Real-Time quantitative polymerase chain reaction analysis of patients with refractory chronic periodontitis

Simone Marconcini, Ugo Covani, Antonio Barone, Orazio Vittorio, Michele Curcio, Serena Barbuti, Fabrizio Scatena, Lamberto Felli, Claudio Nicolini

Research output: Contribution to journalArticle

7 Citations (Scopus)

Abstract

Background: Periodontitis is a complexmultifactorial disease and is typically polygenic in origin.Genes play a fundamental part in each biologic process forming complex networks of interactions. However, only some genes have a high number of interactions with other genes in the network and may, therefore, be considered to play an important role. In a preliminary bioinformatic analysis, five genes that showed a higher number of interactions were identified and termed leader genes. In the present study, we use real-time quantitative polymerase chain reaction (PCR) technology to evaluate the expression levels of leader genes in the leukocytes of 10 patientswith refractory chronic periodontitis and compare the expression levels with those of the same genes in 24 healthy patients. Methods: Blood was collected from 24 healthy human subjects and 10 patients with refractory chronic periodontitis and placed into heparinized blood collection tubes by personnel trained in phlebotomy using a sterile technique. Blood leukocyte cells were immediately lysed by using a kit for total RNA purification from human whole blood. Complementary DNA (cDNA) synthesis was obtained from total RNA and then real-time quantitative PCR was performed. PCR efficiencies were calculated with a relative standard curve derived from a five cDNA dilution series in triplicate that gave regression coefficients >0.98 and efficiencies >96%. The standard curves were obtained using glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and growth factor receptor binding protein 2 (GRB2), casitas B-lineage lymphoma (CBL), nuclear factor-KB1 (NFKB1), and REL-A (gene for transcription factor p65) gene primers and amplified with 1.6, 8, 40, 200, and 1,000 ng/mL total cDNA. Curves obtained for each sample showed a linear relationship between RNA concentrations and the cycle threshold value of real-time quantitative PCR for all genes. Data were expressed as mean ± SE (SEM). The groupswere compared to the analysis of variance.Aprobability value

Original languageEnglish
Pages (from-to)1018-1024
Number of pages7
JournalJournal of Periodontology
Volume82
Issue number7
DOIs
Publication statusPublished - Jul 2011

Fingerprint

Chronic Periodontitis
Real-Time Polymerase Chain Reaction
Genes
Complementary DNA
RNA
Transcription Factor RelA
Leukocytes
Glyceraldehyde-3-Phosphate Dehydrogenases
Phlebotomy
Growth Factor Receptors
Gene Regulatory Networks
Periodontitis
Computational Biology
Lymphoma
Blood Cells
Analysis of Variance
Healthy Volunteers
Carrier Proteins
Technology
Polymerase Chain Reaction

Keywords

  • Bioinformatics
  • Genes
  • Periodontitis
  • Polymerase chain reaction

ASJC Scopus subject areas

  • Periodontics

Cite this

Marconcini, S., Covani, U., Barone, A., Vittorio, O., Curcio, M., Barbuti, S., ... Nicolini, C. (2011). Real-Time quantitative polymerase chain reaction analysis of patients with refractory chronic periodontitis. Journal of Periodontology, 82(7), 1018-1024. https://doi.org/10.1902/jop.2010.100312

Real-Time quantitative polymerase chain reaction analysis of patients with refractory chronic periodontitis. / Marconcini, Simone; Covani, Ugo; Barone, Antonio; Vittorio, Orazio; Curcio, Michele; Barbuti, Serena; Scatena, Fabrizio; Felli, Lamberto; Nicolini, Claudio.

In: Journal of Periodontology, Vol. 82, No. 7, 07.2011, p. 1018-1024.

Research output: Contribution to journalArticle

Marconcini, S, Covani, U, Barone, A, Vittorio, O, Curcio, M, Barbuti, S, Scatena, F, Felli, L & Nicolini, C 2011, 'Real-Time quantitative polymerase chain reaction analysis of patients with refractory chronic periodontitis', Journal of Periodontology, vol. 82, no. 7, pp. 1018-1024. https://doi.org/10.1902/jop.2010.100312
Marconcini, Simone ; Covani, Ugo ; Barone, Antonio ; Vittorio, Orazio ; Curcio, Michele ; Barbuti, Serena ; Scatena, Fabrizio ; Felli, Lamberto ; Nicolini, Claudio. / Real-Time quantitative polymerase chain reaction analysis of patients with refractory chronic periodontitis. In: Journal of Periodontology. 2011 ; Vol. 82, No. 7. pp. 1018-1024.
@article{6a5b75279d8b4b2489c6867a29e6c984,
title = "Real-Time quantitative polymerase chain reaction analysis of patients with refractory chronic periodontitis",
abstract = "Background: Periodontitis is a complexmultifactorial disease and is typically polygenic in origin.Genes play a fundamental part in each biologic process forming complex networks of interactions. However, only some genes have a high number of interactions with other genes in the network and may, therefore, be considered to play an important role. In a preliminary bioinformatic analysis, five genes that showed a higher number of interactions were identified and termed leader genes. In the present study, we use real-time quantitative polymerase chain reaction (PCR) technology to evaluate the expression levels of leader genes in the leukocytes of 10 patientswith refractory chronic periodontitis and compare the expression levels with those of the same genes in 24 healthy patients. Methods: Blood was collected from 24 healthy human subjects and 10 patients with refractory chronic periodontitis and placed into heparinized blood collection tubes by personnel trained in phlebotomy using a sterile technique. Blood leukocyte cells were immediately lysed by using a kit for total RNA purification from human whole blood. Complementary DNA (cDNA) synthesis was obtained from total RNA and then real-time quantitative PCR was performed. PCR efficiencies were calculated with a relative standard curve derived from a five cDNA dilution series in triplicate that gave regression coefficients >0.98 and efficiencies >96{\%}. The standard curves were obtained using glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and growth factor receptor binding protein 2 (GRB2), casitas B-lineage lymphoma (CBL), nuclear factor-KB1 (NFKB1), and REL-A (gene for transcription factor p65) gene primers and amplified with 1.6, 8, 40, 200, and 1,000 ng/mL total cDNA. Curves obtained for each sample showed a linear relationship between RNA concentrations and the cycle threshold value of real-time quantitative PCR for all genes. Data were expressed as mean ± SE (SEM). The groupswere compared to the analysis of variance.Aprobability value",
keywords = "Bioinformatics, Genes, Periodontitis, Polymerase chain reaction",
author = "Simone Marconcini and Ugo Covani and Antonio Barone and Orazio Vittorio and Michele Curcio and Serena Barbuti and Fabrizio Scatena and Lamberto Felli and Claudio Nicolini",
year = "2011",
month = "7",
doi = "10.1902/jop.2010.100312",
language = "English",
volume = "82",
pages = "1018--1024",
journal = "Journal of Periodontology",
issn = "0022-3492",
publisher = "American Academy of Periodontology",
number = "7",

