Spandidos Publications Logo
  • About
    • About Spandidos
    • Aims and Scopes
    • Abstracting and Indexing
    • Editorial Policies
    • Reprints and Permissions
    • Job Opportunities
    • Terms and Conditions
    • Contact
  • Journals
    • All Journals
    • Oncology Letters
      • Oncology Letters
      • Information for Authors
      • Editorial Policies
      • Editorial Board
      • Aims and Scope
      • Abstracting and Indexing
      • Bibliographic Information
      • Archive
    • International Journal of Oncology
      • International Journal of Oncology
      • Information for Authors
      • Editorial Policies
      • Editorial Board
      • Aims and Scope
      • Abstracting and Indexing
      • Bibliographic Information
      • Archive
    • Molecular and Clinical Oncology
      • Molecular and Clinical Oncology
      • Information for Authors
      • Editorial Policies
      • Editorial Board
      • Aims and Scope
      • Abstracting and Indexing
      • Bibliographic Information
      • Archive
    • Experimental and Therapeutic Medicine
      • Experimental and Therapeutic Medicine
      • Information for Authors
      • Editorial Policies
      • Editorial Board
      • Aims and Scope
      • Abstracting and Indexing
      • Bibliographic Information
      • Archive
    • International Journal of Molecular Medicine
      • International Journal of Molecular Medicine
      • Information for Authors
      • Editorial Policies
      • Editorial Board
      • Aims and Scope
      • Abstracting and Indexing
      • Bibliographic Information
      • Archive
    • Biomedical Reports
      • Biomedical Reports
      • Information for Authors
      • Editorial Policies
      • Editorial Board
      • Aims and Scope
      • Abstracting and Indexing
      • Bibliographic Information
      • Archive
    • Oncology Reports
      • Oncology Reports
      • Information for Authors
      • Editorial Policies
      • Editorial Board
      • Aims and Scope
      • Abstracting and Indexing
      • Bibliographic Information
      • Archive
    • Molecular Medicine Reports
      • Molecular Medicine Reports
      • Information for Authors
      • Editorial Policies
      • Editorial Board
      • Aims and Scope
      • Abstracting and Indexing
      • Bibliographic Information
      • Archive
    • World Academy of Sciences Journal
      • World Academy of Sciences Journal
      • Information for Authors
      • Editorial Policies
      • Editorial Board
      • Aims and Scope
      • Abstracting and Indexing
      • Bibliographic Information
      • Archive
    • International Journal of Functional Nutrition
      • International Journal of Functional Nutrition
      • Information for Authors
      • Editorial Policies
      • Editorial Board
      • Aims and Scope
      • Abstracting and Indexing
      • Bibliographic Information
      • Archive
    • International Journal of Epigenetics
      • International Journal of Epigenetics
      • Information for Authors
      • Editorial Policies
      • Editorial Board
      • Aims and Scope
      • Abstracting and Indexing
      • Bibliographic Information
      • Archive
    • Medicine International
      • Medicine International
      • Information for Authors
      • Editorial Policies
      • Editorial Board
      • Aims and Scope
      • Abstracting and Indexing
      • Bibliographic Information
      • Archive
  • Articles
  • Information
    • Information for Authors
    • Information for Reviewers
    • Information for Librarians
    • Information for Advertisers
    • Conferences
  • Language Editing
Spandidos Publications Logo
  • About
    • About Spandidos
    • Aims and Scopes
    • Abstracting and Indexing
    • Editorial Policies
    • Reprints and Permissions
    • Job Opportunities
    • Terms and Conditions
    • Contact
  • Journals
    • All Journals
    • Biomedical Reports
      • Information for Authors
      • Editorial Policies
      • Editorial Board
      • Aims and Scope
      • Abstracting and Indexing
      • Bibliographic Information
      • Archive
    • Experimental and Therapeutic Medicine
      • Information for Authors
      • Editorial Policies
      • Editorial Board
      • Aims and Scope
      • Abstracting and Indexing
      • Bibliographic Information
      • Archive
    • International Journal of Epigenetics
      • Information for Authors
      • Editorial Policies
      • Editorial Board
      • Aims and Scope
      • Abstracting and Indexing
      • Bibliographic Information
      • Archive
    • International Journal of Functional Nutrition
      • Information for Authors
      • Editorial Policies
      • Editorial Board
      • Aims and Scope
      • Abstracting and Indexing
      • Bibliographic Information
      • Archive
    • International Journal of Molecular Medicine
      • Information for Authors
      • Editorial Policies
      • Editorial Board
      • Aims and Scope
      • Abstracting and Indexing
      • Bibliographic Information
      • Archive
    • International Journal of Oncology
      • Information for Authors
      • Editorial Policies
      • Editorial Board
      • Aims and Scope
      • Abstracting and Indexing
      • Bibliographic Information
      • Archive
    • Medicine International
      • Information for Authors
      • Editorial Policies
      • Editorial Board
      • Aims and Scope
      • Abstracting and Indexing
      • Bibliographic Information
      • Archive
    • Molecular and Clinical Oncology
      • Information for Authors
      • Editorial Policies
      • Editorial Board
      • Aims and Scope
      • Abstracting and Indexing
      • Bibliographic Information
      • Archive
    • Molecular Medicine Reports
      • Information for Authors
      • Editorial Policies
      • Editorial Board
      • Aims and Scope
      • Abstracting and Indexing
      • Bibliographic Information
      • Archive
    • Oncology Letters
      • Information for Authors
      • Editorial Policies
      • Editorial Board
      • Aims and Scope
      • Abstracting and Indexing
      • Bibliographic Information
      • Archive
    • Oncology Reports
      • Information for Authors
      • Editorial Policies
      • Editorial Board
      • Aims and Scope
      • Abstracting and Indexing
      • Bibliographic Information
      • Archive
    • World Academy of Sciences Journal
      • Information for Authors
      • Editorial Policies
      • Editorial Board
      • Aims and Scope
      • Abstracting and Indexing
      • Bibliographic Information
      • Archive
  • Articles
  • Information
    • For Authors
    • For Reviewers
    • For Librarians
    • For Advertisers
    • Conferences
  • Language Editing
Login Register Submit
  • This site uses cookies
  • You can change your cookie settings at any time by following the instructions in our Cookie Policy. To find out more, you may read our Privacy Policy.

