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Malin I L Ivarsson, Joakim Dillner, Joyce Carlson, Validity of Maternal Genotypes in DNA from Archival Pregnancy Serum Samples, Clinical Chemistry, Volume 55, Issue 4, 1 April 2009, Pages 842–843, https://doi.org/10.1373/clinchem.2008.116277
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To the Editor:
Much functional genomic research has focused on genotypic data from large cohorts or entire populations(1). The large population-based serum biobanks collected for maternal (pregnancy) screening and stored in many countries could be useful for genetic epidemiologic studies if DNA extracted from archival serum and plasma is of sufficient quantity and quality for successful genotyping analyses(2)(3)(4). Serum collected during pregnancy also contains cell-free DNA from the fetus(5), however, which might affect the outcome of genotyping analyses. We have evaluated the concordance of genotypes between DNA extracted from archival maternal sera with DNA from fresh whole blood from the same women.
The median DNA yield was 15 mg/L (range 1–34 mg/L) using QiaAmp DNA minipreps (Qiagen) on 200 μL fresh whole blood from 137 women and 90 μg/L (range 0–4800 μg/L) using the MagNA Pure LC and Total Nucleic Acid Isolation Kit (MagNAPure; Roche Diagnostics) on 200 μL of 191 archival maternal sera from the same women. DNA yield decreased with increasing serum sample age (P = 0.005, Jonckheere–Terpstra) (Fig. 1 ), with no difference due to first- or second-trimester sampling (P = 0.616, Mann–Whitney). Sample volumes were sufficient for repeat DNA extractions from 175 serum samples. Twenty microliters from each 50-μL extract was evaporated to dryness at 37 °C and used in a 6-μL Y chromosome–specific PCR(5) with 0.0625 μL 40× assay mix, 0.1% BSA (Sigma-Aldrich), and 3.5 mol/L MgCl2 on a 7900 HT Sequence Detection System (Applied Biosystems). Another evaporated 20-μL aliquot was used for multiple displacement amplification (MDA)1 (GenomiPhi V2; GE healthcare) with 0.1% BSA. MDA products were dissolved in 80 μL TE (10 mmol/L Tris, 1 mmol/L EDTA, pH 8.0) by shaking at 4 °C overnight. DNA concentrations were determined using 2 μL template in a F2 g.20210G>A (rs1799963) TaqMan MGB assay, with serum samples supplemented with BSA and MgCl2 as above. All samples were analyzed for 10 high-frequency single nucleotide polymorphisms (SNPs) using 2 μL whole blood extract, 9 μL serum extract, or 2 μL MDA product, 0.075 μL assay mix, BSA, and MgCl2 as above. Seventy-two of 84 serum samples from women carrying male fetuses and 1 of 91 samples from women carrying female fetuses were Y chromosome positive (P = 8E−30). There was no correlation between sample age, gestational age (P = 0.2–0.9, all Pearson χ2), or DNA yield (P = 0.9, Mann–Whitney), and Y chromosome detection.
