Analysis of multiple single nucleotide polymorphisms (SNP) on DNA traces from plasma and dried blood samples.
Reliable analysis of single nucleotide polymorphisms (SNPs) in DNA derived from samples containing low numbers of cells or from suboptimal sources can be difficult. A new procedure to characterize multiple SNPs in traces of DNA from plasma and old dried blood samples was developed. Six SNPs in the Mannose Binding Lectin 2 (MBL2) gene were chosen as targets for analysis. DNA was extracted from plasma obtained from mothers (n=49) and their neonates (n=49) and from old dried blood samples (n=204). Multiple Real-Time SNP analyses in the MBL2 gene were carried out on all samples. Because of very low DNA concentrations in most of the samples, a pre-amplification step was utilized. It was possible to analyze all plasma samples (n=98), including those with very low cell numbers (n=21) and 93% of the old dried blood samples (n=189). Results obtained from pre-amplified samples were in full agreement with neat samples. All possible SNP alleles were present in our population. The frequencies of the different alleles from both plasma and dried blood samples (n=287) were in agreement with earlier studies of the Caucasian population. In conclusion, amplification prior to Real-Time PCR SNP analysis is a convenient, cost effective and useful method to significantly improve the reliable SNP detection in specimens containing very low concentrations or poor quality DNA from suboptimal sources.
Catsburg
Arnold
A
Department of Medical Microbiology and Infection Control, VU University Medical Center, Amsterdam, The Netherlands.
van der Zwet
Wil C
WC
Morré
Servaas A
SA
Ouburg
Sander
S
Vandenbroucke-Grauls
Christina M J E
CM
Savelkoul
Paul H M
PH
D000328:Adult; D001800:Blood Specimen Collection / Q000379:methods*; D004247:DNA / Q000097:blood*; D005260:Female; D005787:Gene Frequency; D005838:Genotype; D006801:Humans; D007231:Infant, Newborn; D037601:Mannose-Binding Lectin / Q000235:genetics*; D009426:Netherlands; D021141:Nucleic Acid Amplification Techniques; D016133:Polymerase Chain Reaction / Q000379:methods*; D020641:Polymorphism, Single Nucleotide; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
C445619:MBL2 protein, human; D037601:Mannose-Binding Lectin; D004247:DNA