    I agree
Search articles by DOI, keyword, author or affiliation
Search
Advanced Search
presentation
Molecular Medicine Reports
Join Editorial Board Propose a Special Issue
Print ISSN: 1791-2997 Online ISSN: 1791-3004
Journal Cover
August-2015 Volume 12 Issue 2

Full Size Image

Sign up for eToc alerts
Recommend to Library

Journals

International Journal of Molecular Medicine

International Journal of Molecular Medicine

International Journal of Molecular Medicine is an international journal devoted to molecular mechanisms of human disease.

International Journal of Oncology

International Journal of Oncology

International Journal of Oncology is an international journal devoted to oncology research and cancer treatment.

Molecular Medicine Reports

Molecular Medicine Reports

Covers molecular medicine topics such as pharmacology, pathology, genetics, neuroscience, infectious diseases, molecular cardiology, and molecular surgery.

Oncology Reports

Oncology Reports

Oncology Reports is an international journal devoted to fundamental and applied research in Oncology.

Experimental and Therapeutic Medicine

Experimental and Therapeutic Medicine

Experimental and Therapeutic Medicine is an international journal devoted to laboratory and clinical medicine.

Oncology Letters

Oncology Letters

Oncology Letters is an international journal devoted to Experimental and Clinical Oncology.

Biomedical Reports

Biomedical Reports

Explores a wide range of biological and medical fields, including pharmacology, genetics, microbiology, neuroscience, and molecular cardiology.

