Improved amplification efficiency on stool samples by addition of spermidine and its use for non-invasive detection of colorectal cancer

Handle URI:
http://hdl.handle.net/10754/556146
Title:
Improved amplification efficiency on stool samples by addition of spermidine and its use for non-invasive detection of colorectal cancer
Authors:
Roperch, Jean-Pierre; Benzekri, Karim; Mansour, Hicham; Incitti, Roberto
Abstract:
Background Using quantitative methylation-specific PCR (QM-MSP) is a promising method for colorectal cancer (CRC) diagnosis from stool samples. Difficulty in eliminating PCR inhibitors of this body fluid has been extensively reported. Here, spermidine is presented as PCR facilitator for the detection of stool DNA methylation biomarkers using QM-MSP. We examined its effectiveness with NPY, PENK and WIF1, three biomarkers which we have previously shown to be of relevance to CRC. Results We determined an optimal window for the amplification of the albumin (Alb) gene (100 ng of bisulfite-treated stool DNA added of 1 mM spermidine) at which we report that spermidine acts as a PCR facilitator (AE = 1680%) for SG RT-PCR. We show that the amplification of methylated PENK, NPY and WIF1 is considerably facilitated by QM-MSP as measured by an increase of CMI (Cumulative Methylation Index, i.e. the sum of the three methylation values) by a factor of 1.5 to 23 fold in individual samples, and of 10 fold in a pool of five samples. Conclusions We contend that spermidine greatly reduces the problems of PCR inhibition in stool samples. This observed feature, after validation on a larger sampling, could be used in the development of stool-based CRC diagnosis tests.
KAUST Department:
Biosciences Core Lab; Computational Bioscience Research Center (CBRC)
Citation:
Improved amplification efficiency on stool samples by addition of spermidine and its use for non-invasive detection of colorectal cancer 2015, 15 (1) BMC Biotechnology
Publisher:
Springer Science + Business Media
Journal:
BMC Biotechnology
Issue Date:
29-May-2015
DOI:
10.1186/s12896-015-0148-6
PubMed ID:
26022272
PubMed Central ID:
PMC4446959
Type:
Article
ISSN:
1472-6750
Additional Links:
http://www.biomedcentral.com/1472-6750/15/41
Appears in Collections:
Articles; Biosciences Core Lab; Computational Bioscience Research Center (CBRC)

Full metadata record

DC FieldValue Language
dc.contributor.authorRoperch, Jean-Pierreen
dc.contributor.authorBenzekri, Karimen
dc.contributor.authorMansour, Hichamen
dc.contributor.authorIncitti, Robertoen
dc.date.accessioned2015-06-02T14:01:10Zen
dc.date.available2015-06-02T14:01:10Zen
dc.date.issued2015-05-29en
dc.identifier.citationImproved amplification efficiency on stool samples by addition of spermidine and its use for non-invasive detection of colorectal cancer 2015, 15 (1) BMC Biotechnologyen
dc.identifier.issn1472-6750en
dc.identifier.pmid26022272en
dc.identifier.doi10.1186/s12896-015-0148-6en
dc.identifier.urihttp://hdl.handle.net/10754/556146en
dc.description.abstractBackground Using quantitative methylation-specific PCR (QM-MSP) is a promising method for colorectal cancer (CRC) diagnosis from stool samples. Difficulty in eliminating PCR inhibitors of this body fluid has been extensively reported. Here, spermidine is presented as PCR facilitator for the detection of stool DNA methylation biomarkers using QM-MSP. We examined its effectiveness with NPY, PENK and WIF1, three biomarkers which we have previously shown to be of relevance to CRC. Results We determined an optimal window for the amplification of the albumin (Alb) gene (100 ng of bisulfite-treated stool DNA added of 1 mM spermidine) at which we report that spermidine acts as a PCR facilitator (AE = 1680%) for SG RT-PCR. We show that the amplification of methylated PENK, NPY and WIF1 is considerably facilitated by QM-MSP as measured by an increase of CMI (Cumulative Methylation Index, i.e. the sum of the three methylation values) by a factor of 1.5 to 23 fold in individual samples, and of 10 fold in a pool of five samples. Conclusions We contend that spermidine greatly reduces the problems of PCR inhibition in stool samples. This observed feature, after validation on a larger sampling, could be used in the development of stool-based CRC diagnosis tests.en
dc.publisherSpringer Science + Business Mediaen
dc.relation.urlhttp://www.biomedcentral.com/1472-6750/15/41en
dc.rightsThis is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.en
dc.subjectMethylated biomarkersen
dc.subjectQM-MSPen
dc.subjectColorectal canceren
dc.subjectSpermidineen
dc.titleImproved amplification efficiency on stool samples by addition of spermidine and its use for non-invasive detection of colorectal canceren
dc.typeArticleen
dc.contributor.departmentBiosciences Core Laben
dc.contributor.departmentComputational Bioscience Research Center (CBRC)en
dc.identifier.journalBMC Biotechnologyen
dc.identifier.pmcidPMC4446959en
dc.eprint.versionPublisher's Version/PDFen
dc.contributor.institutionProfilome, Paris Biotech 24 rue du Faubourg St Jacques, Paris 75014, Franceen
dc.contributor.institutionOncoDiag, Agoranov 96 Bis, Boulevard Raspail, Paris 75006, Franceen
dc.contributor.institutionCentre d’Investigation Clinique (CIC), Henri Mondor Hospital, Créteil, Franceen
kaust.authorMansour, Hichamen
kaust.authorIncitti, Robertoen

Related articles on PubMed

All Items in KAUST are protected by copyright, with all rights reserved, unless otherwise indicated.