Summary
Conclusion: Preparation of DNA from pancreatic juice for subsequent polymerase chain reaction (PCR) is difficult, but manageable. The protocol presented offers a simple and fast solution. This method might be applicable to other complicated samples, such as saliva, wound secretions, or stool washings.
Background: Of all the biological samples used for PCR amplification, pancreatic juice is the most problematic because of the presence of potential inhibitory substances and the amount of nucleases. This demands a DNA preparation procedure that is suitable for routine diagnostic PCR, and is therefore efficient and safe. This is particularly true for pancreatic juice obtained during routine endoscopy.
Methods: We describe here a simple method utilizing modified phenol/chloroform extraction and precipitation directly from native pancreatic juice suitable for diagnostic PCR applications, such as oncogenes.
Results: DNA could be prepared in quantitative amounts from routine endoscopic specimens. DNA could also be prepared from samples kept several days at room temperature.
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References
Löhr M, Brenner DA, Rooney JF, Nelson JA. Application of the polymerase chain reaction in gastrointestinal endoscopy. Endoscopy 1992;24: 779–781.
Hruban RH, van Offershaus GJ, Allison DC, et al. k-ras oncogene activation in adenocarcinoma of the human pancreas. A study of 82 carcinomas using a combination of mutant-enriched polymerase chain reaction analysis and allele-specific oligonucleotide hybridization. Am J Pathol 1993; 143: 545–554.
Kondo H, Sugano K, Fukayama N, et al. Detection of point mutations in the k-ras oncogene at codon 12 in pure pancreatic juice for diagnosis of pancreatic carcinoma. Cancer 1994; 73: 1589–1594.
Heller T, Trautmann B, Zöller-Utz I, et al. Restriktionsenzym-Mismatch-Polymerase-Kettenreaktion zum Nachweis von ki-ras-Onkogen -Mutationen beim Pankreaskarzinom. Dtsch Med Wchnschr 1995; 120: 826–830.
DiMagno EP. Human exocrine pancreatic enzyme secretion, in The Exocrine Pancreas: Biology, Pathobiology, and Diseases. Go VLW, ed. Raven, New York, 1986; pp. 193–210.
Hattfield ARW, Smithies A, Wikins R, Levi AJ. Assessment of ERCP and pure pancreatic juice cytology in patients with pancreatic disease. Gut 1976; 17: 14–21.
Uehara H, Nakaizumi A, Iishi H, et al. Cytologic examination of pancreatic juice for differential diagnosis of benign and malignant mucin-producing tumors of the pancreas. Cancer 1994; 74: 826–833.
Sparmann G, Jäschke A, Müller P, Löhr M, Liebe S, Emmrich J. Tissue homogenization as a key step extracting RNA from human and rat pancreatic tissue. Biotechniques 1997; 22: 408–412.
Rolfs A, Schuller I, Finckh U, Weber-Rolfs I. PCR. Clinical Diagnostics and Research. Springer, Heidelberg, 1993.
Maniatis T, Fritsch EF, Sambrook J. Molecular Cloning. A Laboratory Manual. Cold Spring Harbor Laboratory, Cold Spring Harbor, NY, 1982.
Löhr M, Oldstone MBA. Detection of cytomegalovirus nucleic acid sequences in pancreas in type 2 diabetes. Lancet 1990; 336: 644–648.
Müller P, Heller T, Stelt HJ, et al. Welche Bedeutung haben ras- und p53-Mutationen im Pankreassekret bei Patienten mit chronischer Pankreatitis? Z Gastroenterol 1996; 34: 554/V5 (abstract).
Haliassos A, Chomel JC, Tesson L, et al. Modification of enzymatically amplified DNA for the detection of point mutations. Nucleic Acid Res 1989; 17: 3606.
Trautmann B, Schlitt HJ, Hahn EG, Löhr M. Isolation, culture, and characterization of human pancreatic duct cells. Pancreas 1993; 8: 248–254.
Löhr M, Trautmann B, Göttler M, et al. Human ductal adenocarcinomas of the pancreas express extracellular matrix proteins. Br J Cancer 1994; 69: 144–151.
Belec L, Authier J, Eliezer-Vanerot MC, Piedouillet C, Mohamed AS, Gherardi RK. Myoglobin as a polymerase chain reaction (PCR) inhibitor: a limitation for PCR from skeletal muscle tissue avoided by the use of Thermus thermophilus polymerase. Muscle Nerve 1998; 21: 1064–1067.
Behzadbehbahani A, Klapper PE, Vallely PJ, Cleator GM. Detection of BK virus in urine by polymerase chain reaction: a comparison of DNA extraction methods. J Virol Methods 1997; 67: 161–166.
Wiedbrauk DL, Werner JC, Drevon AM. Inhibition of PCR by aqueous and vitreous fluids. J Clin Microbiol 1995; 33: 2643–2646.
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Müller, P., Jesnowski, R., Liebe, S. et al. Simple method for DNA extraction from pancreatic juice for PCR amplification assays. International Journal of Pancreatology 25, 39–43 (1999). https://doi.org/10.1385/IJGC:25:1:39
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DOI: https://doi.org/10.1385/IJGC:25:1:39