Serum, plasma and paraffin-embedded tissues as sources of DNA for studying cancer susceptibility genes

• Published in: Carcinogenesis (New York) 1997-06, Vol.18 (6), p.1271-1275
• Main Authors: BLÖMEKE, B, BENNETT, W. P, HARRIS, C. C, SHIELDS, P. G
• Format: Article
• Language: English
• Subjects: Arylamine N-Acetyltransferase - genetics; Arylamine N-Acetyltransferase - metabolism; Biological and medical sciences; Biotransformation; Carcinogens - pharmacokinetics; Cytochrome P-450 CYP1A1 - genetics; Cytochrome P-450 CYP1A1 - metabolism; Cytochrome P-450 CYP2E1 - genetics; Cytochrome P-450 CYP2E1 - metabolism; Diverse techniques; DNA - blood; DNA - isolation & purification; DNA, Neoplasm - blood; DNA, Neoplasm - isolation & purification; Exons; Fundamental and applied biological sciences. Psychology; Genetic Predisposition to Disease; Glutathione Transferase - genetics; Glutathione Transferase - metabolism; Humans; Inactivation, Metabolic; Isoenzymes - genetics; Isoenzymes - metabolism; Lung - chemistry; Lung - enzymology; Lung - physiology; Lung Neoplasms - chemistry; Lung Neoplasms - enzymology; Lung Neoplasms - genetics; Molecular and cellular biology; Oncogenes; Paraffin Embedding; Polymerase Chain Reaction - methods; Polymorphism, Genetic; Reproducibility of Results
• ISSN: 0143-3334; 1460-2180; 1460-2180
• DOI: 10.1093/carcin/18.6.1271

The ability to isolate DNA from archived human serum, plasma and paraffin-embedded human tissues enhances opportunities to study breast, lung and other cancer risk factors. We report herein a simple and fast protocol for the extraction of genomic DNA from these sources. Using a phenol-based extraction method, the recovery for DNA is quantitative and reproducible. DNA yields in serum (250 microl) were between 162 and 1060 ng (n = 18 subjects), in plasma (250 microl) were between 165 and 375 ng (n = 5 subjects) and in embedded tissues (5-microm thick sections for ethanol fixed, and between 5- and 20-microm sections for formaldehyde fixation) were between 1 microg and 11.7 microg (n = 32 subjects). The extraction method was combined with newly designed PCR-based assays for cancer susceptibility marker genes such as CYP1A1 (exon 7), CYP2E1 (Dra1, Rsa1), GSTM1 and NAT2 [NAT2*5A (C481T), NAT2*6A (G590A), NAT2*7A (G857A)]. Genotyping results from the serum and paraffin-embedded tissues compared favorably to results from archived freshly frozen tissues, where concordance was 98% for serum, 100% for ethanol-fixed embedded tissues, and 97% for formaldehyde-fixed and paraffin-embedded tissues. This facile method will allow for the use of archived tissue samples of prospective cohort and other studies where intact DNA was not previously available.