}

TY - JOUR

T1 - Real-Time quantitative polymerase chain reaction analysis of patients with refractory chronic periodontitis

AU - Marconcini, Simone

AU - Covani, Ugo

AU - Barone, Antonio

AU - Vittorio, Orazio

AU - Curcio, Michele

AU - Barbuti, Serena

AU - Scatena, Fabrizio

AU - Felli, Lamberto

AU - Nicolini, Claudio

PY - 2011/7

Y1 - 2011/7

N2 - Background: Periodontitis is a complexmultifactorial disease and is typically polygenic in origin.Genes play a fundamental part in each biologic process forming complex networks of interactions. However, only some genes have a high number of interactions with other genes in the network and may, therefore, be considered to play an important role. In a preliminary bioinformatic analysis, five genes that showed a higher number of interactions were identified and termed leader genes. In the present study, we use real-time quantitative polymerase chain reaction (PCR) technology to evaluate the expression levels of leader genes in the leukocytes of 10 patientswith refractory chronic periodontitis and compare the expression levels with those of the same genes in 24 healthy patients. Methods: Blood was collected from 24 healthy human subjects and 10 patients with refractory chronic periodontitis and placed into heparinized blood collection tubes by personnel trained in phlebotomy using a sterile technique. Blood leukocyte cells were immediately lysed by using a kit for total RNA purification from human whole blood. Complementary DNA (cDNA) synthesis was obtained from total RNA and then real-time quantitative PCR was performed. PCR efficiencies were calculated with a relative standard curve derived from a five cDNA dilution series in triplicate that gave regression coefficients >0.98 and efficiencies >96%. The standard curves were obtained using glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and growth factor receptor binding protein 2 (GRB2), casitas B-lineage lymphoma (CBL), nuclear factor-KB1 (NFKB1), and REL-A (gene for transcription factor p65) gene primers and amplified with 1.6, 8, 40, 200, and 1,000 ng/mL total cDNA. Curves obtained for each sample showed a linear relationship between RNA concentrations and the cycle threshold value of real-time quantitative PCR for all genes. Data were expressed as mean ± SE (SEM). The groupswere compared to the analysis of variance.Aprobability value

AB - Background: Periodontitis is a complexmultifactorial disease and is typically polygenic in origin.Genes play a fundamental part in each biologic process forming complex networks of interactions. However, only some genes have a high number of interactions with other genes in the network and may, therefore, be considered to play an important role. In a preliminary bioinformatic analysis, five genes that showed a higher number of interactions were identified and termed leader genes. In the present study, we use real-time quantitative polymerase chain reaction (PCR) technology to evaluate the expression levels of leader genes in the leukocytes of 10 patientswith refractory chronic periodontitis and compare the expression levels with those of the same genes in 24 healthy patients. Methods: Blood was collected from 24 healthy human subjects and 10 patients with refractory chronic periodontitis and placed into heparinized blood collection tubes by personnel trained in phlebotomy using a sterile technique. Blood leukocyte cells were immediately lysed by using a kit for total RNA purification from human whole blood. Complementary DNA (cDNA) synthesis was obtained from total RNA and then real-time quantitative PCR was performed. PCR efficiencies were calculated with a relative standard curve derived from a five cDNA dilution series in triplicate that gave regression coefficients >0.98 and efficiencies >96%. The standard curves were obtained using glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and growth factor receptor binding protein 2 (GRB2), casitas B-lineage lymphoma (CBL), nuclear factor-KB1 (NFKB1), and REL-A (gene for transcription factor p65) gene primers and amplified with 1.6, 8, 40, 200, and 1,000 ng/mL total cDNA. Curves obtained for each sample showed a linear relationship between RNA concentrations and the cycle threshold value of real-time quantitative PCR for all genes. Data were expressed as mean ± SE (SEM). The groupswere compared to the analysis of variance.Aprobability value

KW - Bioinformatics

KW - Genes

KW - Periodontitis

KW - Polymerase chain reaction

UR - http://www.scopus.com/inward/record.url?scp=79960523655&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=79960523655&partnerID=8YFLogxK

U2 - 10.1902/jop.2010.100312

DO - 10.1902/jop.2010.100312

M3 - Article

C2 - 21189087

AN - SCOPUS:79960523655

VL - 82

SP - 1018

EP - 1024

JO - Journal of Periodontology

JF - Journal of Periodontology

SN - 0022-3492

IS - 7

ER -