/µl), yield (2.63 ± 1.17 µg versus 8.00 ± 2.61 µg). The purity of buccal samples however were inconsistent with 16 samples (26.7%) having A260/280 ratios below 1.8 which indicated protein contamination. Genomic DNA purity for all blood samples were within the ideal range with average absorbance ratios of 1.8−2.0. However, all buccal genomic DNA demonstrated 100% genotype call rates for all variants. A complete genotype concordance was also observed between paired genomic DNA samples. Conclusion: Despite related to a reduced quantity and purity, neonatal buccal genomic DNA could generate reliable HRM genotyping results. Therefore, buccal swab collection is a promising alternative to the invasive blood sampling to provide genomic DNA for genetic analysis involving paediatric population.
Highlights
The low yield and quality of buccal-derived genomic DNA have reduced its applicability in various genetic research
Quantity and purity of buccal and blood genomic DNA Buccal genomic DNA was isolated from 60 neonates and compared with blood samples for the genomic DNA yield and quality, and High-resolution melting (HRM) genotyping efficiency
Buccal genomic DNA concentration of as low as 4.36 ng/μl was isolated while the minimum blood genomic DNA concentration was 15.38 ng/μl
Summary
The low yield and quality of buccal-derived genomic DNA have reduced its applicability in various genetic research. The aim of this study was to assess the quantity, purity and genotyping efficiency of genomic DNA isolated from neonatal buccal swabs. Methods: Paired buccal swabs and whole blood samples were collected from 60 neonates with the mean age 5 days (SD=1.57). Results: Buccal swabs provided lower mean genomic DNA concentration (18.78 ± 8.39 ng/μl versus 40.02 ± 13.03 ng/μl), yield (2.63 ± 1.17 μg versus 8.00 ± 2.61 μg). All buccal genomic DNA demonstrated 100% genotype call rates for all variants. Conclusion: Despite related to a reduced quantity and purity, neonatal buccal genomic DNA could generate reliable HRM genotyping results. Buccal swab collection is a promising alternative to the invasive blood sampling to provide genomic DNA for genetic analysis involving paediatric population
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