Molecular and Clinical Oncology

Molecular and Clinical Oncology

International journal addressing all aspects of oncology research, from tumorigenesis and oncogenes to chemotherapy and metastasis.

World Academy of Sciences Journal

World Academy of Sciences Journal

Multidisciplinary open-access journal spanning biochemistry, genetics, neuroscience, environmental health, and synthetic biology.

International Journal of Functional Nutrition

International Journal of Functional Nutrition

Open-access journal combining biochemistry, pharmacology, immunology, and genetics to advance health through functional nutrition.

International Journal of Epigenetics

International Journal of Epigenetics

Publishes open-access research on using epigenetics to advance understanding and treatment of human disease.

Medicine International

Medicine International

An International Open Access Journal Devoted to General Medicine.

Journal Cover
August-2015 Volume 12 Issue 2

Full Size Image

Sign up for eToc alerts
Recommend to Library

  • Article
  • Citations
    • Cite This Article
    • Download Citation
    • Create Citation Alert
    • Remove Citation Alert
    • Cited By
  • Similar Articles
    • Related Articles (in Spandidos Publications)
    • Similar Articles (Google Scholar)
    • Similar Articles (PubMed)
  • Download PDF
  • Download XML
  • View XML
Article

Selection of suitable reference genes for reverse transcription‑quantitative polymerase chain reaction analysis of neuronal cells differentiated from bone mesenchymal stem cells

  • Authors:
    • Yu‑Xi He
    • Yan Zhang
    • Qiwei Yang
    • Chenguang Wang
    • Guanfang Su
  • View Affiliations / Copyright

    Affiliations: Department of Ophthalmology, Second Hospital, Jilin University, Changchun, Jilin 130000, P.R. China
  • Pages: 2291-2300
    |
    Published online on: April 23, 2015
       https://doi.org/10.3892/mmr.2015.3671
  • Expand metrics +
Metrics: Total Views: 0 (Spandidos Publications: | PMC Statistics: )
Metrics: Total PDF Downloads: 0 (Spandidos Publications: | PMC Statistics: )
Cited By (CrossRef): 0 citations Loading Articles...

This article is mentioned in:



Abstract

Reverse transcription‑quantitative polymerase chain reaction (RT‑qPCR) is a technique widely used for the quantification of mRNA transcription, It is well recognized that the reference genes used in RT‑qPCR require appropriate validation to ensure that gene expression is unaffected by experimental conditions. The differentiation of bone mesenchymal stem cells (BMSCs) into neurons is important in the treatment of nerve injury. In gene expression analysis of the differentiation of BMSCs into neuronal cells by, the commonly used reference genes for RNA analysis are often selected without any preliminary evaluation of their suitability. The present study aimed to evaluate the mRNA expression levels of 11 putative reference genes, including ACTB, ARBP, B2M, CYCA, GAPDH, GUSB, HPRT, PPIA, RPL13A, TBP and PGK1, in order to select the most suitable reference genes for RT‑qPCR of the differentiation of neuronal cells by BMSCs. The mRNA expression levels of the 11 putative reference genes were examined using RT‑qPCR in rat BMSCs differentiated into neuronal cells. Normal BMSCs and three types of rat BMSCs, which were chemically induced to differentiate into neurons using neurotrophic cytokines and co‑culture with retinal cells. The geNorm, NormFinder and BestKeeper software programs were used to select the most suitable reference genes. The results of the analyses using the three software programs demonstrated that RPL13A was the most stable among all the groups, while ACTB was the least stable. The combination of CYCA and PPIA reference genes contributed the most to increasing stability. The suitability of selected reference genes requires previous pre‑selection in every investigation. Based on the three software programs, RPL13A, and the combination of CYCA and PPIA were identified as the most suitable reference genes for RT‑qPCR in neuronal cells differentiated from BMSCs.
View Figures

