New protocol for DNA extraction of stool.
Present methods for DNA isolation of stool have various limitations such as the amount of stool used, the requirement of lavage fluids or the use of fresh stool. In this paper, a new method is described for the isolation of human nucleic acids from stool, which is independent from the moment of collection. Fecal samples as dry as possible were collected from 75 patients; two grams of stool were mixed with a lysis buffer containing phenol. DNA yields of crude stool were variable and ranged from 9-1686 micrograms/g of feces. With dot blots in 9 of the 75 cases, the human DNA was identified and ranged from 0.06%-46%. In the remaining 66 cases, human genomic DNA was detected by nested PCR, using human K-ras gene amplification as an example. Amplification products were confirmed for human K-ras with the exonuclease-amplification coupled capture technique (EXACCT). In conclusion, the developed DNA isolation method can be used for the study of large numbers of stool samples, is independent of the age or method of stool collection and is suitable for large-scale screening studies.
Machiels
B M
BM
Maastricht University, The Netherlands.
Ruers
T
T
Lindhout
M
M
Hardy
K
K
Hlavaty
T
T
Bang
D D
DD
Somers
V A
VA
Baeten
C
C
von Meyenfeldt
M
M
Thunnissen
F B
FB
D001483:Base Sequence; D001709:Biotechnology; D015179:Colorectal Neoplasms / Q000175:diagnosis* / Q000235:genetics; D017931:DNA Primers / Q000235:genetics; D004273:DNA, Neoplasm / Q000235:genetics / Q000302:isolation & purification*; D005243:Feces / Q000737:chemistry* / Q000166:cytology; D006801:Humans; D015336:Molecular Probe Techniques; D016133:Polymerase Chain Reaction
D017931:DNA Primers; D004273:DNA, Neoplasm