Figure 1

Figure 2

Figure 3

Figure 4

Figure 5

Figure 6

Figure 7

Figure 8

View References

1 

Bustin SA, Benes V, Garson JA, et al: The MIQE guidelines: minimum information for publication of quantitative real-time PCR experiments. Clin Chem. 55:611–622. 2009. View Article : Google Scholar : PubMed/NCBI

2 

Huggett J, Dheda K, Bustin S and Zumla A: Real-time RT-PCR normalisation; strategies and considerations. Genes Immun. 6:279–284. 2005. View Article : Google Scholar : PubMed/NCBI

3 

Bustin SA and Nolan T: Pitfalls of quantitative real-time reverse-transcription polymerase chain reaction. J Biomol Tech. 15:155–166. 2004.PubMed/NCBI

4 

Zhong Q, Zhang Q, Wang Z, et al: Expression profiling and validation of potential reference genes during Paralichthys olivaceus embryogenesis. Mar Biotechnol (NY). 10:310–318. 2008. View Article : Google Scholar

5 

Bustin SA, Benes V, Nolan T and Pfaffl MW: Quantitative real-time RT-PCR-a perspective. J Mol Endocrinol. 34:597–601. 2005. View Article : Google Scholar : PubMed/NCBI

6 

Dheda K, Huggett JF, Bustin SA, Johnson MA, Rook G and Zumla A: Validation of housekeeping genes for normalizing RNA expression in real-time PCR. Biotechniques. 37:112–114. 116118–119. 2004.PubMed/NCBI

7 

Radonic A, Thulke S, Mackay IM, Landt O, Siegert W and Nitsche A: Guideline to reference gene selection for quantitative real-time PCR. Biochem Biophys Res Commun. 313:856–862. 2004. View Article : Google Scholar : PubMed/NCBI

8 

Suzuki T, Kobayashi K and Sasaki O: Real-time displacement measurement with a two-wavelength sinusoidal phase-modulating laser diode interferometer. Appl Opt. 39:2646–2652. 2000. View Article : Google Scholar

9 

Cordoba EM, Die JV, González-Verdejo CI, Nadal S and Román B: Selection of reference genes in Hedysarum coronarium under various stresses and stages of development. Anal Biochem. 409:236–243. 2011. View Article : Google Scholar

10 

Schmittgen TD and Zakrajsek BA: Effect of experimental treatment on housekeeping gene expression: validation by real-time, quantitative RT-PCR. J Biochem Biophys Methods. 46:69–81. 2000. View Article : Google Scholar : PubMed/NCBI

11 

Santos AR and Duarte CB: Validation of internal control genes for expression studies: effects of the neurotrophin BDNF on hippocampal neurons. J Neurosci Res. 86:3684–3692. 2008. View Article : Google Scholar : PubMed/NCBI

12 

Sun C, Shao J, Su L, et al: Cholinergic neuron-like cells derived from bone marrow stromal cells induced by tricyclodecane-9-yl-xanthogenate promote functional recovery and neural protection after spinal cord injury. Cell Transplant. 22:961–975. 2013. View Article : Google Scholar

13 

Woodbury D, Schwarz EJ, Prockop DJ and Black IB: Adult rat and human bone marrow stromal cells differentiate into neurons. J Neurosci Res. 61:364–370. 2000. View Article : Google Scholar : PubMed/NCBI

14 

Sanchez-Ramos J, Song S, Cardozo-Pelaez F, et al: Adult bone marrow stromal cells differentiate into neural cells in vitro. Exp Neurol. 164:247–256. 2000. View Article : Google Scholar : PubMed/NCBI

15 

Xu C, Su G and Mu D: Retinal cells of rat neonates inducing the bone marrow mesenchymal cells into RGC-like cells. Chin Ophthal Res. 26:174–178. 2008.

16 

Wang C, Su G, Qi S, et al: Study of GAP-43 expression on differentiation of bone mesenchymal stem cells to neuron-like cells. Chin J Lab Diagn. 12:1206–1209. 2008.

17 

Vandesompele J, De Preter K, Pattyn F, et al: Accurate normalization of real-time quantitative RT-PCR data by geometric averaging of multiple internal control genes. Genome Biol. 3:RESEARCH00342002. View Article : Google Scholar : PubMed/NCBI

18 

Andersen CL, Jensen JL and Ørntoft TF: Normalization of real-time quantitative reverse transcription-PCR data: a model-based variance estimation approach to identify genes suited for normalization, applied to bladder and colon cancer data sets. Cancer Res. 64:5245–5250. 2004. View Article : Google Scholar : PubMed/NCBI

19 

Pfaffl MW, Tichopad A, Prgomet C and Neuvians TP: Determination of stable housekeeping genes, differentially regulated target genes and sample integrity: BestKeeper-excel-based tool using pair-wise correlations. Biotechnol Lett. 26:509–515. 2004. View Article : Google Scholar : PubMed/NCBI

20 

Long X, Olszewski M, Huang W and Kletzel M: Neural cell differentiation in vitro from adult human bone marrow mesenchymal stem cells. Stem Cells Dev. 14:65–69. 2005. View Article : Google Scholar : PubMed/NCBI

21 

Feng X, Xiong Y, Qian H, Lei M, Xu D and Ren Z: Selection of reference genes for gene expression studies in porcine skeletal muscle using SYBR green qPCR. J Biotechnol. 150:288–293. 2010. View Article : Google Scholar : PubMed/NCBI

22 

Aramant RB, Seiler MJ and Ball SL: Successful cotransplantation of intact sheets of fetal retina with retinal pigment epithelium. Invest Ophthalmol Vis Sci. 40:1557–1564. 1999.PubMed/NCBI

23 

Gouras P, Flood MT, Kjedbye H, Bilek MK and Eggers H: Transplantation of cultured human retinal epithelium to Bruch’s membrane of the owl monkey’s eye. Curr Eye Res. 4:253–265. 1985. View Article : Google Scholar : PubMed/NCBI

24 

Gimble JM, Zvonic S, Floyd ZE, Kassem M and Nuttall ME: Playing with bone and fat. J Cell Biochem. 98:251–266. 2006. View Article : Google Scholar : PubMed/NCBI

25 

Dezawa M, Kanno H, Hoshino M, et al: Specific induction of neuronal cells from bone marrow stromal cells and application for autologous transplantation. J Clin Invest. 113:1701–1710. 2004. View Article : Google Scholar : PubMed/NCBI

26 

Gregory CA, Prockop DJ and Spees JL: Non-hematopoietic bone marrow stem cells: molecular control of expansion and differentiation. Exp Cell Res. 306:330–335. 2005. View Article : Google Scholar : PubMed/NCBI

27 

Gu S, Xing C, Han J, Tso MO and Hong J: Differentiation of rabbit bone marrow mesenchymal stemcells into corneal epithelial cells in vivo and ex vivo. Mol Vis. 15:99–107. 2009.

28 

Liu C, Xin N, Zhai Y, et al: Reference gene selection for quantitative real-time RT-PCR normalization in the half-smooth tongue sole (cynoglossus semilaevis) at different developmental stages, in various tissue types and on exposure to chemicals. PLoS One. 9:e917152014. View Article : Google Scholar : PubMed/NCBI

29 

McCurley AT and Callard GV: Characterization of housekeeping genes in zebrafish: male-female differences and effects of tissue type, developmental stage and chemical treatment. BMC Mol Biol. 9:1022008. View Article : Google Scholar : PubMed/NCBI

30 

Farrokhi A, Eslaminejad MB, Nazarian H, Moradmand A, Samadian A and Akhlaghi A: Appropriate reference gene selection for real-time Pcr Data normalization during rat mesenchymal stem cell differentiation. Cell Mol Biol (Noisy-le-grand). 58(Suppl): OL1660–1670. 2012.

31 

Swijsen A, Nelissen K, Janssen D, Rigo JM and Hoogland G: Validation of reference genes for quantitative real-time PCR studies in the dentate gyrus after experimental febrile seizures. BMC Res Notes. 5:6852012. View Article : Google Scholar : PubMed/NCBI

32 

Chang E, Shi S, Liu J, et al: Selection of reference genes for quantitative gene expression studies in Platycladus orientalis (Cupressaceae) using real-time PCR. PLoS One. 7:e332782012. View Article : Google Scholar : PubMed/NCBI

33 

Bonefeld BE, Elfving B and Wegener G: Reference genes for normalization: a study of rat brain tissue. Synapse. 62:302–309. 2008. View Article : Google Scholar : PubMed/NCBI

Related Articles

  • Abstract
  • View
  • Download
  • Twitter
Copy and paste a formatted citation
Spandidos Publications style
He YX, Zhang Y, Yang Q, Wang C and Su G: Selection of suitable reference genes for reverse transcription‑quantitative polymerase chain reaction analysis of neuronal cells differentiated from bone mesenchymal stem cells. Mol Med Rep 12: 2291-2300, 2015.
APA
He, Y., Zhang, Y., Yang, Q., Wang, C., & Su, G. (2015). Selection of suitable reference genes for reverse transcription‑quantitative polymerase chain reaction analysis of neuronal cells differentiated from bone mesenchymal stem cells. Molecular Medicine Reports, 12, 2291-2300. https://doi.org/10.3892/mmr.2015.3671
MLA
He, Y., Zhang, Y., Yang, Q., Wang, C., Su, G."Selection of suitable reference genes for reverse transcription‑quantitative polymerase chain reaction analysis of neuronal cells differentiated from bone mesenchymal stem cells". Molecular Medicine Reports 12.2 (2015): 2291-2300.
Chicago
He, Y., Zhang, Y., Yang, Q., Wang, C., Su, G."Selection of suitable reference genes for reverse transcription‑quantitative polymerase chain reaction analysis of neuronal cells differentiated from bone mesenchymal stem cells". Molecular Medicine Reports 12, no. 2 (2015): 2291-2300. https://doi.org/10.3892/mmr.2015.3671
Copy and paste a formatted citation
x
Spandidos Publications style
He YX, Zhang Y, Yang Q, Wang C and Su G: Selection of suitable reference genes for reverse transcription‑quantitative polymerase chain reaction analysis of neuronal cells differentiated from bone mesenchymal stem cells. Mol Med Rep 12: 2291-2300, 2015.
APA
He, Y., Zhang, Y., Yang, Q., Wang, C., & Su, G. (2015). Selection of suitable reference genes for reverse transcription‑quantitative polymerase chain reaction analysis of neuronal cells differentiated from bone mesenchymal stem cells. Molecular Medicine Reports, 12, 2291-2300. https://doi.org/10.3892/mmr.2015.3671
MLA
He, Y., Zhang, Y., Yang, Q., Wang, C., Su, G."Selection of suitable reference genes for reverse transcription‑quantitative polymerase chain reaction analysis of neuronal cells differentiated from bone mesenchymal stem cells". Molecular Medicine Reports 12.2 (2015): 2291-2300.
Chicago
He, Y., Zhang, Y., Yang, Q., Wang, C., Su, G."Selection of suitable reference genes for reverse transcription‑quantitative polymerase chain reaction analysis of neuronal cells differentiated from bone mesenchymal stem cells". Molecular Medicine Reports 12, no. 2 (2015): 2291-2300. https://doi.org/10.3892/mmr.2015.3671
Follow us
  • Twitter
  • LinkedIn
  • Facebook
About
  • Spandidos Publications
  • Careers
  • Cookie Policy
  • Privacy Policy
How can we help?
  • Help
  • Live Chat
  • Contact
  • Email to our Support Team